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Nicoletta BRUNELLO

Professore Ordinario presso: Dipartimento di Scienze della Vita sede ex-Scienze Biomediche


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Pubblicazioni

2021 - Carnosine Protects Macrophages against the Toxicity of Aβ1-42 Oligomers by Decreasing Oxidative Stress [Articolo su rivista]
Caruso, Giuseppe; Benatti, Cristina; Musso, Nicolò; Fresta, Claudia G.; Fidilio, Annamaria; Spampinato, Giorgia; Brunello, Nicoletta; Bucolo, Claudio; Drago, Filippo; Lunte, Susan M.; Peterson, Blake R.; Tascedda, Fabio; Caraci, Filippo
abstract


2021 - Serum metabolic signature of binge-like palatable food consumption in female rats by nuclear magnetic resonance spectroscopy [Articolo su rivista]
Cifani, Carlo; Alboni, Silvia; Mucci, Adele; Benatti, Cristina; Botticelli, Luca; Brunello, Nicoletta; Micioni Di Bonaventura, Maria Vittoria; Righi, Valeria
abstract

Maladaptive eating behavior is a growing public health problem and compulsively eating excessive food in a short time, or binge eating, is a key symptom of many eating disorders. In order to investigate the binge-like eating behavior in female rats, induced by intermittent food restrictions/refeeding and frustration stress, we analyzed for the first time the metabolic profile obtained from serum of rats, through nuclear magnetic resonance (NMR) spectroscopy. In this experimental protocol, rats were exposed to chow food restricting/refeeding and frustration stress manipulation. This stress procedure consists of 15 min exposure to the odor and sight of a familiar chocolate paste, without access to it, just before offering the palatable food. In this model, a "binge-eating episode" was considered the significantly higher palatable food consumption within 2 h in restricted and stressed rats (R + S) than in the other three experimental groups: rats with no food restriction and no stress (NR + NS), only stressed rats (NR + S) or only restricted rats (R + NS). Serum samples from these four different rat groups were collected. The statistical analysis of the 1 H NMR spectral profiles of the four sets of samples pointed to O- and N-acetyl glycoproteins as the main biomarkers for the discrimination of restriction effects. Other metabolites, such as threonine, glycine, glutamine, acetate, pyruvate and lactate, showed trends that may be useful to understand metabolic pathways involved in eating disorders. This study suggested that NMR-based metabolomics is a suitable approach to detect biomarkers related to binge-eating behavior.


2021 - Vortioxetine Prevents Lipopolysaccharide-Induced Memory Impairment Without Inhibiting the Initial Inflammatory Cascade [Articolo su rivista]
Alboni, S.; Benatti, C.; Colliva, C.; Radighieri, G.; Blom, J. M. C.; Brunello, N.; Tascedda, F.
abstract

Vortioxetine is a novel multimodal antidepressant that modulates a wide range of neurotransmitters throughout the brain. Preclinical and clinical studies have shown that vortioxetine exerts positive effects on different cognitive domains and neuroprotective effects. Considering the key role of microglial cells in brain plasticity and cognition, we aimed at investigating the effects of pretreatment with vortioxetine in modulating behavioral and molecular effects induced by an immune challenge: peripheral injection of lipopolysaccharide (LPS). To this purpose, C57BL/6J male mice were first exposed to a 28-day standard diet or vortioxetine-enriched diet, which was followed by an acute immune challenge with LPS. Sickness symptoms and depressive-like behaviors (anhedonia and memory impairment) were tested 6 and 24 h after exposure to LPS, respectively. Moreover, the expressions of markers of immune activation and M1/M2 markers of microglia polarization were measured in the dorsal and ventral parts of the hippocampus. The pretreatment with vortioxetine did not affect both LPS-induced sickness behavior and anhedonia but prevented the deficit in the recognition memory induced by the immune challenge. At the transcriptional level, chronic exposure to vortioxetine did not prevent LPS-induced upregulation of proinflammatory cytokines 6 h after the immune challenge but rather seemed to potentiate the immune response to the challenge also by affecting the levels of expression of markers of microglia M1 phenotype, like cluster of differentiation (CD)14 and CD86, in an area-dependent manner. However, at the same time point, LPS injection significantly increased the expression of the M2 polarization inducer, interleukin 4, only in the hippocampus of animals chronically exposed to vortioxetine. These results demonstrate that a chronic administration of vortioxetine specifically prevents LPS-induced memory impairment, without affecting acute sickness behavior and anhedonia, and suggest that hippocampal microglia may represent a cellular target of this novel antidepressant medication. Moreover, we provide a useful model to further explore the molecular mechanisms specifically underlying cognitive impairments following an immune challenge.


2020 - Lymnaea stagnalis as model for translational neuroscience research: from pond to bench [Articolo su rivista]
Rivi, Veronica; Benatti, C; Colliva, C; Radighieri, G; Brunello, N; Tascedda, F; Blom, Johanna
abstract

The purpose of this review is to illustrate how a reductionistic, but sophisticated, approach based on the use of a simple model system such as the pond snail Lymnaea stagnalis (L. stagnalis), might be useful to address fundamental questions in learning and memory. L. stagnalis, as a model, provides an interesting platform to investigate the dialog between the synapse and the nucleus and vice versa during memory and learning. More importantly, the "molecular actors" of the memory dialogue are well-conserved both across phylogenetic groups and learning paradigms, involving single- or multi-trials, aversion or reward, operant or classical conditioning. At the same time, this model could help to study how, where and when the memory dialog is impaired in stressful conditions and during aging and neurodegeneration in humans and thus offers new insights and targets in order to develop innovative therapies and technology for the treatment of a range of neurological and neurodegenerative disorders.


2020 - P.304 Executive functioning and genetic variation in pediatric patients with cancer [Abstract in Rivista]
Colliva, C.; Dalla Porta, M. F.; Ferrari, M.; Benatti, C.; Cellini, M.; Brunello, N.; Blom, J. M.
abstract


2020 - Should Patients Receiving ACE Inhibitors or Angiotensin Receptor Blockers be Switched to Other Antihypertensive Drugs to Prevent or Improve Prognosis of Novel Coronavirus Disease 2019 (COVID-19)? [Articolo su rivista]
Trifiro, G.; Crisafulli, S.; Ando, G.; Racagni, G.; Drago, F.; Berrino, L.; Re, M.; Bernardini, R.; Chiamulera, C.; D'Avolio, A.; Pani, L.; Clementi, E.; Capuano, A.; Scaglione, F.; Danesi, R.; Cirino, G.; Mugelli, A.; Bonanno, G.; Brunello, N.; Luca, A.; Hrelia, P.; Pistis, M.; Ghelardini, C.; Taglialatela, M.
abstract


2019 - Combined fluoxetine and metformin treatment potentiates antidepressant efficacy increasing IGF2 expression in the dorsal hippocampus [Articolo su rivista]
Poggini, S.; Golia, M. T.; Alboni, S.; Milior, G.; Sciarria, L. P.; Viglione, A.; Bon, G. M.; Brunello, N.; Puglisi-Allegra, S.; Limatola, C.; Maggi, L.; Branchi, I.
abstract

An increasing number of studies show that selective serotonin reuptake inhibitors (SSRIs) exert their therapeutic action, at least in part, by amplifying the influence of the living environment on mood. As a consequence, when administered in a favorable environment, SSRIs lead to a reduction of symptoms, but in stressful conditions, they show limited efficacy. Therefore, novel therapeutic approaches able to neutralize the influence of the stressful environment on treatment are needed. The aim of our study was to test whether, in a mouse model of depression, the combined administration of SSRI fluoxetine and metformin, a drug able to improve the metabolic profile, counteracts the limited efficacy of fluoxetine alone when administered in stressful conditions. Indeed, metabolic alterations are associated to both the onset of major depression and the antidepressant efficacy. To this goal, adult C57BL/6 male mice were exposed to stress for 6 weeks; the first two weeks was aimed at generating a mouse model of depression. During the remaining 4 weeks, mice received one of the following treatments: vehicle, fluoxetine, metformin, or a combination of fluoxetine and metformin. We measured liking- and wanting-type anhedonia as behavioral phenotypes of depression and assessed the expression levels of selected genes involved in major depressive disorder and antidepressant response in the dorsal and ventral hippocampus, which are differently involved in the depressive symptomatology. The combined treatment was more effective than fluoxetine alone in ameliorating the depressive phenotype after one week of treatment. This was associated to an increase in IGF2 mRNA expression and enhanced long-term potentiation, specifically in the dorsal hippocampus, at the end of treatment. Overall, the present results show that, when administered in stressful conditions, the combined fluoxetine and metformin treatment may represent a more effective approach than fluoxetine alone in a short term. Finally, our findings highlight the relevance of polypharmacological strategy as effective interventions to increase the efficacy of the antidepressant drugs currently available.


2019 - Interplay between inflammation and neural plasticity: Both immune activation and suppression impair LTP and BDNF expression [Articolo su rivista]
Golia, M. T.; Poggini, S.; Alboni, S.; Garofalo, S.; Ciano Albanese, N.; Viglione, A.; Ajmone-Cat, M. A.; St-Pierre, A.; Brunello, N.; Limatola, C.; Branchi, I.; Maggi, L.
abstract

An increasing number of studies show that both inflammation and neural plasticity act as key players in the vulnerability and recovery from psychiatric disorders and neurodegenerative diseases. However, the interplay between these two players has been limitedly explored. In fact, while a few studies reported an immune activation, others conveyed an immune suppression, associated with an impairment in neural plasticity. Therefore, we hypothesized that deviations in inflammatory levels in both directions may impair neural plasticity. We tested this hypothesis experimentally, by acute treatment of C57BL/6 adult male mice with different doses of two inflammatory modulators: lipopolysaccharide (LPS), an endotoxin, and ibuprofen (IBU), a nonselective cyclooxygenase inhibitor, which are respectively a pro- and an anti-inflammatory agent. The results showed that LPS and IBU have different effects on behavior and inflammatory response. LPS treatment induced a reduction of body temperature, a decrease of body weight and a reduced food and liquid intake. In addition, it led to increased levels of inflammatory markers expression, both in the total hippocampus and in isolated microglia cells, including Interleukin (IL)-1β, and enhanced the concentration of prostaglandin E2 (PGE2). On the other hand, IBU increased the level of anti-inflammatory markers, decreased tryptophan 2,3-dioxygenase (TDO2), the first step in the kynurenine pathway known to be activated during inflammatory conditions, and PGE2 levels. Though LPS and IBU administration differently affected mediators related with pro- or anti-inflammatory responses, they produced overlapping effects on neural plasticity. Indeed, higher doses of both LPS and IBU induced a statistically significant decrease in the amplitude of long-term potentiation (LTP), in Brain-Derived Neurotrophic Factor (BDNF) expression levels and in the phosphorylation of the AMPA (α-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid) receptor subunit GluR1, compared to the control group. Such effect appears to be dose-dependent since only the higher, but not the lower, dose of both compounds led to a plasticity impairment. Overall, the present findings indicate that acute treatment with pro- and anti-inflammatory agents impair neural plasticity in a dose dependent manner.


2019 - Modulation of neuroplasticity-related targets following stress-induced acute escape deficit [Articolo su rivista]
Benatti, C.; Radighieri, Giulia; Alboni, S.; Blom, J. M. C.; Brunello, N.; Tascedda, F.
abstract

Understanding resilience is a major challenge to improve current pharmacological therapies aimed at complementing psychological-based approaches of stress-related disorders. In particular, resilience is a multi-factorial construct where the complex network of molecular events that drive the process still needs to be resolved. Here, we exploit the acute escape deficit model, an animal model based on exposure to acute unavoidable stress followed by an escape test, to define vulnerable and resilient phenotypes in rats. Hippocampus and prefrontal cortex (PFC), two of the brain areas most involved in the stress response, were analysed for gene expression at two different time points (3 and 24 h) after the escape test. Total Brain-Derived Neurotrophic Factor (BDNF) was highly responsive in the PFC at 24-h after the escape test, while expression of BDNF transcript IV increased in the hippocampus of resistant animals 3 h post-test. Expression of memory enhancers like Neuronal PAS Domain Protein 4 (Npas4) and Activity-regulated cytoskeleton-associated protein (Arc) decreased in a time- and region-dependent fashion in both behavioural phenotypes. Also, the memory inhibitor Protein Phosphatase 1 (Ppp1ca) was increased in the hippocampus of resilient rats at 3 h post-test. Given the importance of neurotrophic factors and synaptic plasticity-related genes for the development of appropriate coping strategies, our data contribute to an additional step forward in the comprehension of the psychobiology of stress and resiliency.


2019 - P.1.04 Expression of histone variants H3.3 and H2a.z in the rat brain: Physiopathological and pharmacological implications [Abstract in Rivista]
Radighieri, G.; Benatti, C.; Zoli, M.; Blom, J. M. C.; Brunello, N.; Tascedda, F.
abstract

In the overall context of epigenetic modifications in charge of managing genome plasticity and dynamics [1, 2], the role of nucleosomal loading of histone variants is becoming increasingly captivating. Two replication-independent isoforms of histones H3 and H2A, namely H3.3 and H2A.Z, have caught attention because of their involvement in neuronal plasticity processes, cognitive functions and behavioral outcomes. In fact, their incorporation/eviction in nucleosomes and their turnover in neurons influence chromatin accessibility and therefore transcription. H3.3 enrichment at gene bodies and promoters of genes involved in synaptic plasticity was proved to be positively correlated with their expression, while learning-induced H2A.Z eviction in specific genes promotes gene transcription, intervening in memory consolidation processes. H3.3 is encoded by H3f3a and H3f3b independent genes, generating identical proteins, namely H3.3A and H3.3B. Notably, H3f3b gene, but not H3f3a, was proved responsive to neuronal activating stimuli as well as environmental triggers and stressful procedures [3]. H2A.Z hypervariants H2A.Z.1 and H2A.Z.2, encoded respectively by H2afz and H2afv genes, regulate both basal and stimulus-induced neuronal gene expression of independent gene sets [4]. Starting from this evidence, the purpose of this study was to characterize basal expression levels of all genes encoding for the histones variants above mentioned in rodent hippocampus and prefrontal cortex (PFC), two brain regions closely related to brain plasticity, cognition and behavior. Adult male rats (n=7) were sacrificed, their brains removed and dissected. Total RNA extraction was performed, followed by total RNA reverse transcription and Real Time PCR, where specific forward and reverse primer were used for each gene encoding for H3.3 (H3f3a and H3f3b), H2A.Z (H2afz and H2afv) and endogenous control GAPDH. Statistical analysis was performed by means of one-way ANOVA; p<0.05 was considered as a threshold for statistically significant difference. Molecular analyses revealed that, for both hippocampus and PFC, H3f3a mRNA was more expressed at the steady-state compared to H3f3b (p<0.001), as happens for H2afz mRNA, which displays higher levels than H2afv (p<0.001). Moreover, comparing hippocampal and PFC mRNA levels for each variant, H3f3a and H3f3b expression was increased in the hippocampus with respect to the prefrontal cortex (p<0.001), and a comparable outcome was showed for H2afv (p<0.001) but not for H2afz (p>0.05). Our results suggest that 1) differential basal expression levels of genes encoding for H3.3 and H2A.Z may underlie unique gene responsiveness following different stimuli, as previously hypothesized by others [3,4], and this may be crucial in highly-responsive, pathological- and environment-related tissues like hippocampus and PFC; 2) striking lower steady-state expression of H3f3b and H2afv genes might imply a major sensitivity to neuronal inputs compared to their correspondent counterparts; 3) higher expression levels in the hippocampus with respect to the PFC might underpin brain-region specific expression and function for histone variants and their isoforms. Together, these data clear the way for further studies meant at investigating stimulus-dependent regulation of H3.3 and H2A.Z gene isoforms expression and their putative involvement in the physiopathology of brain and its diseases [5]. References [1] Rigillo, G., Vilella, A., Benatti, C., Schaeffer, L., Brunello, N., Blom, J.M.C., Zoli, M., Tascedda, F., 2018. LPS-induced histone H3 phospho(Ser10)-acetylation(Lys14) regulates neuronal and microglial neuroinflammatory response. Brain Behav Immun. https://doi.org/10.1016/j.bbi.2018.09.019. [2] Ottaviani, E., Accorsi, A., Rigillo, G., Malagoli, D., Blom, J.M., Tascedda, F., 2013. Epigenetic modification in neurons of the mollusc Pomacea canaliculata after immune challe


2018 - LPS-induced histone H3 phospho(Ser10)-acetylation(Lys14) regulates neuronal and microglial neuroinflammatory response [Articolo su rivista]
Rigillo, Giovanna; Vilella, Antonietta; Benatti, Cristina; Schaeffer, Laurent; Brunello, Nicoletta; Blom, Johanna M. C.; Zoli, Michele; Tascedda, Fabio
abstract

Epigenetic modifications of DNA and histone proteins are emerging as fundamental mechanisms by which neural cells adapt their transcriptional response to environmental cues, such as, immune stimuli or stress. In particular, histone H3 phospho(Ser10)-acetylation(Lys14) (H3S10phK14ac) has been linked to activation of specific gene expression. The purpose of this study was to investigate the role of H3S10phK14ac in a neuroinflammatory condition. Adult male rats received a intraperitoneal injection of lipopolysaccharide (LPS) (830 μg/Kg/i.p., n = 6) or vehicle (saline 1 mL/kg/i.p., n = 6) and were sacrificed 2 or 6 h later. We showed marked region- and time-specific increases in H3S10phK14ac in the hypothalamus and hippocampus, two principal target regions of LPS. These changes were accompanied by a marked transcriptional activation of interleukin (IL) 1β, IL-6, Tumour Necrosis Factor (TNF) α, the inducible nitric oxide synthase (iNOS) and the immediate early gene c-Fos. By means of chromatin immunoprecipitation, we demonstrated an increased region- and time-specific association of H3S10phK14ac with the promoters of IL-6, c-Fos and iNOS genes, suggesting that part of the LPS-induced transcriptional activation of these genes is regulated by H3S10phK14ac. Finally, by means of multiple immunofluorescence approach, we showed that increased H3S10phK14ac is cell type-specific, being neurons and reactive microglia, the principal histological types involved in this response. Present data point to H3S10phK14ac as a principal epigenetic regulator of neural cell response to systemic LPS and underline the importance of distinct time-, region- and cell-specific epigenetic mechanisms that regulate gene transcription to understand the mechanistic complexity of neuroinflammatory response to immune challenges.


2018 - Molecular changes associated with escitalopram response in a stress-based model of depression [Articolo su rivista]
Benatti, Cristina; Alboni, Silvia; Blom, Johanna Maria Catharina; Mendlewicz, Julien; Tascedda, Fabio; Brunello, Nicoletta
abstract

Converging evidence points at hypothalamus-pituitary-adrenal (HPA) axis hyperactivity and neuroinflammation as important factors involved in the etiopathogenesis of major depressive disorder (MDD) and in therapeutic efficacy of antidepressants. In this study, we examined the molecular effects associated with a response to a week-long treatment with escitalopram in the chronic escape deficit (CED) model, a validated model of depression based on the induction of an escape deficit after exposure of rats to an unavoidable stress. We confirmed our previous result that a treatment with escitalopram (10 mg/kg) was effective after 7 days in reverting the stress-induced escape deficit in approximately 50% of the animals, separating responders from non-responders. Expression of markers of HPA axis functionality as well as several inflammatory mediators were evaluated in the hypothalamus, a key structure integrating signals from the neuro, immune, endocrine systems. In the hypothalamus of responder animals we observed a decrease in the expression of CRH and its receptors and an increase in GR protein in total and nuclear extracts; this effect was accompanied by a significant decrease in circulating corticosterone in the same cohort. Hypothalamic IL-1β and TNFα expression were increased in stressed animals, while CXCL2, IL-6, and ADAM17 mRNA levels were decreased in escitalopram treated rats regardless of the treatment response. These data suggest that efficacy of a one week treatment with escitalopram may be partially mediated by a decrease HPA axis activity, while in the hypothalamus the drug-induced effects on the expression of immune modulators did not correlate with the behavioural outcome.


2018 - Neither all anti-inflammatory drugs nor all doses are effective in accelerating the antidepressant-like effect of fluoxetine in an animal model of depression [Articolo su rivista]
Alboni, Silvia; Benatti, Cristina; Capone, Giacomo; Tascedda, Fabio; Brunello, Nicoletta
abstract

Introduction: Non-steroidal anti-inflammatory drugs (NSAIDs) have been studied as possible adjunctive therapy in the treatment of depression. However, administering NSAIDs to increase the effectiveness of antidepressant has yielded inconsistent results. Methods: We evaluated the effect of the co-administration of fluoxetine (5 mg/kg) and flurbiprofen (5 mg/kg) or fluoxetine (5 mg/kg) and celecoxib (5 mg/kg) in the chronic escape deficit (CED) model of depression after 7 days of treatment. The co-administration of fluoxetine plus acetylsalicylic acid (ASA, 45 mg/kg i.p.) was used as a positive control. Moreover, we tested the behavioral effect of different doses (45, 22.5, and 11.25 mg/Kg i.p.) of ASA as potentiating agent of the effect of fluoxetine in the same paradigm. Results: Our study showed that only the co-administration of ASA with fluoxetine was able to revert the stress-induced condition of escape deficit after 7 days of treatment, and that the amplitude of the antidepressant-like effect of ASA was dose dependent. In the same experimental conditions, celecoxib with fluoxetine only partially resolved the stress-induced impaired behavior while flurbiprofen/fluoxetine cotreatment was ineffective. Limitations: Our study is still exploratory, more doses, longer treatment regimens, and different behavioral outcomes must be investigated to draw a clear conclusion. Conclusion: Our results further stress the importance of the type and dose when NSAIDs are associated with antidepressants to ameliorate a clinical response.


2017 - Fluoxetine effects on molecular, cellular and behavioral endophenotypes of depression are driven by the living environment [Articolo su rivista]
Alboni, Silvia; van Dijk, R. M.; Poggini, Silvia; Milior, G.; Perrotta, M.; Drenth, T.; Brunello, Nicoletta; Wolfer, D. P.; Limatola, C.; Amrein, I.; Cirulli, F.; Maggi, L.; Branchi, I.
abstract

Selective serotonin reuptake inhibitors (SSRIs) represent the most common treatment for major depression. However, their efficacy is variable and incomplete. In order to elucidate the cause of such incomplete efficacy, we explored the hypothesis positing that SSRIs may not affect mood per se but, by enhancing neural plasticity, render the individual more susceptible to the influence of the environment. Consequently, SSRI administration in a favorable environment promotes a reduction of symptoms, whereas in a stressful environment leads to a worse prognosis. To test such hypothesis, we exposed C57BL/6 mice to chronic stress in order to induce a depression-like phenotype and, subsequently, to fluoxetine treatment (21 days), while being exposed to either an enriched or a stressful condition. We measured the most commonly investigated molecular, cellular and behavioral endophenotypes of depression and SSRI outcome, including depression-like behavior, neurogenesis, brain-derived neurotrophic factor levels, hypothalamic-pituitary–adrenal axis activity and long-term potentiation. Results showed that, in line with our hypothesis, the endophenotypes investigated were affected by the treatment according to the quality of the living environment. In particular, mice treated with fluoxetine in an enriched condition overall improved their depression-like phenotype compared with controls, whereas those treated in a stressful condition showed a distinct worsening. Our findings suggest that the effects of SSRI on the depression- like phenotype is not determined by the drug per se but is induced by the drug and driven by the environment. These findings may be helpful to explain variable effects of SSRI found in clinical practice and to device strategies aimed at enhancing their efficacy by means of controlling environmental conditions.


2017 - Hippocampus-related effects of fluoxetine treatment under stressful vs enriched conditions [Articolo su rivista]
Alboni, S.; van Dijk, M.; Poggini, S.; Milior, G.; Perrotta, M. L.; Drenth, T.; Brunello, N.; Wolfer, D.; Limatola, C.; Amrein, I.; Cirulli, F.; Maggi, L.; Branchi, I.
abstract


2017 - Hypothalamic expression of inflammatory mediators in an animal model of binge eating [Articolo su rivista]
Alboni, Silvia; Micioni di Bonaventura, Maria Vittoria; Benatti, Cristina; Giusepponi, Maria; Elena, ; Brunello, Nicoletta; Cifani, Carlo
abstract

Binge eating episodes are characterized by uncontrollable, distressing eating of a large amount of highly palatable food and represent a central feature of bingeing related eating disorders. Research suggests that inflammation plays a role in the onset and maintenance of eating-related maladaptive behavior. Markers of inflammation can be selectively altered in discrete brain region where they can directly or indirectly regulate food intake. In the present study we measured expression levels of different components of cytokine systems (IL-1, IL-6, IL-18, TNF- and IFN-ɣ) and related molecules (iNOS and COX2) in the preoptic and anterior-tuberal parts of the hypothalamus of a validated animal model of binge eating. In this animal model, based on the exposure to both food restriction and frustration stress, binge-like eating behavior for highly palatable food is not shown when animals are exposed to the frustration stress during the estrus phase. We found a characteristic down-regulation of the IL-18/ IL-18 receptor system (with increased expression of the inhibitor of the pro-inflammatory cytokine IL18, IL-18BP, together with a decreased expression of the binding chain of the IL-18 receptor) and a three-fold increase in the expression of iNOS specifically in the anterior- tuberal region of the hypothalamus of animals that develop a binge-like eating behavior. Differently, when food restricted animals were stressed during the estrus phase IL-18 expression increased while iNOS expression was not significantly affected. Considering the role of this region of the hypothalamus in controlling feeding related behavior, this can be relevant in eating disorders and obesity. Our data suggest that by targeting centrally selected inflammatory markers, we may prevent that disordered eating turns into a full blown eating disorder.


2017 - Mitochondrial energy metabolism of rat hippocampus after treatment with the antidepressants desipramine and fluoxetine [Articolo su rivista]
Villa, Roberto Federico; Ferrari, Federica; Bagini, Laura; Gorini, Antonella; Brunello, Nicoletta; Tascedda, Fabio
abstract

Alterations in mitochondrial functions have been hypothesized to participate in the pathogenesis of depression, because brain bioenergetic abnormalities have been detected in depressed patients by neuroimaging in vivo studies. However, this hypothesis is not clearly demonstrated in experimental studies: some suggest that antidepressants are inhibitors of mitochondrial metabolism, while others observe the opposite. In this study, the effects of 21-day treatment with desipramine (15 mg/kg) and fluoxetine (10 mg/kg) were examined on the energy metabolism of rat hippocampus, evaluating the catalytic activity of regulatory enzymes of mitochondrial energy-yielding metabolic pathways. Because of the micro-heterogeneity of brain mitochondria, we have distinguished between (a) non-synaptic mitochondria (FM) of neuronal perikaryon (post-synaptic compartment) and (b) intra-synaptic light (LM) and heavy (HM) mitochondria (pre-synaptic compartment). Desipramine and fluoxetine changed the catalytic activity of specific enzymes in the different types of mitochondria: (a) in FM, both drugs enhanced cytochrome oxidase and glutamate dehydrogenase, (b) in LM, the overall bioenergetics was unaffected and (c) in HM only desipramine increased malate dehydrogenase and decreased the activities of Electron Transport Chain Complexes. These results integrate the pharmacodynamic features of desipramine and fluoxetine at subcellular level, overcoming the previous conflicting data about the effects of antidepressants on brain energy metabolism, mainly referred to whole brain homogenates or to bulk of cerebral mitochondria. With the differentiation in non-synaptic and intra-synaptic mitochondria, this study demonstrates that desipramine and fluoxetine lead to adjustments in the mitochondrial bioenergetics respect to the energy requirements of pre- and post-synaptic compartments.


2016 - Changes in the NMR Metabolic Profile of Live Human Neuron-Like SH-SY5Y Cells Exposed to Interferon-α2 [Articolo su rivista]
Valeria, Righi; Schenetti, Luisa; Mucci, Adele; Benatti, Stefania; Tascedda, Fabio; Brunello, Nicoletta; Carmine, Pariante M; Alboni, Silvia
abstract

Interferon (IFN)-α2 is an extensively therapeutically used pro-inflammatory cytokine. Though its efficacy in controlling viral replication and tumor cells proliferation, administration of IFN-α2 is often associated with the development of central side effects. Magnetic resonance spectroscopy studies have demonstrated that IFN-α2 administration affects brain metabolism, however the exact nature of this effect is not completely known. We hypothesized that IFN-α2 can affect metabolic activity of human neuron-like SH-SY5Y cells which possess many characteristics of neurons and represent one of the most used models for studying mechanisms involved in neurotoxicity or neuroprotection. To test our hypothesis we have characterized the metabolic signature of live SH-SY5Y, and their conditioned media, after 24 and 72 h of exposure to vehicle or IFN-α2 (100 ng/ml) by using High Resolution-Magic Angle Spinning (HR-MAS) Nuclear Magnetic Resonance (NMR) spectroscopy. Our results revealed that 1) the use of HR-MAS NMR is ideally suitable for the characterization of the metabolic profile of live cells and their conditioned media without extraction procedures; and 2) a 72 h exposure to IFN-α2 increases the level of metabolites involved in maintaining energetic (including creatine and lactate) and osmotic (such as myo-inositol, scyllo-inositol, taurine and glycerophosphorylcholine) balances in neuron-like cells and of metabolic waste products (namely lactate, ethanol and acetate), glycine and glutamine in their growth media. These results may contribute to gain more knowledge about the IFN-α2 induced effect on the brain and support the interpretation of magnetic resonance spectroscopy studies performed in humans.


2016 - Disease-induced neuroinflammation and depression [Articolo su rivista]
Benatti, Cristina; Blom, Johanna Maria Catharina; Rigillo, Giovanna; Alboni, Silvia; Zizzi, Francesca; Torta, Riccardo; Brunello, Nicoletta; Tascedda, Fabio
abstract

Progression of major depression, a multifactorial disorder with a neuroinflammatory signature, seems to be associated with the disruption of body allostasis. High rates of comorbidity between depression and specific medical disorders, such as, stroke, chronic pain conditions, diabetes mellitus, and human immunodeficiency virus (HIV) infection, have been extensively reported. In this review, we discuss how these medical disorders may predispose an individual to develop depression by examining the impact of these disorders on some hallmarks of neuroinflammation known to be impaired in depressed patients: altered permeability of the blood brain barrier, immune cells infiltration, activated microglia, increased cytokines production, and the role of inflammasomes. In all four pathologies, blood brain barrier integrity was altered, allowing the infiltration of peripheral factors, known to activate resident microglia. Evidence indicated morphological changes in the glial population, increased levels of circulating pro-inflammatory cytokines or increased production of these mediators within the brain, all fundamental in neuroinflammation, for the four medical disorders considered. Moreover, activity of the kynurenine pathway appeared to be enhanced. With respect to the inflammasome NLRP3, a new target whose role in neuroinflammation is emerging as being important, accumulating data suggest its involvement in the pathogenesis of brain injury following stroke, chronic pain conditions, diabetes mellitus or in HIV associated immune impairment. Finally, data gathered over the last 10 years, indicate and confirm that depression, stroke, chronic pain, diabetes, and HIV infection share a combination of underlying molecular, cellular and network mechanisms leading to a general increase in the neuroinflammatory burden for the individual.


2016 - Effect of desipramine and fluoxetine on energy metabolism of cerebral mitochondria [Articolo su rivista]
Villa, Roberto Federico; Ferrari, Federica; Gorini, Antonella; Brunello, Nicoletta; Tascedda, Fabio
abstract

Brain bioenergetic abnormalities in mood disorders were detected by neuroimaging in vivo studies in humans. Because of the increasing importance of mitochondrial pathogenetic hypothesis of Depression, in this study the effects of sub-chronic treatment (21 days) with desipramine (15 mg/kg) and fluoxetine (10 mg/kg) were evaluated on brain energy metabolism. On mitochondria in vivo located in neuronal soma (somatic) and on mitochondria of synapses (synaptic), the catalytic activities of regulatory enzymes of mitochondrial energy-yielding metabolic pathways were assayed.Antidepressants in vivo treatment modified the activities of selected enzymes of different mitochondria, leading to metabolic modifications in the energy metabolism of brain cortex: (a) the enhancement of cytochrome oxidase activity on somatic mitochondria; (b) the decrease of malate, succinate dehydrogenase and glutamate-pyruvate transaminase activities of synaptic mitochondria; (c) the selective effect of fluoxetine on enzymes related to glutamate metabolism.These results overcome the conflicting data so far obtained with antidepressants on brain energy metabolism, because the enzymatic analyses were made on mitochondria with diversified neuronal in vivo localization, i.e. on somatic and synaptic. This research is the first investigation on the pharmacodynamics of antidepressants studied at subcellular level, in the perspective of (i) assessing the role of energy metabolism of cerebral mitochondria in animal models of mood disorders, and (ii) highlighting new therapeutical strategies for antidepressants targeting brain bioenergetics.


2016 - Erratum to: Changes in the NMR Metabolic Profile of Live Human Neuron-Like SH-SY5Y Cells Exposed to Interferon-α2 (J Neuroimmune Pharmacol, 2015, DOI 10.1007/s11481-015-9641-x) [Altro]
Righi, Valeria; Schenetti, Luisa; Mucci, Adele; Benatti, Stefania; Tascedda, Fabio; Brunello, Nicoletta; Pariante, Carmine M.; Alboni, Silvia
abstract

Erratum


2016 - Fluoxetine prevents Aβ1-42-induced toxicity via a paracrine signaling mediated by transforming-growth-factor-β1 [Articolo su rivista]
Caraci, Filippo; Tascedda, Fabio; Merlo, Sara; Benatti, Cristina; Spampinato, Simona F.; Munafò, Antonio; Leggio, Gian Marco; Nicoletti, Ferdinando; Brunello, Nicoletta; Drago, Filippo; Sortino, Maria Angela; Copani, Agata
abstract

Selective reuptake inhibitors (SSRIs), such as fluoxetine and sertraline, increase circulating Transforming-Growth-Factor-β1 (TGF-β1) levels in depressed patients, and are currently studied for their neuroprotective properties in Alzheimer's disease. TGF-β1 is an anti-inflammatory cytokine that exerts neuroprotective effects against β-amyloid (Aβ)-induced neurodegeneration. In the present work, the SSRI, fluoxetine, was tested for the ability to protect cortical neurons against 1 μM oligomeric Aβ1-42-induced toxicity. At therapeutic concentrations (100 nM-1 μM), fluoxetine significantly prevented Aβ-induced toxicity in mixed glia-neuronal cultures, but not in pure neuronal cultures. Though to a lesser extent, also sertraline was neuroprotective in mixed cultures, whereas serotonin (10 nM-10 μM) did not mimick fluoxetine effects. Glia-conditioned medium collected from astrocytes challenged with fluoxetine protected pure cortical neurons against Aβ toxicity. The effect was lost in the presence of a neutralizing antibody against TGF-β1 in the conditioned medium, or when the specific inhibitor of type-1 TGF-β1 receptor, SB431542, was added to pure neuronal cultures. Accordingly, a 24 h treatment of cortical astrocytes with fluoxetine promoted the release of active TGF-β1 in the culture media through the conversion of latent TGF-β1 to mature TGF-β1. Unlike fluoxetine, both serotonin and sertraline did not stimulate the astrocyte release of active TGF-β1. We conclude that fluoxetine is neuroprotective against Aβ toxicity via a paracrine signaling mediated by TGF-β1, which does not result from a simplistic SERT blockade.


2016 - Fluoxetine treatment affects the inflammatory response and microglial function according to the quality of the living environment [Articolo su rivista]
Alboni, Silvia; Poggini, Silvia; Garofalo, Stefano; Milior, Giampaolo; El Hajj, Hassan; Lecours, Cynthia; Girard, Isabelle; Gagnon, Steven; Boisjoly Villeneuve, Samuel; Brunello, Nicoletta; Wolfer, David P; Limatola, Cristina; Tremblay, Marie Ève; Maggi, Laura; Branchi, Igor
abstract

It has been hypothesized that selective serotonin reuptake inhibitors (SSRIs), the most common treatment for major depression, affect mood through changes in immune function. However, the effects of SSRIs on inflammatory response are contradictory since these act either as anti- or pro-inflammatory drugs. Previous experimental and clinical studies showed that the quality of the living environment moderates the outcome of antidepressant treatment. Therefore, we hypothesized that the interplay between SSRIs and the environment may, at least partially, explain the apparent incongruence regarding the effects of SSRI treatment on the inflammatory response. In order to investigate such interplay, we exposed C57BL/6 mice to chronic stress to induce a depression-like phenotype and, subsequently, to fluoxetine treatment or vehicle (21days) while being exposed to either an enriched or a stressful condition. At the end of treatment, we measured the expression levels of several anti- and pro-inflammatory cytokines and inflammatory mediators in the whole hippocampus and in isolated microglia. We also determined microglial density, distribution, and morphology to investigate their surveillance state. Results show that the effects of fluoxetine treatment on inflammation and microglial function, as compared to vehicle, were dependent on the quality of the living environment. In particular, fluoxetine administered in the enriched condition increased the expression of pro-inflammatory markers compared to vehicle, while treatment in a stressful condition produced anti-inflammatory effects. These findings provide new insights regarding the effects of SSRIs on inflammation, which may be crucial to devise pharmacological strategies aimed at enhancing antidepressant efficacy by means of controlling environmental conditions.


2015 - P.2.a.012 Co-administration of fluoxetine with acetylsalicylic acid, but not flurbiprofen or celecoxib, for one week shows an antidepressant-like effect [Abstract in Rivista]
Alboni, Silvia; Benatti, Cristina; Tascedda, Fabio; Brunello, Nicoletta
abstract

Increasing evidence is now demonstrating the involvement of the immune system and in particular of their effectors, cytokines, in the development and progression of depression. In particular, it is worth underlying how pro-inflammatory cytokines appear to be increased in blood or brain of patients with major depression (MD) and that pharmacological use of pro-inflammatory cytokines (i.e. interferon alpha) may induce MD. These data suggest a role for inflammation in the pathogenesis of depression and that anti-inflammatory drugs may be used as an adjunctive therapy in the treatment of MD. However, some studies reported contradictory results and suggest that adverse effects may contraindicate the use of anti-inflammatory agents in the treatment of depression. Nevertheless, non-steroidal anti-inflammatory drugs (NSAIDs) can have different mechanisms of action also depending on the dose. This is true for the therapeutic effects as well as for the unwanted side effects. On this basis, the present study aimed at evaluating the behavioural effect of the co-administration of fluoxetine (FLX, 5 mg/kg i.p.) with different NSAIDs in the chronic escape model of depression (CED). The CED model of depression possesses face, construct and pharmacological validity and is based on the induction, and maintenance, of an escape deficit upon exposing rats to unavoidable stressors. We previously demonstrated that, in this model, the stress-induced impaired behaviour can be resolved by one week of treatment with the co-administration of FLX (5 mg/kg/i.p.) plus acetylsalicylic acid (ASA, 45 mg/kg i.p.) but not FLX alone [1]. Here we evaluated the effect of the co-administration of FLX and flurbiprofen (FLB, an inhibitor of Cox-1 and Cox-2, 5 mg/kg, p.o.) or celecoxib (CLX, a selective COX2 inhibitor, 5 mg/kg, p.o.) in the CED model after 7 days of treatment. The co-administration FLX plus ASA (45mg/kg i.p.) was used as a positive control. Morever we tested the behavioral effect of different doses (45, 22.5 and 11.25 mg/Kg i.p.) of ASA as potentiating agent of the effect of fluoxetine. Our study shows that only the co-administration of ASA with FLX reverted the stress-induced condition of escape deficit after 7 days of treatment. Moreover, the amplitude of the antidepressant-like effect was dose dependent. The percentage of the antidepressant response was about 90%, 60% and 40% for animals receiving FLX (5 mg/kg/i.p.) plus ASA at the dose of 45, 22.5 or 11.25 mg/Kg i.p. respectively. Both flurbiprofen and celecoxib, when administered together with FLX for 7 days, failed to induce an antidepressant-like effects in the CED model. Higher dose of FLB (50 mg/Kg p.o.) and CLX (20 mg/Kg p.o.) were also tested, but they were associated to high mortality rate (80% and 25% respectively). These data demonstrated that neither all NSAIDs, nor all doses, may be useful in the treatment of depression while adverse effects can be potentiated or induced by the co-administration with antidepressants. Unraveling the cellular and molecular mechanisms behind the dissimilar behavioral response elicited by different anti-inflammatory drugs can contribute to understand the role of inflammation in the etiopathogensis of MD and to improve patient care. [1] Brunello, N., Alboni, S., Capone, G., Benatti, C., Blom, J.M., Tascedda, F., Kriwin, P., Mendlewicz, J., 2006 Shortened onset of action of antidepressants in major depression using acetylsalicylic acid augmentation: a pilot open-label study. Int Clin Psychopharmacol. 21:227-31.


2014 - Behavioural and transcriptional effects of escitalopram in the chronic escape deficit model of depression [Articolo su rivista]
Benatti, Cristina; Alboni, Silvia; Blom, Johanna Maria Catharina; Gandolfi, Francesco; Mendlewicz, Julien; Brunello, Nicoletta; Tascedda, Fabio
abstract

The study of depression is facing major challenges: first, the need to develop new drugs with a faster onset of action and second, fulfilling the unmet needs of treatment resistant patients with more effective compounds. The chronic escape deficit (CED) is a valid and useful model of depression and is based on the induction of an escape deficit after exposure of rats to an unavoidable stress. This behavioural model provides a method for evaluating the capacity of a treatment to revert the escape deficit. The majority of antidepressant drugs need to be administered for at least 3-4 weeks in order to revert the escape deficit. A 7-day treatment with escitalopram reverted the stress-induced escape deficit in approximately 50% of the animals. Escitalopram treatment decreased anxiety-related behaviours in stressed animals, by increasing the time spent in the central part of the arena with respect to saline treated stressed animals, without affecting exploratory related behaviours. Gene expression profiling was carried out in the hippocampus to identify new targets associated with the effects of stress or with the different response to escitalopram. By combining a well-validated animal model with gene expression analysis we demonstrated that the CED model may represent a perfect tool for studying treatment-resistant depression.


2014 - Interleukin 18 activates MAPKs and STAT3 but not NF-κB in hippocampal HT-22 cells [Articolo su rivista]
Alboni, Silvia; Montanari, C; Benatti, Cristina; Sanchez Alavez, M; Rigillo, Giovanna; Blom, Johanna Maria Catharina; Brunello, Nicoletta; Conti, B; Pariante, Mc; Tascedda, Fabio
abstract

Interleukin (IL)-18 is a cytokine previously demonstrated to participate in neuroinflammatory processes. Since the components of the IL-18 receptor complex are expressed in neurons throughout the brain, IL-18 is also believed to directly influence neuronal function. Here we tested this hypothesis on mouse hippocampal neurons by measuring the effects of IL-18 on three pathways previously shown to be regulated by this cytokine in non-neuronal cells: the MAPK pathways, p38 and ERK1/2 MAPKs, STAT3 and NF-κB. Experiments were carried out in vitro using the immortalized hippocampal neuronal line HT-22 or in vivo following i.c.v. injection with recombinant mouse IL-18. We showed that IL-18 did not activate NF-κB in HT-22 cells whereas it induced a rapid (within 15min) activation of the MAPK pathways. Moreover, we demonstrated that IL-18 treatment enhanced P-STAT3 (Tyr705)/STAT3 ratio in the nucleus of HT-22 cells after 30-60min of exposure. A similar increase in P-STAT3 (Tyr705)/STAT3 ratio was observed in the whole hippocampus one hour after i.c.v. injection. These data demonstrate that IL-18 can act directly on neuronal cells affecting the STAT3 pathway; therefore, possibly regulating the expression of specific genes within the hippocampus. This effect may help to explain some of the IL-18-induced effects on synaptic plasticity and functionality within the hippocampal system.


2014 - Successful treatment of HIV-1 infection increases the expression of a novel, short transcript for IL-18 receptor α chain [Articolo su rivista]
Nasi, Milena; Alboni, Silvia; Pinti, Marcello; Tascedda, Fabio; Benatti, Cristina; Benatti, Stefania; Gibellini, Lara; DE BIASI, Sara; Borghi, Vanni; Brunello, Nicoletta; Mussini, Cristina; Cossarizza, Andrea
abstract

The importance of interleukin (IL)-18 in mediating immune activation during HIV infection has recently emerged. IL-18 activity is regulated by its receptor (IL-18R), formed by an α and a β chain, the IL-18-binding protein, and the newly identified shorter isoforms of both IL-18R chains. We evaluated gene expression of the IL-18/IL-18R system in peripheral blood mononuclear cells from HIV+ patients. Compared with healthy donors, IL-18 expression decreased in patients with primary infection. The IL-18Rα short transcript expression was strongly upregulated by successful highly active antiretroviral therapy. HIV progression and its treatment can influence the expression of different components of the complex IL-18/IL-18R system.


2013 - Chronic antidepressant treatments resulted in altered expression of genes involved in inflammation in the rat hypothalamus [Articolo su rivista]
Alboni, Silvia; Benatti, Cristina; Montanari, Claudia; Tascedda, Fabio; Brunello, Nicoletta
abstract

To gain insight into the possible immune targets of antidepressant, we evaluated the expression of several inflammatory mediators in the hypothalamus of rats chronically (28 days) treated with the serotonin selective reuptake inhibitor fluoxetine (5mg/kg, i.p.) or the tricyclic compound imipramine (15 mg/kg, i.p.). We focused our attention on the hypothalamus as it plays a key role in determining many of the somatic symptoms experienced by depressed patients. This brain region, critical also for expression of motivated behaviours, participates in the control of the hypothalamic-pituitary-adrenal axis activity and in stress response as well as coordinates physiological functions such as sleep and food intake that have been found altered in a high percentage of depressed patients. Notably, hypothalamus is a key structure for brain cytokine expression and function as it integrates signals from the neuro, immune, endocrine systems. By means of quantitative Real Time PCR experiments we demonstrated that a chronic treatment with either fluoxetine or imipramine resulted in a reduction of IL-6 and IFN-γ mRNAs and increased IL-4 mRNA expression in the rat hypothalamus. Moreover, we demonstrated that hypothalamic expression of members of IL-18 system was differentially affected by chronic antidepressant treatments. Chronically administered fluoxetine decreased IL-8 and CX3CL1 hypothalamic expression, while a chronic treatment with imipramine decreased p11 mRNA. Our data suggest that a shift in the balance of the inflammation toward an anti-inflammatory state in the hypothalamus may represent a common mechanism of action of both the chronic treatments with fluoxetine and imipramine.


2013 - Interferon alpha exposure increases the expression of the enzymes belonging to the kynurenine pathway in an in vitro model of human neurons: SH-SY5Y cells [Abstract in Rivista]
Alboni, Silvia; Benatti, Cristina; Claudia, Montanari; Benatti, Stefania; Tascedda, Fabio; Cannazza, Giuseppe; Pariante Carmine, M; Brunello, Nicoletta
abstract

The past two decades have witnessed a burgeoning area of pre-clinical and clinical research linking psychiatric illnesses – particularly major depression (MD) – to activation of the inflammatory immune system. One of the stronger evidence supporting a causal role for inflammation in leading MD comes from reports indicating that depressive symptoms frequently develop in patients undergoing immunotherapy with cytokines, such as interferon (IFN)-α, for the treatment of malignancies or chronic viral infection. Although INF-alpha- induced effects on the brain made of IFN-α a model to study the influence of pro-inflammatory cytokines in the CNS and behavior the molecular mechanisms underlying these effects are far from being fully understood. It has been proposed that IFN-α may contribute to the etiology of MD by inducing indolamine 2,3-dioxygenase (IDO) expression and thus unbalancing in the tryptophan/kynurenine metabolism toward the production of neurotoxic metabolites and\or reducing serotonin (5-HT) availability. IDO catalyzes the initial rate-limiting step in tryptophan degradation along the kynurenine pathway (KP). Kynurenine, the initial product of tryptophan degradation, is further catalysed into neurotoxic end-products through steps catalyzed by kynurenine 3-monooxygenase (KMO) and kynureninase (Kynu). However, Kynurenine can also be catabolised by kynurenine aminotransferase (KAT), into kynurenic acid, a potentially neuroptotective agent. A role for a disturbance in the equilibrium between neurotoxic/ neuropoptective end KP endproducts producing an alteration in the neuroprotective–neurodegenerative balance in the brain of patients with MD, has been proposed in the neurodegeneration hypothesis of depression. Given that we previously demonstrated that IFN-α induces toxic effects in an in vitro model of human neurons (human SH-SY5Y neuroblastoma cells) we were aim to investigate the effects of IFN-α on KP in these cells. Our studies show that IFN-α exposure increased the expression of all the kynurenergic enzymes investigated (IDO, KMO, Kynu and KAT). More particularly strongly induced the expression of IDO mRNA (more than 900 –fold) in SH-SY5Y cells. Similar effects on kynurenergic enzyme expression were also observed when SH-SY5Y cells where differentiated with all-trans retinoic acid (in presence of neurotrophic support and in serum deprived conditions). We also demonstrated that INF-α decreased 5-HT levels whereas increased the kynurenine levels in the medium of both differentiated as well not differentiated SH-SY5Y cells.


2013 - Metabolic changes induced by interferon-α exposure in an in vitro model of human neurons [Abstract in Rivista]
Alboni, Silvia; Schenetti, Luisa; Brunello, Nicoletta; Pariante Carmine, M; Righi, Valeria
abstract

The human neuroblastoma SH-SY5Y cell line is a third successive subclone of the SK-N-SH line, originally established from a bone marrow biopsy of a neuroblastoma patient. These cells possess many characteristics of neurons, and they represent one of the most-used models for studying cellular events and mechanisms involved in neurotoxicity and neurodegeneration or even in neuroprotection. We have been using these cells as a tools to evaluate the largely unexplored effects induced by interferon (IFN)- alpha (a clinically used type I IFN) exposure on neurons. Interferons are cytokines endowed with a pleiotropic spectrum of biological properties, including immunomodulation, antiviral and pro-inflammatory activity. Beside the periphery, and cells of the immune system, type I IFNs may have broad-ranging actions also in the brain, affecting neuronal differentiation, survival and synaptic plasticity. We recently demonstrated that exposure to IFN-α induces neurotoxic effects in a time e dose dependent manner in SH-SY5Y cells by impairing mitochondrial integrity and activity, recruitment of Bcl-2 family members, induction of oxidative stress (increases reactive oxygen species). Indeed following 72 hours exposure to this cytokine we found increased early apoptosis (Alboni et al., 2013). This prompted us to investigate the changes in the metabolic profile of live SH-SY5Y cells exposed to IFN-α (72 h) using 1H High Resolution-Magic Angle Spinning nuclear magnetic resonance (HR-MAS NMR) spectroscopy. Firstly, this technique enabled us to characterize the metabolic signature of intact SH-SY5Y cells. Several metabolites, including amino acids, osmolites, phospholipids, organic acids, sugars and polyols have been identified. Moreover, we found that human neuronal cells exposed to IFN-α for 72 h had significantly increased concentration of lactate (% 93.9), taurine (% 117.7), myo-inositol (% 123.2), scyllo-inositol (% 117.4), glycerolphosphocholine (% 135.2) and creatine (% 159.9) compared with the vehicle-treated control cells. These data provide the demonstration that IFN-α exposure induces metabolic changes in human neuron-like cells. Moreover, these results may contribute to explain IFN-α-induced central side-effects often observed following IFN-α treatment for viral infection and malignancies.


2013 - Metabolic characterization of SH-SY5Y cells and effects of interferon-alpha exposure [Abstract in Atti di Convegno]
Alboni, Silvia; Schenetti, Luisa; Mucci, Adele; Brunello, Nicoletta; Valeria, Righi
abstract

The human neuroblastoma SH-SY5Y cell line is a third successive subclone of the SK-N-SH line, originally established from a bone marrow biopsy of a neuroblastoma patient. These cells possess many characteristics of neurons, and they represent one of the most-used models for studying cellular events and mechanisms involved in neurotoxicity and neurodegeneration or even in neuroprotection. Interferons are cytokines endowed with a pleiotropic spectrum of biological properties, including immunomodulation, antiviral and proinflammatory activity. Interferon (IFN)-α is a type I IFN that may have broad-ranging actions in the brain, affecting neuronal differentiation, survival and synaptic plasticity. We previously demonstrated that a 72 hours exposure to IFN-α induces early apoptosis in SH-SY5Y cells. This prompted us to investigate the metabolic profile of the SH-SY5Y cells using HR-MAS NMR Spectroscopy after a 72 hours exposure to IFN-α to explore the metabolic changes that characterize these cells. Moreover, since a metabolic characterization of this extensively used cell clone is still lacking, we analyzed the metabolic profile of the SH-SY5Y in standard growth conditions. Results show some interesting changes in metabolites, such as choline containing compounds, creatine and glutamate. Our goal will be to relate the metabolic changes to IFN-α exposure.


2013 - N-acetyl-cysteine prevents toxic oxidative effects induced by IFN-α in human neurons. [Articolo su rivista]
Alboni, Silvia; Gibellini, Lara; Montanari, Claudia; Benatti, Cristina; Benatti, Stefania; Tascedda, Fabio; Brunello, Nicoletta; Cossarizza, Andrea; Pariante, Carmine M.
abstract

Currently IFN-α is widely used for effective treatment of viral infections and several malignancies. However, IFN-α can cause neuropsychiatric disturbances and mental impairments, including fatigue, insomnia, depression, irritability and cognitive deficits. Molecular and cellular mechanisms leading to such side-effects are still poorly understood. Neurons seem to be an important target in mediating cellular effects induced by exposure to this cytokine, but so far little is known about IFN-α-induced effects on these cells. We have investigated the ability of IFN-α (2-100 ng/ml) to induce damage and toxicity to the human neuroblastoma SH-SY5Y cell line, commonly used for studying such phenomena, and the mechanisms underlying these effects. After 24 h treatment, IFN-α increased mitochondrial activity, whereas cell density was reduced in a dose- and time-dependent manner. This effect did not depend on reduced cell proliferation, but rather the activation of apoptosis, as revealed by an increased Bax:Bcl-2 mRNA ratio after 72-h IFN-α exposure. At this time-point, IFN-α also reduced the expression of the brain-derived neurotrophic factor gene, and induced an increase in reactive oxygen species (ROS). A co-treatment with N-acetyl-cysteine (NAC; 5 mm), a potent antioxidant and mitochondrial modulator, was able to counteract all of these IFN-α-induced effects. These findings demonstrated that IFN-α induces neurotoxicity and apoptosis that is, in part, very likely due to mitochondrial damages and production of ROS. We suggest that NAC, already tested for the treatment of psychiatric disorders, may be useful to prevent IFN-α-induced central side-effects in a safe and effective way.


2012 - Interferon-alpha exposure increases the expression of enzymes of the kynurenine pathway and induces apoptosis in a model of human neurons [Abstract in Rivista]
Alboni, Silvia; Benatti, Cristina; Montanari, Claudia; Tascedda, Fabio; C. M., Pariante; Brunello, Nicoletta
abstract

Major depression (MD) is associated with a profound unbalance between the nervous-, the endocrine- and the immune- systems. This suggests the possibility that molecules that regulate the homeostasis of these systems may contribute to the development of MD. For instance, some cytokines, important neuro-endocrineimmuno modulators, have been proposed to have a role in MD as supported by the observation that activation of immune system with therapeutically used cytokines may induce MD. IFN-a is an innate immune cytokine with potent antiviral and anti-proliferative properties that is used to treat viral infections, such as hepatitis C, and certain cancers. Despite therapeutic efficacy in these illnesses, it has been observed that IFN-a exposure may be associated with important side effects including neuropsychological and behavioural changes that overlap with MD. Although IFN-a- induced effects on the brain make IFN-alpha a model to study the influence of pro-inflammatory cytokines in the CNS and behavior, the molecular mechanisms underlying these effects are far from being fully understood. It has been proposed that cytokines may contribute to the etiology of MD by inducing indolamine 2,3-dioxygenase (IDO) expression. IDO catalyzes the initial rate-limiting step in tryptophan (TPR) degradation along the kynurenine pathway (KP). Kynurenine, the initial product of TPR degradation, is further catalysed into neurotoxic end-products through steps catalyzed by kynurenine 3-monooxygenase (KMO) and kynureninase (Kynu). However, Kynurenine can also be catabolised by kynurenine aminotransferase (KAT), into kynurenic acid, a potentially neuroprotective agent. A role for a disturbance in the KP in the neuroprotective–neurodegenerative balance in the brain of patients with MD, has been proposed in the neurodegeneration hypothesis of depression. This prompted us to investigate the effects of IFN-a on the expression of the IDO, KMO, Kynu and KAT mRNAs in an in vitro model of human neurons: SH-SY5Y (human neuroblastoma) cells. We also evaluated the IFN-a-induced effect on cell viability and number in these cells. Finally, because cell number results from the balance between cell proliferation and cell elimination, we measured cell proliferation and apoptosis in SH-SY5Y cells after IFN-a exposure. The same experiments were performed in differentiated SH-SY5Y cells with retinoic acid (RA) and Brain- Derived Neurotrophic Factor (BDNF). Our studies show that IFN-a exposure increased the expression of all the kynurenergic enzymes investigated (with an unbalance of the KP toward the synthesis of neurotoxic end-products) and in more particularly strongly induced the expression of IDO mRNA (more than 900-fold) in SH-SY5Y cells. We also demonstrated that IFN-a reduced in a dose- (2, 20, 50 and 100 ng/ml) and time- (24, 48 and 72 hrs) dependent manner the cell number and induces apoptosis in SH-SY5Y cells. Similar results were obtained in SHSY5Y RA/BDNF differentiated cells. Together our results clearly enlighten the cytotoxic effects of IFN-a in this in vitro model of human neurons. Moreover, our findings provide further information on the molecular pathways involved in cytokine-induced effects in the brain and add a piece to the puzzle of what and how these factors or pathways may contribute to the pathogenesis of MD.


2012 - Transcriptional profiles underlying vulnerability and resilience in rats exposed to an acute unavoidable stress [Articolo su rivista]
Benatti, Cristina; Valensisi, Cristina; Blom, Johanna Maria Catharina; Alboni, Silvia; Montanari, Claudia; Ferrari, Francesco; Tagliafico, Enrico; Julien, Mendlewicz; Brunello, Nicoletta; Tascedda, Fabio
abstract

A complex interplay between gene and environment influences the vulnerability or the resilience to stressful events. In the acute escape deficit (AED) paradigm, rats exposed to an acute unavoidable stress (AUS) develop impaired reactivity to noxious stimuli. Here we assessed the behavioral and molecular changes in rats exposed to AUS. A genome-wide microarray experiment generated a comprehensive picture of changes in gene expression in the hippocampus and the frontal cortex of animals exposed or not to AUS. Exposure to AUS resulted in two distinct groups of rats with opposite behavioral profiles: one developing an AED, called “stress vulnerable,” and one that did not develop an AED, called “stress resilient.” Genome-wide profiling revealed a low percentage of overlapping mechanisms in the two areas, suggesting that, in the presence of stress, resilience or vulnerability to AUS is sustained by specific changes in gene expression that can either buffer or promote the behavioral and molecular adverse consequences of stress. Specifically, we observed in the frontal cortex a downregulation of the transcript coding for interferon-β and leukemia inhibitory factor in resilient rats and an upregulation of neuroendocrine related genes, growth hormone and prolactin, in vulnerable rats. In the hippocampus, the muscarinic M2 receptor was downregulated in vulnerable but upregulated in resilient rats. Our findings demonstrate that opposite behavioral responses did not correspond to opposite regulatory changes of the same genes, but resilience rather than vulnerability to stress was associated with specific changes, with little overlap, in the expression of patterns of genes.


2012 - cAMP-dependent binding proteins and endogenous phosphorylation after antidepressant treatment [Capitolo/Saggio]
Racagni, G.; Tinelli, D.; Bianchi, E.; Brunello, N.; Perez, J.
abstract

Among the most significant changes induced in monoaminergic neurones by chronic antidepressant treatment are: reduction in tyrosine hydroxylase activity, decrease in the ability of NA to stimulate the activity of adenylate cyclase, and reduction in the concentration of noradrenergic and serotonergic receptors. Many synaptic and transsynaptic mechanisms may participate in the desensitization of neurotransmitter receptors after chronic treatment with antidepressants. The most commonly described effector mechanism beyond the second messengers depends on protein phosphorylation mediated by activation of specific protein serine-threonine kinases. Components of the protein phosphorylation system are associated with the cytoskeleton. This chapter investigates whether the cAMP-dependent phosphorylation system associated with microtubules, which are constituents of neuronal cytoskeleton, could be an intracellular target for antidepressants acting on NA and 5-HT neurones.


2011 - Central effects of a local inflammation in three commonly used mouse strains with a different anxious phenotype [Articolo su rivista]
Benatti, Cristina; Alboni, Silvia; Montanari, Claudia; Caggia, Federica; Tascedda, Fabio; Brunello, Nicoletta; Blom, Johanna Maria Catharina
abstract

As in humans, genetic background in rodents may influence a peculiar set of behavioural traits such as sensitivity to pain and stressors or anxiety-related behaviours. Therefore, we tested the hypothesis that mice with different genetic backgrounds [outbred (CD1), inbred (C57BL/6J) and hybrid (B6C3F1) adult male mice] display altered reactivity to pain, stress and anxiety related behaviours. We demonstrated that B6C3F1 mice displayed the more anxious phenotype with respect to C57BL/6J or CD1 animals, with the latter being the less anxious strain when tested in an open field and on an elevated plus maze. No difference was observed across strains in thermal sensitivity to a radiant heat source. Mice were then treated with a sub-plantar injection of the inflammatory agent Complete Freund's Adjuvant (CFA), 24h later they were hyperalgesic with respect to saline exposed animals, irrespective of strain. We then measured intra-strain differences and CFA-induced inter-strain effects on the expression of various genes with a recognized role in pain and anxiety: BDNF, IL-6, IL-1β, IL-18 and NMDA receptor subunits in the mouse thalamus, hippocampus and hypothalamus. The more anxious phenotype observed in B6C3F1 hybrid mice displayed lower levels of BDNF mRNA in the hippocampus and hypothalamus when compared to outbred CD1 and C57BL/6J inbred mice. CFA led to a general decrease in central gene expression of the evaluated targets especially in CD1 mice, while BDNF hypothalamic downregulation stands out as a common effect of CFA in all three strains evaluated


2011 - Constitutive and LPS-regulated expression of interleukin-18 receptor beta variants in the mouse brain [Articolo su rivista]
Alboni, Silvia; Montanari, Claudia; Benatti, Cristina; Blom, Johanna Maria Catharina; Simone, Ml; Brunello, Nicoletta; Caggia, Federica; Guidotti, G; Marcondes, Mc; Sanchez Alavez, M; Conti, B; Tascedda, Fabio
abstract

Interleukin (IL)-18 is a pro-inflammatory cytokine that is proposed to be involved in physiological as well as pathological conditions in the adult brain. IL-18 acts through a heterodimer receptor comprised of a subunit alpha (IL-18Rα) required for binding, and a subunit beta (IL-18Rβ) necessary for activation of signal transduction. We recently demonstrated that the canonical alpha binding chain, and its putative decoy isoform, are expressed in the mouse central nervous system (CNS) suggesting that IL-18 may act on the brain by directly binding its receptor. Considering that the co-expression of the beta chain seems to be required to generate a functional receptor and, a short variant of this chain has been described in rat and human brain, in this study we have extended our investigation to IL-18Rβ in mouse. Using a multi-methodological approach we found that: (1) a short splice variant of IL-18Rβ was expressed in the CNS even if at lower levels compared to the full-length IL-18Rβ variants, (2) the canonical IL-18Rβ is expressed in the CNS particularly in areas and nuclei belonging to the limbic system as previously observed for IL-18Rα and finally (3) we have also demonstrated that both IL-18Rβ isoforms are up-regulated in different brain areas three hours after a single lipopolysaccharide (LPS) injection suggesting that IL-18Rβ in the CNS might be involved in mediating the endocrine and behavioral effects of LPS. Our data highlight the considerable complexity of the IL-18 regulation activity in the mouse brain and further support an important central role for IL-18.


2011 - Stress induces altered CRE/CREB pathway activity and BDNF expression in the hippocampus of glucocorticoid receptor-impaired mice [Articolo su rivista]
Alboni, Silvia; Tascedda, Fabio; Corsini, Daniela; Benatti, Cristina; Caggia, Federica; Capone, Giacomo; Barden, N; Blom, Johanna Maria Catharina; Brunello, Nicoletta
abstract

The gene coding for the neurotrophin Brain-Derived Neurotrophic Factor (BDNF) is a stress-responsive gene. Changes in its expression may underlie some of the pathological effects of stress-related disorders like depression. Data on the stress-induced regulation of the expression of BDNF in pathological conditions are rare because often research is conducted using healthy animals. In our experiments, we used transgenic mice with glucocorticoid receptor impaired (GR-i) expression in the hypothalamus created as a tool to study the neuroendocrine changes occurring in stress-related disorders. First, under basal condition, GR-i mice displayed lower levels of BDNF exons IX and IV and decreased CRE(BDNF) binding activity with respect to wild-type (WT) mice in the hippocampus. Then, we exposed GR-i and WT mice to an acute restraint stress (ARS) to test the hypothesis that GR-i mice display: 1] different ARS induced expression of BDNF, and 2] altered activation of signaling pathways implicated in regulating BDNF gene expression in the hippocampus with respect to WT mice. Results indicate that ARS enhanced BDNF mRNA expression mainly in the CA3 hippocampal sub-region of GR-i mice in the presence of enhanced levels of pro-BDNF protein, while no effect was observed in WT mice. Moreover, ARS reduced CREB signaling and binding to the BDNF promoter in GR-i mice but enhanced signaling and binding, possibly through ERK1/2 activation, in WT mice. Thus, life-long central GR dysfunction resulted in an altered sensitivity at the transcriptional level that may underlie an impaired response to an acute psycho-physical stress.


2010 - Erratum: HTR1B as a risk profile maker in psychiatric disorders: A review through motivation and memory (European Journal of Clinical Pharmacology DOI:10.1007/s00228-009-0724-6) [Articolo su rivista]
Drago, A.; Alboni, S.; Brunello, N.; De Ronchi, D.; Serretti, A.
abstract


2010 - HTR1B as a risk profile maker in psychiatric disorders:a review through motivation and memory [Articolo su rivista]
Antonio, Drago; Alboni, Silvia; Brunello, Nicoletta; Diana De, Ronchi; Alessandro, Serretti
abstract

Purpose Serotonin receptor 1B (HTR1B) is involved in theregulation of the serotonin system, playing different roles inspecific areas of the brain. We review the characteristics ofthe gene coding for HTR1B, its product and the functionalrole of HTR1B in the neural networks involved inmotivation and memory; the central role played by HTR1Bin these functions is thoroughly depicted and show HTR1Bto be a candidate modulator of the mnemonic andmotivationally related symptoms in psychiatric illnesses.Methods In order to challenge this assessment, we analyzehow and how much the genetic variations located in thegene that codes for HTR1B impacts on the psychiatricphenotypes by reviewing the literature on this topic.Results We gathered partial evidence arising from geneticassociation studies, which suggests that HTR1B plays arelevant role in substance-related and obsessive compulsivedisorders. On the other hand, no solid evidence for otherpsychiatric disorders was found. This finding is quitestriking because of the heavy impairment of motivationand of mnemonic-related functions (for example, recallbias) that characterize major psychiatric disorders.Conclusions The possible reasons for the contrast betweenthe prime relevance of HTR1B in regulating memory andmotivation and the limited evidence brought by geneticassociation studies in humans are discussed, and somesuggestions for possible future directions are provided.


2010 - Time-dependent effects of escitalopram on brain derived neurotrophic factor (BDNF) and neuroplasticity related targets in the central nervous system of rats [Articolo su rivista]
Alboni, Silvia; Benatti, Cristina; Capone, Giacomo; Corsini, Daniela; Caggia, Federica; Tascedda, Fabio; Mendlewicz, J; Brunello, Nicoletta
abstract

Chronic treatment with antidepressants affects several proteins linked to neuroplasticity, particularly brain derived neurotrophic factor (BDNF): this leads eventually to their therapeutic effects. It is possible that also for putative early therapeutic onset, antidepressants may act by promoting cellular adaptations linked to neuroplasticity. Escitalopram, known to be already effective in preclinical models of depression after 7 days, allowed us to investigate whether two effective treatment regimens (7 and 21 days) may contribute to synaptic plasticity by acting on BDNF signalling. We focused our attention on two regulators of BDNF transcription, CREB and CaRF (calcium responsive factor), and on kinases, CaMKII, ERK1/2 and p38 MAPK, linked to BDNF that play a distinctive role in synaptic plasticity. We evaluated whether the effects of escitalopram on these targets may be different in brain areas involved in the depressive symptomatology (hippocampus, frontal and prefrontal cortex). Here we demonstrate that escitalopram regulates intracellular pathways linked to neuroplasticity at both the time points evaluated in an area-specific manner. While the two escitalopram-treatment regimens failed to affect gene expression in the rat frontal cortex, 7days of treatment with escitalopram activated intracellular pathways linked to BDNF and increased the levels of Pro-BDNF in the rat prefrontal cortex. Moreover, 21 days of treatment with escitalopram decreased CREB/BDNF signalling while increasing p38 levels in the rat hippocampus. Even if further experiments with different antidepressant strategies will be needed, our data suggest that escitalopram efficacy may be mediated by early and late effects on synaptic plasticity in selective brain areas.


2009 - Chronic treatment with the selective NOP receptor antagonist [Nphe1,Arg14,Lys15]N/OFQ-NH2 (UFP-101) reverses the behavioural and biochemical effects of unpredictable chronic mild stress in rats [Articolo su rivista]
Vitale, Giovanni; Ruggieri, Valentina; Filaferro, Monica; Frigeri, Claudio; Alboni, Silvia; Tascedda, Fabio; Brunello, Nicoletta; Guerrini, R; Cifani, C; Massi, M.
abstract

Introduction The present study was designed to assess the antidepressant effects of UFP-101, a selective nociceptin/orphanin FQ peptide (NOP) receptor antagonist, in a validated animal model of depression: the chronic mild stress (CMS). Materials and methods and Results UFP-101 (5, 10 and 20 nmol/rat; i.c.v., once a day for 21 days) dose- and time-dependently reinstated sucrose consumption in stressed animals without affecting the same parameter in non-stressed ones. In the forced swimming test, UFP-101 reduced immobility of stressed rats from day 8 of treatment. After a 3-week treatment, rats were killed for biochemical evaluations. UFP-101 abolished increase in serum corticosterone induced by CMS and reverted changes in central 5-HT/5-HIAA ratio. The behavioural and biochemical effects of UFP-101 mimicked those of imipramine, the reference antidepressant drug, administered at the dose of 15 mg/kg (i.p.). Co-administration of nociceptin/orphanin FQ (5 nmol/rat, from day 12 to 21) prevented the effects of UFP-101. Brain-derived neurotrophic factor mRNA and protein in hippocampus were not reduced by CMS nor did UFP-101 modify these parameters. Discussion and Conclusion This study demonstrated that chronic treatment with UFP-101 produces antidepressant-like effects in rats subjected to CMS supporting the proposal that NOP receptors represent a candidate target for the development of innovative antidepressant drugs.


2009 - Early neonatal inflammation affects adult pain reactivity and anxiety related traits in mice: genetic background counts [Articolo su rivista]
Benatti, Cristina; Alboni, Silvia; Capone, Giacomo; Corsini, Daniela; Caggia, Federica; Brunello, Nicoletta; Tascedda, Fabio; Blom, Johanna Maria Catharina
abstract

Protracted or recurrent pain and inflammation in the early neonatal period may cause long-lasting changes in central neural function. However, more research is necessary to better characterize the long-term behavioral sequelae of such exposure in the neonatal period. Objectives: (1) to study whether timing of postnatal exposure to persistent inflammation alters responsiveness to thermal pain in the adult animal; (2) to assess whether animals experiencing early postnatal chronic inflammation display altered anxiety related behavior; (3) to study the importance of genetic background. Newborn mice (outbred strain, CD1 and F1 hybrid strain, B6C3F1) received an injection of Complete Freund's Adjuvant (CFA) or saline on either postnatal day 1 or 14 (PND1; PND14) into the left hind paw. Pain to radiant heat and anxiety were examined in 12-week-old adult animals. Adult baseline PWL was significantly decreased in CD1 mice exposed to CFA on PND 1 and 14 as compared to their saline treated counterparts. B6C3F1 mice exposed to CFA on PND14 showed markedly reduced baseline PWL compared to the PND14 saline group. Persistent inflammation experienced by B6C3F1 mice on PND1 failed to affect baseline adult thermal responsiveness. Adult mice, CD1 and B6C3F1, displayed low anxiety traits only if they had been exposed to persistent inflammation on PND1 and not on PND14. Our research suggests a role for genetic background in modulating long-term behavioral consequences of neonatal persistent inflammation: the data support the hypothesis that pain experienced very early in life differentially affects adult behavioral and emotional responsiveness in outbred (CD1) and hybrid mice (B6C3F1).


2009 - Gene expression profile of the hippocampus of a behavioural model of depression [Abstract in Rivista]
Valensisi, Cristina; Caggia, Federica; Alboni, Silvia; Benatti, Cristina; Ferrari, F; Mendlewicz, J; Blom, Johanna Maria Catharina; Brunello, Nicoletta; Tascedda, Fabio
abstract

Although the neurobiological basis of depression has not been fully elucidated, numerous studies have emphasized that in the etiology of depression stress may be the most significant cause, together with genetic vulnerability. Stress induces a coordinated and complex response that is adaptive and integral to survival. The brain's ability to adapt and change over time is refered to neuroplasticity and long-term plasticity in the brain requires changes in gene expression. However, exposure to intense or chronic stressors leads to an increased risk for the development of stress-related disorders including major depression. Numerous studies demonstrate that neuronal atrophy and loss of plasticity occur in hippocampus in response to stress and depression. Therefore, the hippocampal region may play a central role in depressive illness. Likewise changes in gene expression underlying the plasticity of hippocampal structures appear to be relevant in undenstanding the molecular and cellular mechanisms involved in the etiology as well as the treatment of depression, and the mechanisms leading vulnerability or resilience to stress. In fact, humans display a remarkable heterogeneity in their responses to stress and adversity. Although we are beginning to understand how maladaptive neurobiological changes may contribute to depression, relatively little is known about the molecular mechanisms that may underlie stress resilience. Here we set out to investigate the changes in the gene expression profile underlying the effects of stress on the hippocampus using a behavioural paradigm of depression, the chronic escape deficit model [1], which is based on the modified reactivity of rats to external stimuli, the escape deficit, induced by exposure to intense and unavoidable stress. The chronic escape deficit model starts as an acute escape deficit which can be indefinitely sustained by repeated administration of mild stressors. This approach has proved to be a valid and useful model of depression because it consider depressive symptoms like behavioural despair. We performed gene expression profiling in the rat hippocampus, using GeneChip Rat Exon Array (Affymetrix). Using this new platform we carried out analyses of gene expression on three different levels: gene, transcript and exon level analyses. The behavioural results showed that exposure to intense and unavoidable stressful procedure induced escape deficit only in 60% of them. Whereas the animals remaining display a behaviour apparently identical to control animals which did not undergo the stressful procedure. Comparing gene expression profiles and performing functional analysis on differently expressed genes we have indicated multiple pathways that may be involved in the underlying mechanisms of stress condition associated with escape deficit. Moreover we identified possible cellular functions and biological processes that could represent targets that may contribute to mediate the effects of stress on the hippocampal plasticity. Such as, gene expression profiling of stress-vulnerable and stress-resilient animals revealed distinct transcriptional profiles, suggesting that resilient behaviour represents an active neurobiological process and not simply the absence of vulnerability.


2009 - Mapping of the full length and the truncated interleukin-18 receptor alpha in the mouse brain [Articolo su rivista]
Alboni, Silvia; Cervia, D; Ross, B; Montanari, Claudia; Gonzalez, As; Sanchez Alavez, M; Marcondes, Mc; De Vries, D; Sugama, S; Brunello, Nicoletta; Blom, Johanna Maria Catharina; Tascedda, Fabio; Conti, B.
abstract

The cytokine IL-18 acts on the CNS both in physiological and pathological conditions. Its action occurs through the heterodimeric receptor IL-18Rα β. To better understand IL-18 central effects, we investigated in the mouse brain the distribution of two IL-18Rα transcripts, a full length and an isoform lacking the intracellular domain hypothesized to be a decoy receptor. Both isoforms were expressed in neurons throughout the brain primarily with overlapping distribution but also with some unique pattern. These data suggest that IL-18 may modulate neuronal functions and that its action may be regulated through expression of a decoy receptor.


2009 - Microarray analysis in hippocampus of rats treated with escitalopram in the chronic escape deficit model of depression [Abstract in Rivista]
Caggia, Federica; Valensisi, Cristina; Alboni, Silvia; Benatti, Cristina; Corsini, Daniela; F., Ferrari; Tagliafico, Enrico; J., Mendlewicz; Tascedda, Fabio; Brunello, Nicoletta
abstract

Currently, the biological bases of depression and the molecular mechanisms underlying antidepressant action are not completely understood. Valuable tools to better understand the pathophysiology of this disease are behavioural models of depression eventually combined with genome-wide gene expression analysis. The Chronic Escape Deficit (CED) is a validated behavioural model of depression, based on the induction of an escape deficit after exposure of rats to an unavoidable stress. This model allows to evaluate the capacity of a treatment to revert the escape deficit. The antidepressant drugs tested in CED model need to be administered for at least 3−4 weeks in order to revert the escape deficit [1,2]. In this study, we demonstrated that already after one week of treatment with Escitalopram, a widely used SSRI, 50% of the animals responded reverting the escape deficit induced. Moreover, the other 50% of treated animals did not respond also after 3−4 weeks of treatment. Since in the CED model the behavioural alteration is induced by stress application and reverted by escitalopram treatment in only half of animals, the aims of our study were two fold: (i) to investigate transcriptional changes activated by stress; (ii) to study the different gene expression pattern involved into mechanisms of the response and not response to the pharmacological treatment. To address these issues we performed a microarray experiment in the rat hippocampus using Affymetrix GeneChip Rat Exon 1.0 ST evaluating both gene-level and exon-level expression profiling on the whole genome. Total RNA extracted from hippocampus of each animal was utilized to chip a single array using the Affymetrix protocols. 20 single arrays were utilized for data analysis and divided into five replicates for each experimental group (control, stress, stress-escitalopram responders and stress escitalopram-not responders). Using two parallel analyses (gene level and exon level) of raw data files carried out in Expression Console software using iterPLIER algorithms, we identified genes and exons that were differentially regulated in each pairwise comparison considered. The exons identified in this study were examinated for their biological association to gene ontology (GO) categories using eGOn software. Moreover, all exons differentially expressed were also uploaded into Ingenuity Pathways Analysis (Ingenuity® Systems, www.ingenuity.com) in order to identify molecular pathways and functions related to stress and escitalopram response. Our results suggest that stress may exert a negative effect on gene transcription since the largest number of genes was downregulated. Moreover from our data it seems that a different pattern of gene expression exhibits between animals that respond and that did not respond to escitalopram treatment. Functional analysis of exon dataset, arising from stress protocol and escitalopram treatment, reflects interesting different biological features. More specifically, the biological functions regard both molecular and cellular functions, such as cellular growth and proliferation, gene expression and signal transduction, as well as involvement of central neurotransmission and immune response. We believe that this pharmacogenomic approach will be helpful to understand the molecular mechanisms involved in the pathogenesis of depression as well as in the response to antidepressant drugs.


2008 - Impaired stress-induced regulation of brain derived neurotrophic factor expression in hippocampus of glucocorticoid receptor impaired mice: Model of depression [Abstract in Rivista]
Alboni, Silvia; Corsini, Daniela; Caggia, Federica; Benatti, Cristina; Capone, Giacomo; Barden, N; Blom, Johanna Maria Catharina; Tascedda, Fabio; Brunello, Nicoletta
abstract

Objective: The gene codifying for the neurotrophin Brain Derived Nurotrophic Factor (BDNF) is a stress-responsive gene and alteration in its expression may be important in producing some of the pathophysiological effects of stress in the hippocampus as seen in stress-related pathologies like depression. While the effects of stress procedures on the regulation of BDNF expression was widely investigated in hippocampus of healthy control animals, the stress-induced effects on BDNF hippocampal expression in “pathological” condition are still lacking. In order to deepen our knowledge in the understanding of the effects of an acute stressful procedure on molecular targets of synaptic plasticity we used transgenic mice with impaired glucocorticoid receptor (GR-i) expression that represent an animal model of depression. The hypothesis was tested that a single period of restraint stress (6 hours) affects BDNF mRNA expression in the hippocampus of GR-i mice differently than in wild-type (WT) mice. Methods: Using real time RT-PCR we evaluated the effects of a 6 hours acute stress on the levels of BDNF coding exon VIII and the activity regulated BDNF exon IV mRNA. In the WT and in the GR-i animals, the hippocampal levels of the two BDNF exons, immediately after the stress ended, were significantly lower in stressed animals with the respect to respective control unstressed animals. However, for the BDNF exon IV mRNA the reduction is most pronounced inWT animals and two-way ANOVA followed by Bonferroni posttest revealed a significant interaction between stress response and genotype at the level of BDNF exon IV mRNA expression. Results: The BDNF gene is a very complex gene regulated by a wide array of stimuli and signalling pathways. An electophoresis mobility shift assay (EMSA) was used to study DNA-binding activity of two transcription factors with an important role in controlling synaptic plasticity most likely trough an involvement BDNF: the cAMP responsive element binding (CREB) protein and the nuclear factor kB (Nf-kB). Taken together, our results show a different binding activity of these transcription factors in GR-i mice with respect to WT mice following acute stressful conditions. Conclusion: The identification of the molecular mechanisms activated by stress in GR-i mice model of depression may contribute to the development of new strategies that reducing neuron vulnerability to stress and prevent neurophatologocal alteration in the hippocampus.


2008 - Microarray analysis of the chronic escape deficit model of depression: Effects of escitalopram treatment in hippocampus [Abstract in Rivista]
Caggia, Federica; Valensisi, Cristina; Alboni, Silvia; Benatti, Cristina; Corsini, Daniela; Ferrari, F; Tagliafico, Enrico; Mendlewicz, J; Brunello, Nicoletta; Tascedda, Fabio
abstract

Objective: Currently, the biological bases of depression and the molecular mechanisms underlying antidepressant action are not completely understood. Behavioural models of depression and genome-wide gene expression analysis can be relevant to better understand the pathophysiology of this disease. Chronic escape deficit is a valid and useful model of depression and is based on the induction of an escape deficit after exposure of rats to unavoidable stress. This behavioural model allows to evaluate the capacity of a treatment to revert the escape deficit. The majority of antidepressant drugs need to be administered for at least 3−4 weeks in order to revert the escape deficit. In this study, we demonstrated that only one week of treatment with Escitalopram, a widely used SSRI, is effective in the chronic escape deficit model of depression. Also, our study demonstrated that only 50% of the animals receiving ESC responded to the treatment. The mechanisms underlying the action of escitalopram are still poorly understood and the molecular targets and pathways involved remain to be identified. In order to identify the biological target involved in the response to escitalopram, we performed a microarray experiment using the chronic escape deficit model of depression after a 7 day treatment with escitalopram. Methods: Gene expression patterns in the rat hippocampus were analyzed using Affymetrix GeneChip Rat Exon 1.0 ST evaluating both gene-level and exon-level expression profiling on the whole genome. Total RNA extracted from hippocampus of each treated animal was utilized to chipping a single array using the Affymetrix protocols. 20 single arrays were utilized for data analysis and divided into five replicates for each experimental group (naive, stress, escitalopram responders and not responders). With two parallel analyses (gene level and exon level) of raw data files carried out in Expression Console software using iterPLIER algorithms, we identified various transcripts that were differentially regulated in each pairwise comparison. In order to identify biological processes and signalling networks regulated by escitalopram response, we performed a functional analysis using Ingenuity web tool. Results: Functional annotation of selected genes reflected interesting different biological features between escitalopram responders and not responders. More specifically, the biological functions regard cellular growth and proliferation, gene expression and signal transduction. Conclusion: We believe that this pharmacogenomic approach will be helpful to understand the molecular mechanisms involved in the pathogenesis of depression as well as in the response to antidepressant drugs.


2008 - Molecular effects of subchronic andchronic treatment with escitalopram inthe rat central nervous system [Abstract in Rivista]
Benatti, Cristina; Alboni, Silvia; Capone, Giacomo; Corsini, Daniela; Caggia, Federica; Blom, Johanna Maria Catharina; Tascedda, Fabio; Brunello, Nicoletta
abstract

A clear understanding of the mechanisms behind depressionand its treatment is a critical issue for amelioratethe effectiveness of existing antidepressants. Acutely,antidepressant drugs increase synaptic concentrations ofmonoamines, but clinical efficacy requires several weeksof continuous treatment, proposing a key role for timedependentneural adaptations, perhaps induced by acutesynaptic actions, in their therapeutic efficacy.Escitalopram is the S(+) enantiomer of citalopram, oneof the most widely prescribed serotonin selective reuptakeinhibitor (SSRIs) antidepressants. In the chronic mildstress model of depression sucrose intake was alreadynormalized after one week of treatment.We evaluated the effects of a subchronic or a chronictreatment with escitalopram on expression levels of someof the main targets of antidepressant drugs such as theneurotrophin Brain Derived Neurotrophic Factor (BDNF),the transcription factors cAMP Response Element Binding(CREB) [1] Protein and Calcium Responsive Factor(CaRF).Sprague-Dawley rats were treated for 7 days (subchronic)or 21 days (chronic) with either escitalopram(10 mg/kg die i.p) or saline (1 mL/kg die); BDNF, CREBand CaRF mRNAs were evaluated using RNAse ProtectionAssay in hippocampus and prefrontal cortex.No difference was observed on BDNF, CREB andCaRF expression in the hippocampus of rats treatedsubchronically with escitalopram with respect to the grouptreated with saline. In contrast a significant inductionof BDNF mRNA was observed in prefrontal cortexof escitalopram-treated animals with respect to salinetreated ones. CaRF expression patterns were similar.Escitalopram for 7 days caused a significant induction ofCaRF mRNA with respect to the group treated with saline(p<0.05; Dunnett t-test), on the other hand CREB mRNAremained unaffected. Following a chronic treatment withEscitalopram, BDNF, CREB and CaRF mRNA levels weresignificantly decreased with respect to the group treatedwith saline in hippocampus (p<0.05; Dunnett t-test),while a 21 day treatment with escitalopram failed toproduce changes in gene expression in prefrontal cortex.These results showed that escitalopram is able todifferentially affect BDNF, CREB and CaRF expressionwith respect to treatment duration and that the observedeffects are area-specific.Consequently, to further investigate the possiblemolecular mechanisms underlying the observed effectson gene expression we evaluated by western blottingsome of the main signalling pathways regulating CREB aswell as BDNF expression, such as p38 MAPK (Mitogen-Activated Protein Kinase), CaMKII (Calcium CalmodulineKinase), ERK 1/2 (Extracellular Signal-Regulated Kinase)and CREB itself [1].Our study demonstrates that:1. A subchronic treatment with escitalopram inducesBDNF and CaRF expression in prefrontal cortexprobably through activation of p38 MAPK signallingpathway.2. A 21 day escitalopram treatment reduces hippocampalBDNF, CaRF, CREB gene expression and also CREBphosphorylated nuclear levels.Spatially distinct signalling pathways may be involvedin mediating the differential effect on gene expressionobserved following either a subchronic or a chronictreatment with escitalopram.


2007 - Combined effect of antidepressant and anti-inflammatory drugs in an animal model of depressio [Abstract in Rivista]
Brunello, Nicoletta; Alboni, Silvia; Benatti, Cristina; Corsini, Daniela; Capone, Giacomo; Tascedda, Fabio; J., Mendlewicz
abstract

The delay in the onset of action of antidepressants remains one of the unsolved issues in the treatment of depression despite the availability of safe and effective drugs. It has been known for years that depression may share identical symptoms as those in inflammatory reaction and that immune function and inflammation markers are altered in psychiatric patients. This suggests to use anti-inflammatory drugs as an adjunctive therapy for depression. We previously demonstrated that the combination of acetylsalicylic acid (ASA) with Fluoxetine (FLX) accelerated and potentiated the effect of the antidepressant in the chronic escape model of depression (Brunello et al., 2006). These results, together with preliminary clinical data in major depressed nonresponder patients, suggest that ASA might accelerate the onset of action of SSRIs (Mendlewicz et al., 2006). Preclinical and clinical studies have recently reported that Escitalopram (ESC), the active enantiomer of citalopram, shows a faster onset of action compared to other antidepressants. The aim of our study was to compare the effect of one week of combined treatment with ESC plus ASA vs. ESC alone in the chronic escape deficit model of depression. Significant response after one week of treatment was present in about 50% of the animals receiving ESC (10 mg/kg/day) alone and in about 75% of the rats receiving ESC plus ASA (45 mg/kg/day). These results suggest that the co-administration of ASA with ESC increased the response to treatment in reverting the behavioural despair induced by stress in rats, thus confirming the specific effect of combined therapy.


2007 - Farmacologia e neurobiologia nei disturbi d'ansia e nella depressione [Articolo su rivista]
Brunello, Nicoletta; Benatti, Cristina
abstract


2006 - Acetylsalicylic acid accelerates the antidepressant effect of fluoxetine in a rat model of depression [Abstract in Rivista]
Capone, Giacomo; Alboni, Silvia; Benatti, Cristina; Tascedda, Fabio; Blom, Johanna Maria Catharina; J., Mendlewicz; Brunello, Nicoletta
abstract

Depression currently ranks fourth among the major causesof disability worldwide and by 2020, it is estimated thatunipolar major depression will rank second as a sourceof lost disability-adjusted life years (DALYs) worldwide(Murray and Lopez, 1997). To date however no singleagent is effective in all patients treated, probably due tothe different neurobiological alterations occurring for thedisorder and to individual differences in pharmacogeneticand pharmacodynamic parameters.Different therapies have been proposed to amelioratethe clinical responses of antidepressant drugs throughaugmentation or combination strategies.Another achievement in the development of newtreatments is to reduce the latency of clinical effect ofantidepressant drugs known to be characterized by 4-6weeks lag phase.Evidence has accumulated suggesting that majordepression is associated with dysfunction of inflammatorymediators and that psychiatric symptoms may occur duringinflammatory diseases. Moreover antidepressants show ananti-inflanmlatory action possibly related to their clinicalefficacy. In fact, an improvement in psychiatric symptomshas been recently reported in patients treated with antiinflammatorydrugs for other indications.These data imply that inflammation may be involved inthe pathogenesis of depression and that anti-inflanm~atorydrugs may be used as an adjunctive therapy.Among anti-inflammatory drugs acetylsalicylic acid(ASA) has been shown to act on the brain serotonergicsystem and to have a neuroprotective effect in vitro aswell as in vivo.Aim of the present study was to evaluate the effectsof combined treatment fluoxetine (FLX) plus ASA andASA alone in a behavioural model of depression:the chronic escape deficit (Gambarana et al., 2001).The chronic escape deficit model is based on themodified reactivity of rats to external stimuli inducedby exposure to unavoidable stress and allows evaluatingthe capacity of a treatment to revert the condition ofescape deficit. In this model, FLX alone needs to beadministered for at least 3 weeks in order to revert thiscondition.Our results showed that ASA (45 mg/kg) did not possessantidepressant properties in the chronic escape deficitmodel at any time tested. A combined treatment of FLX(5 mg/kg) and ASA (45 mg/kg) completely reverted thecondition of escape deficit as early as after 7 days, theeffect being already partially present after 4 days. Theeffect was maintained after 14 and 21 days of treatment.In the same experimental condition the effect of FLX(5 mg/kg) was significant only at 21 days, as previouslydemonstrated by other groups.The exact nature of the mechanisms underlying theabove behavioural effects are still unknown, neverthelessseveral hypotheses can be formulated. Further biochemicaland genetic researches could help to clarify the targets ofaction of the combined treatment FLX plus ASA for thedevelopment of more active and faster acting molecules.


2006 - Acetylsalicylic acid accelerates the antidepressant effect of fluoxetine in the chronic escape deficit model of depression [Articolo su rivista]
Brunello, Nicoletta; Alboni, Silvia; Capone, Giacomo; Benatti, Cristina; Blom, Johanna Maria Catharina; Tascedda, Fabio; Kriwin, P; Mendlewicz, J.
abstract

Evidence has accumulated suggesting that major depression is associated with dysfunction of inflammatory mediators. Moreover, antidepressants show an antiinflammatory action possibly related to their clinical efficacy. An improvement in psychiatric symptoms has been recently reported in patients treated with antiinflammatory drugs for other indications. These data imply that inflammation may be involved in the pathogenesis of depression and that anti-inflammatory drugs may be used as an adjunctive therapy. The aim of the present study was to evaluate the behavioural effect of the co-administration of acetylsalicylic acid (ASA, 45 mg/kg or 22.5 mg/kg) and fluoxetine (FLX, 5 mg/kg) in the chronic escape deficit model of depression. The chronic escape deficit model is based on the modified reactivity of rats to external stimuli induced by exposure to unavoidable stress and allows evaluation of the capacity of a treatment to revert the condition of escape deficit. In this model, FLX alone needs to be administered for at least 3 weeks to revert this condition. Our results show that combined treatment of fluoxetine and ASA completely reverted the condition of escape deficit by as early as 7 days, the effect being already partially present after 4 days. The effect was maintained after 14 and 21 days of treatment. ASA alone was ineffective at any time tested and the effect of fluoxetine was significant only at 21 days. These results, together with clinical data from preliminary results, suggest that ASA might accelerate the onset of action of selective serotonin reuptake inhibitor antidepressants.


2006 - Behavioural and molecular effects of the combined treatment fluoxetine plus acetylsalicylic acid in a rat model of depression [Abstract in Rivista]
Alboni, Silvia; Capone, Giacomo; Benatti, Cristina; Tascedda, Fabio; Blom, Johanna Maria Catharina; Mendlewicz, J; Brunello, Nicoletta
abstract

Current treatments for depression are inadequate for many patients, and different therapies have been proposed to ameliorate the clinical responses of antidepressant drugs through augumentation or combination strategies. Another achievement in the development of new treatments is to reduce the latency of clinical effect of antidepressant drugs known to be characterized by 4−6 weeks lag phase. Evidence has accumulated suggesting that major depression is associated with dysfunction of inflammatory mediators and that psychiatric symptoms may occur during inflammatory diseases. Moreover antidepressants show an anti-inflammatory action possibly related to their clinical efficacy. In fact, an improvement in psychiatric symptoms has been recently reported in patients treated with anti-inflammatory drugs for other indications. These data imply that inflammation may be involved in the pathogenesis of depression and that anti-inflammatory drugs may be used as an adjunctive therapy. Among the anti-inflammatory drugs the Acetylsalicylic acid has been shown to act on the brain serotonergic system and to have a neuroprotective effect toward brain damage. Aim of the present study was to evaluate the effects of combined treatment fluoxetine (FLX) plus acetylsalicylic acid (ASA) in a behavioural model of depression: the chronic escape deficit. The chronic escape deficit model is based on the modified reactivity of rats to external stimuli induced by exposure to unavoidable stress and allows evaluating the capacity of a treatment to revert the condition of escape deficit. In this model, FLX alone needs to be administered for at least 3 weeks in order to revert this condition. Our results showed that ASA (45 mg/kg) did not possess antidepressant properties in the chronic escape deficit model at any time tested. A combined treatment of FLX (5 mg/kg) and ASA (45 mg/kg) completely reverted the condition of escape deficit as early as after 7 days, the effect being already partially present after 4 days. The effect was maintained after 14 and 21 days of treatment. In the same experimental condition the effect of FLX (5 mg/kg) was significant only at 21 days, as previously demonstrated by other groups. The exact nature of the mechanisms underlying the above behavioural effects are still unknown, nevertheless several hypotheses can be formulated. Moreover, because a role for the neurotrophin BDNF was proposed in the clinical response to antidepressant treatment, we have determined the effect of combined treatment FLX plus ASA on hippocampal BDNF mRNA and protein in the same behavioural model. Ours data demonstrated that the hippocampal levels of BDNF mRNA were significantly increased with respect to control groups (naive and stressed) only in the animals responding (number of escape 10 out of 30 trials) to the combined treatment. Further biochemical and genetic researches could help to clarify the targets of action of the combined treatment FLX plus ASA for the development of more active and faster molecules.


2006 - Early postnatal chronic inflammation produces long-term changes in pain behavior and N-methyl-D-aspartate receptor subtype gene expression in the central nervous system of adult mice [Articolo su rivista]
Blom, Johanna Maria Catharina; Benatti, Cristina; Alboni, Silvia; Capone, Giacomo; Ferraguti, Chiara; Brunello, Nicoletta; Tascedda, Fabio
abstract

The objective of this study was to test whether postnatal chronic inflammation resulted in altered reactivity to pain later in life when reexposed to the same inflammatory agent and whether this alteration correlated with brain-region-specific patterns of N-methyl-D-aspartate (NMDA) receptor subtype gene expression. Neonatal mouse pups received a single injection of complete Freund's adjuvant (CFA) or saline into the left hind paw on postnatal day 1 or 14. At 12 weeks of age, both neonatal CFA- and saline-treated animals received a unilateral injection of CFA in the left hind paw. Adult behavioral responsiveness of the left paw to a radiant heat source was determined in mice treated neonatally with saline or CFA before and after receiving CFA as adults. Twenty-four hours later, brains were dissected and NMDA receptor subunit gene expression was determined in four different brain areas by using an RNase protection assay. The results indicated that NMDA receptor subtype gene expression in adult mice exposed to persistent neonatal peripheral inflammation was brain region specific and that NMDA gene expression and pain reactivity differed according to the day of neonatal CFA exposure. Similarly, adult behavioral responsiveness to a noxious radiant heat source differed according to the age of neonatal exposure to CFA. The data suggest a possible molecular basis for the hypothesis that chronic persistent inflammation experienced early during development may permanently alter the future behavior and the sensitivity to pain later in life, especially in response to subsequent or recurrent inflammatory events


2006 - Effects of acute stress on brain-derived neurotrophic factor in the hippocampus of transgenic mouse model of depression [Abstract in Rivista]
Alboni, Silvia; Blom, Johanna Maria Catharina; Corsini, Daniela; Benatti, Cristina; Capone, Giacomo; Ferraguti, Chiara; N., Barden; Tascedda, Fabio; Brunello, Nicoletta
abstract

Brain-Derived Neurotrophic Factor (BDNF) is a member of the neurotrophin family which includes a group of molecules important for the development and the maintenance of the nervous system. Since BDNF is highly expressed in the hippocampus, the action and regulation of this neurotrophin in this area has become subject of intense study. The gene codifying for BDNF is a stress responsive gene and alterations in its expression may be important in regulating some of the physiological and pathophysiological effects of chronic and acute stress in the hippocampus. Different studies show that several types of stress reduce BDNF expression in the hippocampus of control animals [Smith et al., 1995] and these works led to a neurotrophic hypothesis of depression [Nestler et al., 2002]. Nevertheless, the effect of stress on BDNF gene expression may differ between a "normal" and a "pathological" brain. In our study, we used transgenic mice with glucocorticoid receptor impaired (GRi) expression created [Pepin et al., 1992], as a tool to study the neuroendocrine changes observed in stress-related disorders, such as major depression. This GRi mouse model is characterized by dysfunctional glucocorticoid inhibitory feedback and an excessive activation of the hypothalamic pituitar~adrenal (HPA) axis, that can be restored by antidepressant drugs' treatment. The hypothesis was tested that a single period of 30 minutes of restraint stress affects BDNF expression in the hippocampus of GRi mice differently than in wildtype (WT) mice. Using RNase protection assay and in situ hybridization we had assessed the BDNF mRNA hippocampal levels, while the levels of BDNF and its precursor, pro-BDNF were analyzed by western blotting. Our results indicated that 30 minutes of restraint enhanced BDNF mRNA expression in the CA3 hippocampal subregion of GRi mice; the same stress procedure induced also a statistically significant increase of pro- BDNF level in hippocampus of GRi mice. No effect of acute stress was observed in the WT at the level of the expression of BDNE Moreover, we evaluated the effects of restraint on signalling pathways implicated in the regulation of BDNF expression (mitogen-activated protein kinase and calcium/calmodulin-dependent kinase cascades) that converge on the phosphorylation of CREB that we found down-regulated in the hippocampus of GRi mice and up-regulated in WT mice. Our data suggest that, in the presence of psychophysiological stress (restraint stress), GRi mice display altered hippocampal regulation in BDNF gene expression. Thus, life-long central GR dysfunction may negatively affects neural functioning by limiting the capacity to cope with change or acute stress, which could be a predisposing or determining factor in depression. Understanding the mechanisms underlying the induction of BDNF mRNA and accumulation of pro-BDNF in the hippocampus of GRi mice, may help to clarify the molecular basis of action of this neurotrophin and contribute to the development of new strategies reducing the vulnerability of neurons to stress, thus preventing neuropathological alterations in the hippocampus.


2006 - Shortened onset of action of antidepressants in major depression using acetylsalicylic acid augmentation: a pilot open-label study [Articolo su rivista]
Mendlewicz, J; Kriwin, P; Oswald, P; Souery, D; Alboni, Silvia; Brunello, Nicoletta
abstract

Based on our preclinical data showing a potential accelerating effect of acetylsalicylic acid (ASA) in combination with fluoxetine in an animal model of depression, we examined the effect of ASA augmentation therapy on selective reuptake inhibitors (SSRI) in major depressed non-responder patients. Twenty-four non-responder patients having received at least 4 weeks of an adequate SSRI treatment were included in a pilot open-label study. Participants were treated openly during 4 weeks with 160 mg/day ASA in addition to their current antidepressant treatment. The combination SSRI-ASA was associated with a response rate of 52.4%. Remission was achieved in 43% of the total sample and 82% of the responder sample. In the responder group, a significant improvement was observed within week 1 (mean Hamilton Depression Rating Scale-21 items at day 0 = 29.3 +/- 4.5, at day 7 = 14.0 +/- 4.1; P < 0.0001) and remained sustained until day 28. Despite limitations due to the open nature of this study, our preliminary results confirm our preclinical findings and are in favour of an accelerating effect of ASA in combination with SSRIs in the treatment of major depression. Potential physiological and biochemical mechanisms may involve an anti-inflammatory and/or neurotrophic effect.


2005 - Neonatal persistent inflammation alters pain response and NMDA receptor expression in adult mice [Abstract in Rivista]
Benatti, Cristina; Alboni, Silvia; Ferraguti, Chiara; Tascedda, Fabio; Blom, Johanna Maria Catharina; Brunello, Nicoletta
abstract

Infant pain is of critical interest, especially with respect to premature infants and other high-risk neonates that experience many invasive and traumatic procedures early in development. The early neonatal period is characterized by great plasticity and reorganization. Sustained activation of central nervous circuits, caused by protracted and recurrent pain, may cause long-lasting changes in central neural function thus affecting developmental outcome and behavioural responsiveness to pain or stress later in life. However little is known about the neurobiological substrates underlying this ``memory'' process. The aim of our study was twofold:to study whether timing of postnatal exposure to a persistent inflammatory insult alters the responsiveness to thermal pain in the adult animal;given the role of the NMDA receptor in pain processing as well as in learning and memory, to examine if NMDA receptor subtype gene expression in specific areas of the cns is influenced by neonatal inflammation.Methods: Newborn mice received a single injection of Complete Freund's Adjuvant (CFA) or saline on either postnatal day 1, 3 or 14 (P1, P3 and P14) into the left hind paw. At twelve weeks of age paw withdrawal latency (PWL) of each animal was tested both in basal condition and 24h after an unilateral injection of 100 μL of CFA in the left hind paw. Mice were then killed by cervical dislocation and cerebral areas were removed. Using a sensitive RNAse protection assay, NMDA receptor subunit (NR1, NR2A, NR2B, NR2C) gene expression was evaluated in different brain areas; all data were processed by one-way ANOVA (p < 0.05).Results: Baseline paw withdrawal latency was significantly decreased in animals exposed to CFA at day 1 and 14 as compared to their saline exposed counterparts. Animals exposed to CFA at postnatal day 3 showed a significant increase in paw withdrawal latency with respect to saline injected animals. Twenty-four hours later a unilateral injection of CFA into the left hind paw, a significant decrease in paw withdrawal latency was observed in all experimental groups with respect to baseline values. PWL of P1 saline treated animals after CFA exposure was significantly higher than P3 and P14 saline treated mice. Adult mice exposed to an injection with CFA on postnatal day 1 exhibited reduced expression of the NMDA receptor subtype NR1 and NR2C in the hippocampus while mRNA levels for NR2A and NR2B did not differ between CFA treated and untreated mice. Exposure to CFA on postnatal day 3 and 14 did not affect adult expression levels of NMDA receptor subunits in the hippocampus. NMDA receptor subunit expression displayed a different profile in the thalamus. Exposure to CFA at P1 and P3 did not alter NMDA receptor subunit expression while exposure to CFA at P14 resulted in enhanced expression of the NR2A and NR2B subunits.Conclusions: These findings indicate that changes in NMDA receptor subtype gene expression in adult mice exposed to persistent neonatal peripheral inflammation are brain region specific and that NMDA gene expression and pain reactivity differ according to the day of neonatal exposure to CFA.


2005 - New combination therapies from animal to human [Abstract in Rivista]
Brunello, Nicoletta; Alboni, Silvia; Benatti, Cristina; Capone, Giacomo; Tascedda, Fabio; Blom, Johanna Maria Catharina; J., Mendlewicz
abstract

Evidence has accumulated suggesting that major depression is associated with dysfunction of inflammatory mediators. Antidepressants interfere with the synthesis and release of cytokines and do not exert behavioral effects in animal models of depression when hippocampal neurogenesis is blocked, a phenomenon which is occurring in the presence of inflammation. The anti-inflammatory drug acetylsalicylic acid (ASA), besides inhibiting the cyclooxigenase pathway, interacts with central serotonergic system, by increasing serotonin levels in cortex and reducing the density of different serotonin receptor subtypes. These neurochemical effects suggest a role of ASA in the treatment of depression. Therefore we studied the effect of ASA and fluoxetine combined treatment in a behavioral model of depression. The chronic escape deficit model is based on the modified reactivity of rats to external stimuli induced by exposure to unavoidable stress and allows evaluating the capacity of a treatment to revert the condition of escape deficit. Any kind of antidepressant drug needs to be administered for at least 3 weeks in order to revert this condition. The combined treatment of fluoxetine and ASA completely reverted the condition of escape deficit as early as after 7 days, the effect being already partially present after 4 days. The effect was maintained after 14 and 21 days of treatment. ASA alone was ineffective at any time tested and the effect of fluoxetine was significant only at 21 days. Given these preclinical results, an open clinical study has been started using the combination SSRI-ASA in treatment resistant depressed patients. Preliminary results suggest a potential accelerating effect of ASA in combination to SSRI.


2004 - Chronic treatment with desipramine and fluoxetine modulate BDNF, CaMKK alpha and CaMKK beta mRNA levels in the hippocampus of transgenic mice expressing antisense RNA against the glucocorticoid receptor [Articolo su rivista]
J., Vinet; Carra, Serena; Blom, Johanna Maria Catharina; Brunello, Nicoletta; N., Barden; Tascedda, Fabio
abstract

Antidepressants up-regulate the cAMP response element binding protein (CREB) and the brain-derived neurotrophic factor (BDNF) in hippocampus and these effects contribute to the protection of hippocampal neurons from stressful stimuli such as high glucocorticoid levels. CREB can be activated by both protein kinase A and by Ca2+-calmodulin-dependent protein kinases (CaMKs), which are in turn phosphorylated by their upstream activators CaMKKalpha and CaMMKKbeta. Using in situ hybridization, we examined the effects of chronic treatment with fluoxetine (FLU) or desipramine (DMI) on BDNF, CaMKKalpha and CaMKKbeta mRNAs in the hippocampus of wild-type (Wt) and transgenic (TG) mice characterized by glucocorticoid receptor (GR) dysfunction. Basal levels of CaMKKbeta were down regulated in the CA3 region of TG mice. DMI decreased the expression of both CaMKKalpha and CaMMKKbeta in the CA3 region of Wt mice. FLU up-regulated BDNF mRNA levels in the CA3 of TG animals while both FLU and DMI increased BDNF gene expression in the dentate gyrus (DG) of TG animals. Our results demonstrate a different regulation of BDNF expression by antidepressant drugs in the hippocampus of Wt and TG animals. Moreover, for the first time, a role for CaMKKs in the mechanism of action of antidepressant agents, at least in the hippocampus, is reported. These data are discussed in view of interactions existing between CaMK pathway and GR-mediated gene transcription.


2004 - Escitalopram: meccanismo d’azione e profili clinico [Articolo su rivista]
Brunello, Nicoletta; Cannazza, Giuseppe; Auguglia, E.
abstract

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2004 - Mood stabilizers: protecting the mood ... protecting the brain [Articolo su rivista]
Brunello, Nicoletta
abstract

The mechanism underlying the therapeutic action of mood stabilizers in bipolar disorder is not completely understood. Thediscovery that anticonvulsant agents, such as valproate (VPA), were effective in the treatment of bipolar disorder suggested acommon biochemical mechanism(s) with lithium. Recent research has focused on how VPA and lithium change the activities ofcellular signal transduction systems, especially the cyclic AMP and phosphoinositide second messenger pathways. Despitebeing structurally dissimilar, VPA produces effects on the protein kinase C (PKC) signalling pathway that are similar to lithium,although the VPA effects appear to be largely independent of myo-inositol. Furthermore, the therapeutic benefit of either drugrequire a prolonged administration suggesting alterations at the genomic level. Studies have revealed that both VPA and lithiumaltered the expression of several early inducible genes belonging to the AP-1 family of transcription factors; this family isresponsible for controlling the expression of a number of genes including cytoprotective proteins such as the anti-apoptoticprotein, bcl-2. Evidence shows that chronic administration of VPA or lithium can stimulate bcl-2 expression as well as inhibitGSK-3h activity, which renders a cell less susceptible to apoptosis. Thus, the mood stabilizers may act to restore the balanceamong aberrant signalling pathways in specific areas of the brain and prevent degeneration


2004 - Regulation of CREB function in rat frontal cortex after combined treatment with Fluoxetine and Olanzapine [Abstract in Rivista]
Capone, Giacomo; Alboni, Silvia; Blom, Johanna Maria Catharina; Ferraguti, Chiara; Brunello, Nicoletta; Tascedda, Fabio
abstract

Statement of the Study: Generally, drugs used in the treatment of depression exert their therapeutic effect after 4/6 weeks and only in 60–65% of patients. The search for an adequate and faster treatment of major depression is one of the main challenges in neuropsychopharmacology. Recently, a clinical study of treatmentresistant depressed patients without a psychotic component, showed that after only one week of treatment, Fluoxetine (a selective serotonin reuptake inhibitor antidepressant) and Olanzapine (an atypical antipsychotic agent) produced a higher level of improvement than either monotherapy alone (Shelton et al., American Journal of Psychiatry 158(1), 131–134, 2001). Furthermore, preclinical data, using microdialysis, indicated that the combination of Olanzapine and Fluoxetine resulted in an increase in the extracellular levels of dopamine and norepinephrine in the rat prefrontal cortex, an effect that was significantly bigger than after treatment with either drug alone (Zhang et al., Neuropsychopharmacology 23(3),250– 262, 2000). However, it is not yet completely understood which intracellular signaling pathway could be involved in the fast response (seen in clinical trials) to Fluoxetine plus Olanzapine co-treatment. Methods: Since antidepressant and antipsychotic drugs affect the cyclic adenosine monophosphate (cAMP) pathway, including the expression of the cAMP response element binding protein (CREB), the levels of CREB mRNA and CREB nuclear protein, total and phosphorylated, were studied in the frontal cortex of rats using RNase protection assay and Western Blotting analysis respectively. Four experimental groups were used: rats were treated for one, five or ten days with either saline, Fluoxetine (ip 10 mg/Kg), Olanzapine (sc 1 mg/Kg) or combined Fluoxetine plus Olanzapine (10 mg/Kg and 1 mg/Kg). Summary of Results: Our results show that the level of phosphorylated CREB Ser133 was significantly increased in the frontal cortex of rats receiving the combined treatment regimen (Fluoxetine plus Olanzapine) for five days. No effect was observed in acutely and ten day treated rats. Conclusion: This specific effect on CREB phosphorylation levels after five days of combined treatment with Fluoxetine plus Olanzapine might represent one of the mechanisms underlying the faster response to this therapy recently observed in several clinical trials.


2004 - Restraint stress increases the expression of brain derived neurotrophic factor in the hippocampus of a mouse model of depression [Abstract in Rivista]
Alboni, Silvia; Benatti, Cristina; Blom, Johanna Maria Catharina; Ferraguti, Chiara; Tascedda, Fabio; Barden, N; Brunello, Nicoletta
abstract

Statement of the study: Brain-Derived Neurotrophic Factor (BDNF) is a member of the neurotrophin family which includes a group of molecules important for the development and the maintenance of the nervous system. Since BDNF is highly expressed in the hippocampus, the action and regulation of BDNF in this particular area has become subject of intense study. Single or repeated immobilization stress markedly reduces both BDNF mRNA and protein levels in rat hippocampus. Consequently, BDNF is considered a stress-responsive gene, and it has been recently suggested that alterations in the expression of this growth factor may be important in regulating some of the physiological and pathophysiological effects of stress on the hippocampus. Stress-induced changes observed in the hippocampus of experimental animals resulted in a novel hypothesis attributing a central role to neurotrophic factors in both the etiology of depression and as well as in its treatment. However, the effects of stress on neurotrophic factors in the hippocampus of depressed patients remain unknown. In fact, the expression pattern of a large array of genes affected by depression or antidepressant drugs, such as BDNF, may differ between a normal and a pathological brain. Methods: In these experiments, we used transgenic (TG) mice deficient in glucocorticoid receptor (GR) functioning. This TG mouse was created as a model to study the neuroendocrine changes occurring in stress-related disorders, such as major depression. We evaluated the hypothesis that a single period of 30 minutes of restraint stress affects BDNF mRNA expression in the hippocampus of TG mice differently than in WT mice. Summary of results: BDNF mRNA was significantly increased by the stress procedure only in the hippocampus of TG mice, the induction being specific for the CA3 subregion as revealed by in situ hybridization. Moreover, we found that stress down-regulated CREB phosphorylation in the hippocampus of TG mice whereas it upregulated the level of phosphorylated CREB Ser133 in WT mice. Conclusion: These data suggest that, in the presence of emotional stress, lifelong central glucocorticoid receptor dysfunction results in altered hippocampal sensitivity, with respect to the level of neurotrophic gene expression. The understanding of the mechanisms through which psychological stress (such as restraint stress) induces BDNF mRNA in the hippocampus of TG mice, may help to clarify the biological and molecular basis of the action of neurotrophic factors and may contribute to the development of new strategies that will ultimately reduce the vulnerability of neurons and prevent neuropathological alterations in the hippocampus.


2004 - Serotonin-enhancing drugs [Articolo su rivista]
Brunello, N.
abstract

Through its projections to various brain areas, serotonin participates to the control of numerous functions, like sleep, mood regulation, fear and anxiety, aggressiveness, motivation and reward, learning and memory, hunger, sexual activity, circadian rhythm regulation, neuroendocrine regulation, stress response and pain sensitivity. The multiplicity of the effects mediated by serotonin derives from the interaction of this neurotransmitter with a wide variety of membrane receptors localized in both the nervous system (central and peripheral) and peripheral organs. In fact, such receptors were localized also in the heart and other sites of the cardiovascular system, as well as in the gastrointestinal system. Recent genetic and molecular biology studies lead to the identification of at least 15 different serotonin receptor subtypes that ensure enormous variability in the response associated to the serotonergic system. However, despite the elevated number of receptors, serotonin receptor pharmacology is relatively recent, and few are the drugs currently marketed which act at a serotonin receptor level. Hence, the drugs which are better known as serotonergic system modulators are not the agonists or antagonists of the various serotonin receptor subtypes, but the serotonin transporter inhibitors. These are drugs that block neuronal serotonin reuptake, although with different potency and selectivity, and are effective in treating mood, depressive, anxiety, and eating disorders.


2003 - Cellular mechanisms and second messengers: relevance to the psychopharmacology of bipolar disorders [Articolo su rivista]
Brunello, Nicoletta; Tascedda, Fabio
abstract

The discovery of lithium's efficacy as a mood-stabilizing agent revolutionized the treatment of patients with bipolar disorder and after five decades, lithium continues to be the mainstay of treatment for bipolar disorder. Recent research on the molecular mechanism underlying the therapeutic effect of lithium has focused on how it changes the activities of cellular signal transduction systems, especially the cyclic AMP and phosphomositide second-messenger systems. Considerable data suggest that carbamazepine and valproate (VPA) are an alternative or adjunctive treatment to lithium. VPA, despite being dissimilar structurally to lithium, shares most of the effects of lithium at the level of protein kinase C (PKC). Like lithium, VPA reduces the activity of PKC and reduces the protein levels of different PKC isoforms, however the effects of VPA appear to be largely independent of inositol. The ton-term efficacy of VPA and lithium in bipolar disorder suggested that modulation of gene expression might be an important target for these drugs. Both VPA and lithium altered the expression of the early inducible genes for c-fos and cjun thus promoting the expression of specific proteins. The genes known to be regulated by the AP-1 family of transcription factors include genes for various neuropeptides, neurotrophins, receptors, transcription factors, enzymes, proteins that bind to cytoskeletal elements, and cytoprotective proteins such as bcl-2. In conclusion chronic treatment with lithium and other mood stabilizers, by regulating transcriptional factors, may modulate the expression of a variety of genes that compensate for aberrant signalling associated with the pathophysiology of bipolar disorder.


2003 - Chiralità e farmaci antidepressivi [Articolo su rivista]
Brunello, Nicoletta; Cannazza, Giuseppe
abstract

Chiralità e Farmaci antidepressivi


2003 - Cloning of mouse Ca2+/calmodulin-dependent protein kinase kinase beta (CaMKKbeta) and characterization of CaMKKbeta and CaMKKalpha distribution in the adult mouse brain. [Articolo su rivista]
Vinet, Jonathan; Carra, Serena; Blom, Johanna Maria Catharina; Harvey, M; Brunello, Nicoletta; Barden, N; Tascedda, Fabio
abstract

The Ca(2+)/calmodulin-dependent protein kinase kinases alpha and beta (CaMKKs alpha and beta) are novel members of the CaM kinase family. The CaMKKbeta was cloned from mouse brain. The deduced amino acid sequence shared 96.43% homology with the rat CaMKKbeta. Both the alpha and beta isoforms were widely distributed throughout the adult mouse brain. Additionally, all peripheral tissues examined displayed CaMKK alpha and beta expression.


2003 - Noradrenaline in mood and anxiety disorders: Basic and clinical studies [Articolo su rivista]
Brunello, N.; Blier, P.; Judd, L. L.; Mendlewicz, J.; Nelson, C. J.; Souery, D.; Zohar, J.; Racagni, G.
abstract

The neurotransmitter noradrenaline is known to be involved in a range of physiological and psychological processes, and dysfunctions of this neurotransmitter system have been implicated in a range of psychiatric disorders. However, the clinical utility of targeting the noradrenergic system in the treatment of such disorders has been somewhat overshadowed by the availability of agents selective for the serotonin system. A number of antidepressants with increased, but varying, selectivity for the noradrenergic system have become available in recent years, including mirtazapine, bupropion and, most recently, the first truly selective noradrenaline reuptake inhibitor, reboxetine. This review brings together current thinking on the role of noradrenaline in the aetiology and therapy of mood disorders to encourage a rational, evidence-based approach to the treatment of such disorders, and to provide suggestions and guidelines for future research in the area. Int Clin Psychopharmacol 18:191-202


2002 - Altered regulation of CREB by chronic antidepressant administration in the brain of transgenic mice with impaired glucocorticoid receptor function. [Articolo su rivista]
Blom, Johanna Maria Catharina; Tascedda, Fabio; Carra, Serena; Ferraguti, Chiara; Barden, N; Brunello, Nicoletta
abstract

Various effects of antidepressant drugs on gene transcription have been described and altered gene expression has been proposed as being a common biological basis underlying depressive illness. One target for the common action of antidepressants is a modifying effect on the regulation of postreceptor pathways and genes related to the cAMP cascade. Recent studies have demonstrated that long-term antidepressant treatment resulted in sustained activation of the cyclic adenosine 3',5'-monophosphate system and in increased expression of the transcription factor cAMP response element binding protein (CREB). A transgenic animal model of depression with impaired glucocorticoid receptor function was used to investigate the effect of chronic antidepressant treatments on CREB expression in different brain areas. Wild-type and transgenic mice received one administration of saline, desipramine, or fluoxetine, daily for 21 days. The effects of antidepressants on CREB mRNA were analyzed using a sensitive RNase protection assay. Antidepressant treatment resulted in a neuroanatomically and animal specific expression pattern of CREB. Our findings suggest that life-long central glucocorticoid receptor dysfunction results in an altered sensitivity with respect to the effects of antidepressants on the expression of CREB.


2002 - Cognitive deficits and changes in gene expression of NMDA receptors after prenatal methylmercury exposure [Articolo su rivista]
Baraldi, Mario; Zanoli, Paola; Tascedda, Fabio; Blom, Johanna Maria Catharina; Brunello, Nicoletta
abstract

Previous studies showed learning and memory deficit in adult rats that were prenatally exposed to methylmercury chloride (MMC) in an advanced stage of pregnancy (15 days). Under these conditions, the cognitive deficits found at 60 days of age paralleled particularly changes in the N-methyl-D-aspartate (NMDA) receptor characteristics. In the present study, we report the behavioral effects of a single oral dose of MMC (8 mg/kg) administered earlier at gestational day 8. The use of different learning and memory tests (passive avoidance, object recognition, water maze) showed a general cognitive impairment in the in utero-exposed rats tested at 60 days of age compared with matched controls. Considering the importance of the glutamatergic receptor system and its endogenous ligands in learning and memory process regulation, we surmised that MMC could affect the gene expression of NMDA receptor subtypes. The use of a sensitive RNase protection assay allowed the evaluation of gene expression of two families of NMDA receptors (NR-1 and NR-2 subtypes). The result obtained in 60-day-old rats prenatally exposed to MMC, showed increased mRNA levels of the NR-2B subunit in the hippocampus but not in the frontal cortex. The data suggest that the behavioral abnormalities of MMC-exposed rats might be ascribed to a neurotoxic effect of the metal that alters the gene expression of a specific NMDA receptor subunit in the hippocampus.


2002 - The role of noradrenaline and selective noradrenaline reuptake inhibition in depression [Articolo su rivista]
Brunello, Nicoletta; J., Mendlewicz; S., Kasper; B., Leonard; S., Montgomery; Jc, Nelson; E., Paykel; M., Versiani; G., Racagni
abstract

Depression is a common disorder that impacts on all aspects of a person's life. For the past 10 years, clinicians have focused on serotonin in their treatment of depression. This is largely due to the growing acceptance of the efficacy and safety of the selective serotonin reuptake inhibitors (SSRIs) in comparison with older tricyclic antidepressants (TCAs). However, evidence for a role of noradrenaline in depression has been accumulating for some time, beginning with the discovery that drugs which either caused or alleviated depression acted to alter noradrenaline metabolism. Until recently, the role of noradrenaline in depression was predicted from clinical experience with noradrenergic TCAs (desipramine, nortriptyline and protriptyline) and selective serotonin and noradrenaline reuptake inhibitors (venlafaxine, milnacipran). The licensing of reboxetine, a selective noradrenaline reuptake inhibitor now allows the role of noradrenaline in depression to be investigated directly. This review presents key data from the literature that support a role for noradrenaline in depression taking into account neurophysiology, psychopharmacology and clinical trial data. (C) 2002 Elsevier Science B.V./ECNP. All rights reserved.


2001 - Effect of reboxetine treatment on brain cAMP- and calcium/calmodulin-dependent protein kinases [Articolo su rivista]
S., Mori; M., Popoli; Brunello, Nicoletta; G., Racagni; J., Perez
abstract

Previous studies showed that the type II Ca2+/calmodulin- and cAMP-dependent protein kinases (CaMKII and PKA) are affected by long-term antidepressant treatment in presynaptic and somatodendritic compartments, respectively. This study describes the long-term effects of the selective noradrenaline reuptake inhibitor reboxetine on PKA and CaMKII. in both the microtubule and subsynaptosomal fractions of rat brain. Unlike other antidepressants, chronic reboxetine induced in the cerebrocortical soluble and microtubule fractions a decrease in the [P-32]cAMP binding to the type II PKA regulatory subunit. No change in the cAMP-dependent endogenous phosphorylation of the protein substrate, microtubule-associated protein 2 was observed. In the hippocampal subsynaptosomal fractions (synaptic vesicles and synaptosomal membranes) reboxetine induced a robust increase in the activity but not in the expression of CaMKII. An increase in the calcium/calmodulin-dependent phosphorylation of presynaptic substrates was also detected. These findings showed that reboxetine modulates post-receptor signal transduction systems in rat brain. (C) 2001 Elsevier Science Ltd. All rights reserved.


2001 - Modulation of glutamate receptors in response to the novel antipsychotic olanzapine in rats. [Articolo su rivista]
Tascedda, Fabio; Blom, Johanna Maria Catharina; Brunello, Nicoletta; Zolin, K; Gennarelli, Massimo; Colzi, A; Bravi, D; Carra, Serena; Racagni, G; Riva, M. A.
abstract

BACKGROUND:A disturbance in glutamate neurotransmission has been hypothesized in schizophrenia. Hence, the beneficial effects of pharmacological treatment may be related to adaptive changes taking place in this neurotransmitter system.METHODS:In this study, we investigated the modulation of ionotropic and metabotropic glutamate receptors in the rat brain following acute or chronic exposure to the novel antipsychotic olanzapine.RESULTS:In accordance with the clear distinction between classical and atypical drugs, olanzapine did not alter glutamate receptor expression in striatum. Chronic, not acute, exposure to olanzapine was capable of up-regulating hippocampal mRNA levels for GluR-B and GluR-C, two alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (AMPA)-forming subunits. This effect could be relevant for the improvement of schizophrenic alterations, which are thought to depend on dysfunction of the glutamatergic transmission within the hippocampal formation. We also found that the expression of group II glutamate metabotropic receptors was up-regulated in the frontal cortex after chronic exposure to clozapine, and to a lesser extent olanzapine, but not with haloperidol.CONCLUSIONS:The adaptive mechanisms taking place in glutamatergic transmission might prove useful in ameliorating some of the dysfunction observed in the brain of schizophrenic patients.


2001 - Posttraumatic stress disorder: Diagnosis and epidemiology, comorbidity and social consequences, biology and treatment [Articolo su rivista]
Brunello, N.; Davidson, J. R. T.; Deahl, M.; Kessler, R. C.; Mendlewicz, J.; Racagni, G.; Shalev, A. Y.; Zohar, J.
abstract

Epidemiological studies clearly indicate that posttraumatic stress disorder (PTSD) is becoming a major health concern worldwide even if still poorly recognized and not well treated. PTSD commonly co-occurs with other psychiatric disorders, and several symptoms overlap with major depressive disorders, anxiety disorders and substance abuse; this may contribute to diagnostic confusion and underdiagnosis. This anxiety disorder provokes significant occupational, psychiatric, medical and psychosocial disability, and its consequences are enormously costly, not only to the survivors and their families, but also to the health care system and society. Work impairment associated with PTSD is very similar to the amount of work impairment associated with major depression. The pathophysiology of PTSD is multifactorial and involves dysregulation of the serotonergic as well as the noradrenergic system. A rational therapeutic approach should normalize the specific psychobiological alterations associated with PTSD. This can be achieved through the use of antidepressant drugs, mainly of those that potentiate serotonergic mechanisms. Recent double-blind placebo-controlled studies report the efficacy of selective serotonin reuptake inhibitors. Several cognitive-behavioral and psychosocial treatments have also been reported to be efficacious and could be considered when treating PTSD patients. Copyright


2001 - Serine/threonine kinases as molecular targets of antidepressants: Implications for pharmacological treatment and pathophysiology of affective disorders [Articolo su rivista]
Popoli, M.; Mori, S.; Brunello, N.; Perez, J.; Gennarelli, M.; Racagni, G.
abstract

It is currently a widely accepted opinion that adaptive. plastic changes in the molecular and cellular components of neuronal signaling systems correlate with the effects on mood and cognition observed after long-term treatment with antidepressant drugs. Protein phosphorylation represents a key step for most signaling systems, and it is involved in the regulation of virtually all cellular functions. Two serine/threonine kinases. Ca2+/calmodulin-dependent protein kinase II and cyclic AMP-dependent protein kinase. have been shown to be activated in the brain following antidepressant treatment. The changes in kinase activity are mirrored by changes in the phosphorylation of selected protein substrates in subcellular compartments (presynaptic terminals and microtubules), which, in rum, ma), contribute to the modulation of synaptic transmission observed with antidepressants. The molecular consequences of protein kinase activation may account for some of the alterations in neural function induced by antidepressants, and may suggest novel possible strategies of pharmacological intervention. (C) 2001 Elsevier Science Inc. All rights reserved.


2000 - Depression and sleep disorders: clinical relevance, economic burden and pharmacological treatment [Articolo su rivista]
Brunello, Nicoletta; R., Armitage; I., Feinberg; E., Holsboer Trachsler; D., Leger; P., Linkowski; Wb, Mendelson; G., Racagni; B., Saletu; Al, Sharpley; F., Turek; E., Van Cauter; J., Mendlewicz
abstract

A wide range of studies have been published over the past two decades that involve the intersection of sleep EEG, insomnia, psychiatric illness (especially depressive disorders) and psychopharmacology. Much of value has been discovered, but there have also been false starts and contradictory results. There is in fact strong evidence that insomnia is associated with medical and psychiatric illness and that the sleepiness associated with insomnia is the cause of many accidents. Thus, the direct (visits to doctors, cost of sleeping medication, complications from use of these medications) and indirect (accidents, quality of life) costs of insomnia are enormous and constitute a major public health problem in the industrialized countries. Believing that it is now timely to assess the state of this important research area, a consensus conference was convened on June 26-28, 1998, in Porto Cervo (Italy) to attempt to clarify the important issues and findings on the clinical effect of the different classes of antidepressant drugs on sleep quality in depression. The participants´ consensus on some of the main topics is presented with the hope that this discussion and analysis will contribute to productive research in this important field. Copyright


2000 - Second messenger-regulated protein kinases in the brain: Their functional role and the action of antidepressant drugs [Articolo su rivista]
M., Popoli; Brunello, Nicoletta; J., Perez; G., Racagni
abstract

Depression has been treated pharmacologically for over three decades, but the views regarding the mechanism of action of antidepressant drugs have registered recently a major change. It was increasingly appreciated that adaptive changes in postreceptor signaling pathways, rather than primary action of drugs on monoamine transporters, metabolic enzymes, and receptors, are connected to therapeutic effect. For some of the various signaling pathways affected by antidepressant treatment, it was shown that protein phosphorylation, which represents an obligate step for most pathways, is markedly affected by long-term treatment. Changes were reported to be induced in the function of protein kinase C, cyclic AMP-dependent protein kinase, and calcium/calmodulin-dependent protein kinase, For two of these kinases (cyclic AMP- and calcium/calmodulin-dependent), the changes have been studied in isolated neuronal compartments (microtubules and presynaptic terminals). Antidepressant treatment activates the two kinases and increases the endogenous phosphorylation of selected substrates (microtubule-associated protein 2 and synaptotagmin). These modifications may be partly responsible for the changes induced by antidepressants in neurotransmission. The changes in protein phosphorylation induced by long-term antidepressant treatment may contribute to explain the therapeutic action of antidepressants and suggest new strategies of pharmacological intervention.


2000 - Social phobia: diagnosis and epidemiology, neurobiology and pharmacology, comorbidity and treatment [Articolo su rivista]
Brunello, Nicoletta; JA den, Boer; Ll, Judd; S., Kasper; Je, Kelsey; M., Lader; Y., Lecrubier; Jp, Lepine; Rb, Lydiard; J., Mendlewicz; Sa, Montgomery; G., Racagni; Mb, Stein; Hu, Wittchen
abstract

Social phobia is a common disorder associated with significant psychosocial impairment, representing a substantial public health problem largely determined by the high prevalence, and the lifelong chronicity. Social phobia starts in early childhood or adolescence and is often comorbid with depression, other anxiety disorders, alcohol and substance abuse or eating disorders. This: cascade of comorbidity, usually secondary to social phobia, increases the disability associated with the condition. The possibility that social phobia may be a trigger for later developing comorbid disorders directs attention to the need for early effective treatment as a preventive measure. The most recent drug class to he investigated for the psychopharmacological treatment of social phobia is the SSRI group for which there is growing support. The other drugs classes that have been evaluated are monoamine oxidase inhibitors (MAOIs), benzodiazepines, and beta-blockers, The SSRIs represent a new and attractive therapeutic choice for patients with generalized social phobia. Recently the fil st, large scale, placebo-controlled study to assess the efficacy of drug treatment in generalized social phobia has been completed with paroxetine, Paroxetine was more effective in reducing the symptoms than placebo and was well tolerated. Many now regard SSRIs as the drugs of choice in social phobia because of their effectiveness and because they avoid the problems of treatment with benzodiazepines or classical MAOIs. (C) 2000 Elsevier Science B.V. All rights reserved.


1999 - Dysthymia: clinical picture, extent of overlap with chronic fatigue syndrome, neuropharmacological considerations, and new therapeutic vistas [Articolo su rivista]
Brunello, Nicoletta; H., Akiskal; P., Boyer; G. L., Gessa; R. H., Howland; S. Z., Langer; J., Mendlewicz; M. P., de Souza; G. F., Placidi; G., Racagni; S., Wessely
abstract

Dysthymia, as defined in the American Psychiatric Association and International Classification of Mental Disorders, refers to a prevalent form of subthreshold depressive pathology with gloominess, anhedonia, low drive and energy, low self-esteem and pessimistic outlook. Although comorbidity with panic, social phobic, and alcohol use disorders has been described; the most significant association is with major depressive episodes. Family history is loaded with affective, including bipolar, disorders. The latter finding explains why dysthymia, especially when onset is in childhood, can lead to hypomanic switches, both spontaneously and upon pharmacologic challenge in as many as 30%. Indeed, antidepressants from different classes - tricyclic antidepressants (TCAs), monoamine oxidase inhibitors (MAOIs), reversible inhibitors of monoamine oxidase A (RIMAs), selective serotonin-reuptake inhibitors (SSRIs) and, more recently, amisulpride, and spanning noradrenergic, serotonergic as well as dopaminergic mechanisms of action - have been shown to be effective against dysthymia in an average of 65% of cases. This is a promising development because social and characterologic disturbances so pervasive in dysthymia often, though not always, recede with continued pharmacotherapy beyond acute treatment. Despite symptomatic overlap of dysthymia with chronic fatigue syndrome - especially with respect to the cluster of symptoms consisting of low drive, lethargy, lassitude and poor concentration - neither the psychopathologic status, nor the pharmacologic response profile of the latter syndrome is presently understood. Chronic fatigue today is where dysthymia was two decades ago. We submit that the basic science - clinical paradigm that has proven so successful in dysthymia could, before too long, crack down the conundrum of chronic fatigue as well. At a more practical level, we raise the possibility that a subgroup within the chronic fatigue group represents a variant of dysthymia. (C) 1999 Elsevier Science B.V. All rights reserved.


1999 - Physiology to functionality: The brain and neurotransmitter activity [Relazione in Atti di Convegno]
Racagni, G.; Brunello, N.
abstract

The monoamine hypothesis of depression proposes that the biological basis for depression is a deficiency in the neurotransmitters serotonin or noradrenaline, or both. Over the past 30 years this hypothesis has been refined as more experimental and clinical evidence has emerged. The selective serotonin reuptake inhibitors in particular have made a significant contribution to our understanding of the role of serotonin in depression. Our knowledge of the noradrenergic system is less complete, mainly because, until recently, no equivalent agents were available for this system. With the advent of reboxetine, the first selective noradrenaline reuptake inhibitor, attention is again focusing on the role of noradrenaline in depression. To an extent, the action of the selective inhibitors can be predicted through knowledge of the neuroanatomy of the central and peripheral nervous systems. With regard to depression, the most important pathways are those of the serotonergic and noradrenergic neurones projecting to the prefrontal cortex, from the raphe nucleus and locus coeruleus, respectively. However, increasing the levels of the monoamines in the central nervous system affects many other pathways and a wide range of serotonin and adrenergic receptors, leading to a multiplicity of potential beneficial and adverse events. In addition, the complex intracellular responses are beginning to be examined, again with the aid of the selective antidepressants.


1999 - Regulation of ionotropic glutamate receptors in the rat brain in response to the atypical antipsychotic seroquel (quetiapine fumarate). . [Articolo su rivista]
Tascedda, Fabio; E., Lovati; Blom, Johanna Maria Catharina; P., Muzzioli; Brunello, Nicoletta; G., Racagni; M. A., Riva
abstract

The interplay between dopamine and glutamate appears to be relevant in the etiopathology of schizophrenia. Although currently used antipsychotics do not interact with glutamatergic receptors, previous results have demonstrated that the expression profile of ionotropic glutamate receptors can be regulated by drugs such as haloperidol or clozapine. In the present investigation, the mRNA levels for NMDA and AMPA receptor subunits were measured after chronic treatment with the novel antipsychotic agent Seroquel (quetiapine fumarate, quetiapine) as compared to haloperidol and clozapine. Similarly to the prototype atypical clozapine, quetiapine reduced the mRNA expression for NR-1 and NR-2C, two NMDA forming subunits, in the nucleus accumbens. Furthermore, quetiapine, but not haloperidol or clozapine, increased the hippocampal expression for the AMPA subunits GluR-B and GluR-C. The differences between classical and atypical antipsychotics, as well as among the novel agents, might be relevant for specific aspects of their therapeutic activity and could provide valuable information for the role of glutamate in specific symptoms of schizophrenia.


1998 - Guidelines for depot antipsychotic treatment in schizophrenia [Articolo su rivista]
Kane, J. M.; Aguglia, E.; Altamura, A. C.; Gutierrez, J. L. A.; Brunello, N.; Fleischhacker, W. W.; Gaebel, W.; Gerlach, J.; Guelfi, J. -D.; Kissling, W.; Lapierre, Y. D.; Lindstrom, E.; Mendlewicz, J.; Racagni, G.; Carulla, L. S.; Schooler, N. R.
abstract

These guidelines for depot antipsychotic treatment in schizophrenia were developed during a two-day consensus conference held on July 29 and 30, 1995 in Siena, Italy. Depot antipsychotic medications were developed in the 1960s as an attempt to improve the long-term treatment of schizophrenia (and potentially other disorders benefiting from long-term antipsychotic medication). Depot drugs as distinguishable from shorter acting intramuscularly administered agents can provide a therapeutic concentration of at least a seven day duration in one parenteral dose. The prevention of relapse in schizophrenia remains an enormous public health challenge worldwide and improvements in this area can have tremendous impact on morbidity, mortality and quality of life, as well as direct and indirect health care costs. Though there has been debate as to what extent depot (long-acting injectable) antipsychotics are associated with significantly fewer relapses and rehospitalizations, in our view when all of the data from individual trials and metaanalyses are taken together, the findings are extremely compelling in favor of depot drugs. However in many countries throughout the world fewer than 20% of individuals with schizophrenia receive these medications. The major advantage of depot antipsychotics over oral medication is facilitation of compliance in medication taking. Non-compliance is very common among patients with schizophrenia and is a frequent cause of relapse. In terms of adverse effects, there are not convincing data that depot drugs are associated with a significantly higher incidence of adverse effects than oral drugs. Therefore in our opinion any patient for whom long-term antipsychotic treatment is indicated should be considered for depot drugs. In choosing which drug the clinician should consider previous experience, personal patient preference, patients history of response (both therapeutic and adverse effects) and pharmacokinetic properties. In conclusion the use of depot antipsychotics has important advantages in facilitating relapse prevention. Certainly pharmacotherapy must be combined with other treatment modalities as needed, but the consistent administration of the Former is often what enables the latter.


1998 - International Academy for Biomedical and Drug Research Consensus Conference, San Juan, December 8-9, 1996: Novel therapeutic indications of antidepressant drugs: Introduction [Articolo su rivista]
Brunello, N.
abstract


1998 - Rationale for the development of noradrenaline reuptake inhibitors [Relazione in Atti di Convegno]
Brunello, N.; Racagni, G.
abstract

Elucidation of the mechanism of action of antidepressants led to the hypothesis that depression is caused by dysfunction in either the noradrenergic or serotonergic neurotransmitter systems. As inconsistencies in studies designed to confirm this hypothesis arose, the consensus on the biological basis of depression is being refined. The need for better tolerated and effective antidepressants has resulted in the development of agents with more specific receptor binding profiles than the tricyclic antidepressants. These newer antidepressants selectively inhibit the reuptake of noradrenaline (selective NARIs), serotonin (SSRIs) or both (SNRIs). They are useful tools for describing changes in neuroreceptors and intracellular events that occur during antidepressant pharmacotherapy. Reboxetine, a selective NARI, down-regulates beta-adrenergic receptors and desensitises noradrenaline-coupled adenylate cyclase. It also affects cAMP- and Ca2+/calmodulin-dependent phosphorylation systems in a different manner to tricyclic antidepressants and SSRIs. This implies that although different classes of antidepressants may affect common central pathways, the ways in which they do this are distinctive. In conclusion, reboxetine, a selective NARI which is well tolerated and effective in the treatment of depression, has provided us with a new insight into the action of antidepressants and thus will help us to refine our theory of the biological basis of depression.


1998 - The role of reboxetine, a selective noradrenaline reuptake inhibitor, in the treatment of depression: Discussion [Articolo su rivista]
Brunello, N.; Ragagni, G.; Szabadi, E.; Bradshaw, C. M.; Boston, P. F.; Langely, R. W.; Hindmarch, I.; Ban, T. A.; Gaszner, P.; Aguglia, E.; Batista, R.; Costillo, A.; Lipscey, A.; Macher, J. P.; Torrez Ruiz, A.; Vergara, L.
abstract


1998 - cAMP-dependent phosphorylation system after short and long-term administration of moclobemide [Articolo su rivista]
S., Mori; R., Zanardi; M., Popoli; S., Garbini; Brunello, Nicoletta; E., Smeraldi; G., Racagni; J., Perez
abstract

Accumulating evidence suggested that signal transduction cascade including protein phosphorylation is implicated in the neurochemical action of antidepressant agents. Clinical data indicated that moclobemide, a short acting and reversible inhibitor of monoamino oxidase type A, is an effective antidepressant medication. However, little is known about the intracellular effects of this compound. Thus, in the present study we assessed the binding of cAMP to cAMP-dependent protein kinase (PKA) in rat cerebral cortex following short and long-term administration of moclobemide. The results showed that 21 days of treatment with moclobemide significantly increased the specific [P-32]-cAMP covalent binding into the soluble 52-54 kDa cAMP-receptor. This effect was not seen following 1, 5 and 12 days of treatment. These findings suggest that PKA could be implicated in the biochemical effects of moclobemide. (C) 1998 Elsevier Science Ltd. All rights reserved.


1997 - Practical issues in clinical research of schizophrenia [Articolo su rivista]
Brunello, Nicoletta; L. L., Judd; S. Z., Langer; J., Mendlewicz; G., Racagni
abstract

none


1996 - Cyclic GMP inhibition of metabotropic glutamate receptor-induced phosphoinositide hydrolysis in mesencephalic neurons [Articolo su rivista]
A., Ambrosini; L., Bresciani; Brunello, Nicoletta; G., Racagni
abstract

The effect of cGMP on metabotropic glutamate receptor-induced stimulation of phosphoinositide hydrolysis in mesencephalic neuronal cultures was evaluated by cell incubation with the stable analogue dibutyryl-cGMP (10 mu M). A complete blockade of (1S,3R)-1-aminocyclopentane-1,3-dicarboxylic acid- or quisqualate-induced inositol phosphate formation was observed. Ionotropic glutamate receptors in mesencephalic neurons activate cGMP formation and, through this intracellular messenger, they might control mGluR activity. Copyright


1996 - Effect of idebenone on in vivo serotonin release and serotonergic receptors in young and aged rats [Articolo su rivista]
C., Scavini; A., Rozza; E., Lanza; L., Favalli; G., Racagni; Brunello, Nicoletta
abstract

The effect of idebenone on the serotonergic system was evaluated in the aging rat by measuring the kinetic constants of H-3-5HT and H-3-ketanserin binding sites in the cerebral cortex of rats at 3, 15 and 24 months of age following acute and subchronic administration of the drug. Idebenone displayed no in vitro affinity toward any population of serotonin receptors and did not modify their kinetic parameters after a single dose of 100 mg/kg at any age tested. A subchronic treatment with the drug for 21 days at the dose of 30 mg/kg did not induce any relevant change in 3- and 15-month-old rats, whereas it significantly increased the density of both H-3-5HT and H-3-ketanserin binding sites in 24-month-old rats, where a lower number of receptors is detected as a consequence of aging. This effect was rather specific, since under the same experimental conditions no changes were detected in the density of cortical beta-adrenergic receptors in aged animals. In microdialysis studies, acute administration with idebenone did not affect 5HT and 5HIAA release at any age. Conversely, the pattern of serotonin metabolism was significantly modified in aged rats following repeated treatment with idebenone and was partially restored to a value similar to the one observed in young animals. These results suggest that idebenone, a putative neuroprotective agent which has been shown to improve brain metabolism in ischemic conditions, might also attenuate age-associated neuronal damage, acting probably on several neurotransmitter systems which undergo selective modification during aging.


1995 - Critical issues in the treatment of affective disorders [Articolo su rivista]
Brunello, N.; Burrows, G. D.; Jonsson, B.; Judd, L. L.; Kasper, S.; Keller, M. B.; Kupfer, D. J.; Lecrubier, Y.; Mendlewicz, J.; Montgomery, S. A.; Nemeroff, C. B.; Preskorn, S.; Racagni, G.; Rush, A. J.
abstract


1995 - Introduction [Articolo su rivista]
Brunello, N.
abstract


1995 - Metabotropic glutamate receptors negatively coupled to adenylate cyclase inhibit NMDA receptor activity and prevent neurotoxicity in mesencephalic neuron in vitro [Articolo su rivista]
A., Ambrosini; L., Bresciani; S., Fracchia; Brunello, Nicoletta; G., Racagni
abstract

The functional effects of G protein-linked glutamate receptor activation have been studied in mouse mesencephalic neurons in vitro. We have been able to identify two receptor classes, one linked to phosphoinositide hydrolysis and another that inhibits adenylate cyclase. The agonist (1S,3R)-aminocyclo-pentane-1,3-dicarboxylate (ACPD) affected the two responses with similar potency (EC(50) = 2 and 7 mu M, respectively). In contrast, (2S,3S,4S)-alpha-(carboxycyclopropyl)glycine selectively decreased adenylate cyclase activity (EC(50) = 150 nM), without interfering with the phosphoinositide pathway. Activation of ion channel-linked glutamate receptors in mesencephalic neurons leads to cGMP formation. In this study, we demonstrate that cell pretreatment with ACPD or (2S,3S,4S)-alpha-(carboxycyclopropyl)glycine prevented, in a dose-dependent fashion, N-methyl-D-aspartate (NMDA)-induced cGMP formation but not the kainate-stimulated response. The pharmacological profile suggests that receptors that are negatively coupled to adenylate cyclase are responsible for this effect. Coexposure of neurons to ACPD and Ba2+, a K+ channel blocker, counteracted the ACPD-induced blockade of NMDA receptors, suggesting that activation of K+ conductances could be involved in the post-transduction events triggered by metabotropic receptors in the mesencephalon. Neuronal treatment with NMDA for 10 min caused a reduction in mitochondrial activity. Direct inhibition of nitric oxide synthase with the inhibitor N-G-nitro-L-arginine or removal of extracellular nitric oxide with reduced hemoglobin did not prevent this metabolic impairment, thus excluding a role for nitric oxide in this test for excitotoxicity. On the contrary, the mitochondrial function was maintained when neurons exposed to NMDA were preincubated with metabotropic receptor agonists. To summarize, our results suggest that metabotropic receptors that are negatively coupled to adenylate cyclase exert modulatory control specifically on NMDA receptor activity. This event could also contribute to the reduction of neurotoxic effects due to NMDA receptor hyperactivity.


1995 - New insights into the biology of schizophrenia through the mechanism of action of clozapine. [Articolo su rivista]
Brunello, Nicoletta; C., Masotto; L., Steardo; R., Markstein; G., Racagni
abstract

Many studies have detected in the brain of schizophrenic patients various morphological and structural abnormalities in various regions and in particular in the cortical and limbic areas. These abnormalities might in part result from neurodevelopmental disturbances suggesting that schizophrenia might have organic causes. These abnormalities may be the primary event in schizophrenia and be responsible for altered dopaminergic, but not only dopaminergic, neurotransmission in these regions. If schizophrenia is in some way strictly related to brain morphological abnormalities it becomes hard to believe that a curative treatment will ever be possible. Considering this scenario, treatment of schizophrenia will be restricted to symptomatic and preventive therapy and therefore, more effective and better tolerated antipsychotics are necessary. The widely used classical antipsychotic drugs present some disadvantages. They do not improve all symptoms of schizophrenia, are nor effective in all patients, produce a number of unpleasant and serious, and partly irreversible, motor side effects. The atypical antipsychotic clozapine constitutes a major advance in particular for patients nor responding to conventional neuroleptics. To explain the unique therapeutic effect of clozapine many hypotheses have been proposed. Most of the explanations given so far assume that the D-2 blockade is the basis for the antipsychotic activity of clozapine and that the difference in respect to other antipsychotics is due to the contribution of other receptor interactions. Considering the dopaminergic receptor in particular the recently discovered D-4 receptor subtype, it has been observed that even if several classical neuroleptics exhibit high affinity to the D-4 receptor, clozapine is more selective for this subtype compared to D-2 receptors. Moreover clozapine, differently from all other conventional neuroleptics, is a mixed but weak D-1/D-2 antagonist. This observation has prompted speculation that the synergism between D-1 and D-2 receptors might allow antipsychotic effects to be achieved below the threshold for unwanted motor side effects. Probably the D-1 antagonistic activity exerted by clozapine at low doses enhances preferentially the extracellular concentration of dopamine in specific areas of the brain, such as the prefrontal cortex, where a dopaminergic hypoactivity has been suggested to be in part responsible for negative symptoms of schizophrenia. The clozapine enhancement of dopaminergic activity in this brain area might explain its efficacy against schizophrenia negative symptoms. However, it cannot be excluded that the affinities displayed by clozapine for other nondopaminergic receptors also contribute to its unique therapeutic profile. The various hypotheses mentioned in this review need to be further validated or disproved. The only way to do that is developing new drugs where the postulated mechanistic profile is specifically realized and to clinically test these compounds.


1995 - Relationships between conformational behaviour and binding affinity towards β1 and β2 adrenoceptors of some chiral phenoxypropanolamines with bulky N-substituents [Articolo su rivista]
Villa, L.; Villa, A. M.; Pallavicini, M.; Romeo, S.; Valoti, E.; Ferri, V.; Iuliano, E.; Brunello, N.
abstract

The optical isomers of a series of phenoxypropanolamine compounds with N-substituents bulkier than isopropyl have been synthesized, and their binding affinity towards beta(1) and beta(2)-adrenoceptors has been determined. A computational study, including a Molecular Dynamics (MD) simulation and quenching in water and a GRID analysis provided some useful suggestions for possible interpretation patterns for the different affinity exhibited by the compounds studied.


1995 - Synthesis of 1,4-disubstituted-2-methylpiperazines and study of their analgesic properties [Articolo su rivista]
Barlocco, D.; Brunello, N.; Bernardi, M.; Cignarella, G.
abstract

A series of N,N-disubstituted-2-methylpiperazines (3a-e) has been synthesized and compounds tested both in vitro and in vivo for their analgesic properties. Binding studies showed that the new compounds are devoid of relevant affinity towards μ receptors. However, compound 3a displayed significant analgesic properties both in the hot plate test and in the mouse phenyl-p-benzoquinone induced abdominal constriction test (MAC), whereas the other derivatives were found active only in MAC test. Moreover, it should be noted that compound 3a elicited a Straub-tail reaction also at the lowest administered dose (10 mg/kg, ip) and this effect was antagonized by naloxone.


1994 - Current understanding of the mechanism of action of classic and newer antidepressant drugs [Articolo su rivista]
Brunello, N.; Langer, S. Z.; Perez, J.; Racagni, G.
abstract

Until 1980, the two major classes of antidepressant drugs were the tricyclics (TCAs) and the monoamine oxidase inhibitors (MAOIs). During the 1980s several additional classes of drugs appeared. Among these, selective serotonin reuptake inhibitors (SSRIs) have a distinct and well‐characterized mechanism of action, and have become among the most widely prescribed antidepressant drugs. A common factor in the action of antidepressants with different chemical structures is the ability to increase the synaptic availability of one or more neurotransmitters, as a result either of blockade of the reuptake process or inhibition of metabolic degradation. However, the delay in clinical response to antidepressants suggests that, even if the acute pharmacologic effects of antidepressants may be essential for efficacy, symptom resolution requires some adaptive changes in neurotransmission. More recent studies, therefore, have focused on the signal transduction process beyond the receptor level, and on the regulation of gene expression of specific proteins. The side‐effect profiles of antidepressants are related to their acute mechanisms of action. TCAs lack specificity of their pharmacological action, and are associated with antimuscarinic, antihistaminic, and alpha‐adrenergic blocking actions, that account for many unwanted side effects but do not contribute to their therapeutic profile. The SSRIs, as a result of their selective action to inhibit serotonin reuptake, appear to have more tolerable side effects than TCAs and are less likely to be lethal when taken in overdose. SSRIs have little or no affinity for neurotransmitter receptors. The most common side effect is gastrointestinal discomfort. Depression 2:119–126 (1994/1995). © 1995 Wiley‐Liss, Inc. Copyright © 1994 Verlag Chemie, GmbH


1994 - Molecular biology of dopaminergic receptors: Pharmacological implications [Articolo su rivista]
Fumagalli, F.; Brunello, N.; Racagni, G.
abstract

Molecular biology techniques have had a tremendous impact on the studies of neurotransmitter receptors, in particular of dopamine receptors. Until recently only two dopamine receptors were described on the basis of pharmacological and biochemical experiments: the D1 and D2 receptors, differently coupled to adenyl cyclase and other second messenger systems. Molecular cloning studies have revealed the existence of five functional dopamine receptors. Several studies have been conducted regarding the distribution of these different receptor mRNAs and the expression of dopamine receptors in transfected cell lines in order to better understand the functional significance of the individual receptors. At present, little is known about the physiological and pharmacological importance of this receptor heterogeneity. Future reseach should allow us to get a better insight in the physical structure of these receptors in order to prepare highly selective dopaminergic ligands to use in the therapies related to dysfunction of the dopaminergic system. In fact this heterogeneity gives the possibility for developing selective antagonists for use in clinical therapeutics. Recently clinical studies have been focused on drugs like clozapine and risperidone which can modulate not only dopamine receptors but also serotonergic receptors (5HT2) having less extrapyramidal side effects characteristic of the blockage of D2 receptors.


1994 - Molecular properties of the WB4101 enantiomers and of its chiral methyl derivatives for α1-adrenoceptor recognition [Articolo su rivista]
Villa, L.; Valoti, E.; Villa, A. M.; Pallavicini, M.; Ferri, V.; Iuliano, E.; Brunello, N.
abstract

The optical isomers of the well known alpha(1) - antagonist WB4101 and of its derivatives with a methyl group in the oxyethyl moiety were prepared for the evaluation of their alpha - adrenoceptors binding affinity. By means of a detailed computational analysis, the present work shows that the introduction of a methyl group affects the behaviour of WB4101 in different ways. A limitation of the conformational freedom in certain regions of the torsional subspace of the potential energy function, differences in the reactivity of the protonated species towards a model proton acceptor and the quality of the superposition with the rigid template for alpha(1) antagonists, corynanthine, are examined and discussed in order to select a candidate bioactive form and possible features which act as modulators of the recognition process at the the alpha(1) - adrenoceptors.


1993 - Changes of lymphocyte beta‐adrenergic receptors and platelet serotonin reuptake sites in aging and dementias [Articolo su rivista]
Rozza, A.; Scavini, C.; Steardo, L.; Guaita, A.; Favalli, L.; Racagni, G.; Brunello, N.
abstract

We have studied the kinetic constants of β;‐adrenergic receptors in lymphocytes and of serotonin uptake sites in platelets from adult and elderly volunteers and from aged patients with a diagnosis of multi‐infarct dementia (MID) or senile dementia of Alzheimer's type (SDAT). The results showed that in the physiological aging process, β;‐adrenergic receptor density is reduced and 5HT transport system is enhanced. MID is associated to a loss of β;‐adrenoceptor affinity and a slight increase in the capacity of 5HT uptake. The kinetic constants of both β;‐adrenergic receptors in lymphocytes and 5HT uptake sites in platelets from SDAT patients were comparable to those observed in the adult population, being thus different from those found in the aged matched volunteers. These changes could be interpreted as resulting from compensatory mechanisms that replaced altered plasma monoamine concentrations. The apparent absence of such changes in SDAT patients could suggest the lack of these compensatory effects. Copyright © 1993 John Wiley &amp; Sons, Ltd.


1993 - Evidence for the existence of cAMP-dependent protein kinase phosphorylation system associated with specific phosphoproteins in stable microtubules from rat cerebral cortex [Articolo su rivista]
Perez, J.; Tinelli, D.; Cagnoli, C.; Pecin, P.; Brunello, N.; Racagni, G.
abstract

Cyclic AMP is a second messenger by which different extracellular signals are transduced into biological responses. Within the cell, most of the effects of cAMP are mediated through the cAMP protein kinase which appears to be localized in specific compartments of the cell near to their substrate proteins. In the present study, we have investigated the possible association of cAMP-dependent protein kinase, its substrate proteins and RII binding proteins in stable microtubules from rat cerebral cortex. The results show that in this fraction there is a cAMP binding protein of 52-54 kDa. This cAMP receptor is in the inactive holoenzyme form, since the addition of cAMP (5 μM) induces an increase in the endogenous phosphorylation of different stable microtubules polypeptides, which is completely inhibited in the presence of a specific protein kinase inhibitor (PKI 5-24 1 μM). Interestingly, overlay binding assay reveals that beside MAP2, 32P/R II is able to bind stable microtubule proteins of Mr 150 and 75 kDa which, according to their electrophoretic mobility, can also be endogenous substrates for the enzyme. We conclude that cAMP-dependent phosphorylation system is indeed associated with stable microtubules from rat cerebral cortex. © 1993.


1993 - Heterologous sensitization of adenylate cyclase activity by serotonin in the rat cerebral cortex [Articolo su rivista]
Rovescalli, A. C.; Brunello, N.; Perez, J.; Vitali, S.; Steardo, L.; Racagni, G.
abstract

In vitro exposure of rat cerebrocortical slices to μM concentrations of serotonin (5HT) results in an increased response of adenylate cyclase to isoproterenol (ISO). No change in the affinity of the β-adrenoceptor toward the agonist was found after 5HT exposure when measuring ISO displacement of [3H]CGP 12177 binding. A similar increase of adenylate cyclase response was also found when using VIP as a stimulatory agent. The dose-response curve of adenylate cyclase to the GTP analogue, GppNHp, was modified by 5HT, which promotes a significantly higher maximal response without altering the potency of GppNHp. Forskolin-stimulated adenylate cyclase activity was not affected by 5HT. Serotonergic 5HT2 receptors are involved in the sensitization of adenylate cyclase to GppNHp, since the selective 5HT2 antagonist ketanserin inhibits the effect of 5HT, whereas the 5HT2 agonist DOI mimics 5HT. The involvement of 5HT2 receptor-coupled activation of protein kinase C is also demonstrated: direct protein kinase C activators such as phorbol esters and s,n-dioctanoylglycerol behave in the same manner as 5HT, while the protein kinase C inhibitor CGP 41251 prevents 5HT from increasing adenylate cyclase responsiveness to GppNHp. Moreover, in vitro exposure of cortical slices to 5HT results in reduced inhibition of adenylate cyclase by somatostatin. Since no change was observed at the receptor level and in the direct stimulation of the catalytic subunit of the enzyme, we propose that 5HT might accomplish the sensitization of adenylate cyclase through protein kinase C by inactivating the inhibitory coupling protein Gi and facilitating the interaction of the exogenous GppNHp with the stimulatory coupling protein Gs. © 1993.


1993 - Protein phosphorylation as intracellular target in the action of MAO-A inhibitors and other antidepressants [Relazione in Atti di Convegno]
Perez, J.; Cagnoli, C.; Mori, S.; Tinelli, D.; Migliavacca, E.; Brunello, N.; Racagni, G.
abstract


1992 - Age-related variations in relative abundance of alternative spliced D2 receptor mRNAs in brain areas of two rat strains [Articolo su rivista]
Della Vedova, F.; Fumagalli, F.; Sacchetti, G.; Racagni, G.; Brunello, N.
abstract

Age-related reduction in the steady-state levels of messenger RNA for D2javax.xml.bind.JAXBElement@698be68c and D2javax.xml.bind.JAXBElement@3b75e9a8, the alternatively spliced form of dopamine D2 receptors, was observed in different rat brain areas using the sensitive reserve transcription (RT)-polymerase chain reaction (PCR) technique. In both Sprague-Dawley and Wistar aged rats, the decrease was more pronounced in the D2javax.xml.bind.JAXBElement@6e2a8528 isoform mRNA thus leading to a changed ratio in striatum as well as in the hyppocampus. © 1992.


1992 - Biochemical and molecular studies after long term administration of 5HT reuptake inhibitors: comparison with other antidepressants [Articolo su rivista]
Perez, J.; Tinelli, D.; Cagnoli, C.; Pecin, P.; Brunello, N.; Racagni, G.
abstract


1992 - New biochemical hypotheses on the mechanism of action of antidepressant drugs: cAMP - Dependent phosphorylation system [Relazione in Atti di Convegno]
Racagni, G.; Brunello, N.; Tinelli, D.; Perez, J.
abstract


1992 - cAMP-dependent phosphorylation system, a biochemical target in the action of antidepressant drugs [Relazione in Atti di Convegno]
Brunello, N.; Perez, J.; Tinelli, D.; Cagnoli, C.; Fumagalli, F.; Racagni, G.
abstract


1991 - Localization of nerve growth factor receptor mRNA in contused rat spinal cord by in situ hybridization [Articolo su rivista]
Reynolds, M. E.; Brunello, N.; Mocchetti, I.; Wrathall, J. R.
abstract

Northern blot analysis using a probe for the low-affinity nerve growth factor receptor (NGFR) revealed that a mild contusive injury induces the expression of NGFR mRNA in rat spinal cord with a maximal expression at 7 days post-injury. We have now localized this induction using in situ hybridization and found the highest concentration of NGFR mRNA at the lesion epicenter. The location and pattern of autoradiographic grains were compared with that of various cell types at the injury site as determined by immunocytochemical studies. The results suggest that cells associated with blood vessels at the epicenter are induced to express NGFR mRNA at 7 days post-injury. © 1991.


1991 - cAMP binding proteins in the rat cerebral cortex after administration of selective 5-HT and NE reuptake blockers with antidepressant activity [Articolo su rivista]
Perez, J.; Tinelli, D.; Bianchi, E.; Brunello, N.; Racagni, G.
abstract

This study was undertaken to evaluate the cyclic adenosine monophosphate (cAMP) binding proteins in the cerebral cortex of rat short- and long-term administration with antidepressants. Prolonged treatment with different antidepressants that inhibit serotonin or norepinephrine uptake such as fluoxetine and the (+) enantiomer of oxaprotiline, respectively, was able to induce an increase in the photoactivated incorporation of 8-N3-[32P]cAMP into a protein band with apparent molecular weight of 52,000 in both soluble and crude microtubule fraction. On the contrary, chronic treatment with the (-) enantiomer of oxaprotiline, which does not affect monoamine uptake, failed to produce this effect. Moreover, no changes were observed after acute or in vitro addition of antidepressants, suggesting that modification in the cAMP binding may be related to adaptive changes elicited by prolonged antidepressants treatment. In conclusion, our studies indicate that the cAMP binding protein associated with the crude microtubule fraction could be an intracellular target for the action of antidepressant drugs.


1990 - BIOCHEMICAL AND MOLECULAR CHANGES IN RAT CEREBRAL CORTEX AFTER CHRONIC ANTIDEPRESSANT TREATMENT: ‘IN VITRO AND’‘IN VIVO’ STUDIES [Articolo su rivista]
Brunello, N.; Perez, J.; Tinelli, D.; Rovescalli, A. C.; Racagni, G.
abstract


1990 - Increased nerve growth factor receptor mRNA in contused rat spinal cord [Articolo su rivista]
Brunello, N.; Reynolds, M.; Wrathall, J. R.; Mocchetti, I.
abstract

Peripheral nerve injury induces the expression of nerve growth factor receptor (NGFR). To determine whether a similar induction results from injury of the spinal cord, NGFR mRNA content was determined using Northern blot hybridization analysis of total RNA from spinal cords of rats contused in the mid-thoracic region. By four days after contusion NGFR mRNA was significantly increased in the thoracic segments that included the injury site. The induction was maximal at 7 days, about 5- to 7-fold the level of uninjured controls, and remained 4 times higher than controls at 14 and 28 days after injury. These results suggest that axotomy in the CNS may also trigger the molecular mechanism(s) leading to up-regulation of NGFR expression. © 1990.


1990 - Influence of chronic morphine treatment on dopaminergic sensitivity [Articolo su rivista]
Lucchelli, A.; Boselli, C.; Brunello, N.
abstract


1990 - Modification of transducing and intracellular mechanisms in rat cerebral cortex after chronic treatment with antidepressant drugs [Articolo su rivista]
Brunello, N.; Tinelli, D.; Bianchi, E.; Rovescalli, A. C.; Racagni, G.
abstract


1990 - Receptor modulation and cAMP dependent phosphorylation system after repeated antidepressant administration [Relazione in Atti di Convegno]
Racagni, G.; Perez, J.; Tinelli, D.; Bianchi, E.; Cagnoli, C.; Brunello, N.
abstract


1989 - Behavioural and neurochemical changes produced by lefetamine in two inbred strains of mice [Articolo su rivista]
Cagiano, R.; De Salvia, M. A.; Tattoli, M.; Lacomba, C.; Brunello, N.; Racagni, G.; Cuomo, V.
abstract


1989 - Effect of Different Photoperiod Exposure on [3H]Imipramine Binding and Serotonin Uptake in the Rat Brain [Articolo su rivista]
Rovescalli, A. C.; Brunello, N.; Riva, M.; Galimberti, R.; Racagni, G.
abstract

Abstract: Seasonal rhythmicity in the occurrence of acute depressive episodes and the therapeutic efficacy of light exposure suggest the possible involvement of the pineal gland or other biological oscillators in the pathophysiology of depressive illness. We have performed studies to clarify whether different light/dark (LD) cycle schedules may induce changes in the biochemical targets of antidepressants in the rat CNS. In particular, we have investigated the effect of short‐ (LD 8: 16) or long‐day (LD 14:10) photoperiods on different biochemical parameters of serotonergic neurons. A significant increase in the density of [3H]imipramine ([3H]IMI) binding and in the Vmax of 5‐[3H]hydroxytryptamine (5‐[3H]HT) uptake was found in the hypothalamus of LD 8:16‐ with respect to LD 14:10‐exposed rats, whereas no difference was found in the kinetic properties of postsynaptic 5‐HT receptors and in 5‐HT metabolism in the hypothalami and cerebral cortices of rats exposed to the two different photoperiods. A seasonal rhythm of [3H]IMI binding sites and 5‐HT uptake seems to exist only in certain brain areas, such as the hypothalamus, because no differences were found in the cerebral cortex of LD 14:10‐ and LD 8:16‐accustomed rats. [3H]IMI binding and 5‐HT uptake were significantly increased in the hypothalamus of rats accustomed to a light/dark‐inverted cycle (DL 10:14) and killed 6 h after the stopping of lighting in comparison to rats exposed to normal LD 14:10 cycles and killed 6 h after the beginning of lighting. Therefore, a circadian modification of the serotonergic presynaptic sites seems to be present and related to light/dark exposure. Because the existence of endogenous compounds able to modulate [3H]IMI binding and 5‐HT uptake, other than 5‐HT, has been postulated in the mammalian brain, the involvement of these substances in the periodic changes observed could be suggested. Copyright © 1989, Wiley Blackwell. All rights reserved


1989 - Effect of reboxetine, a new antidepressant drug, on the central noradrenergic system: Behavioural and biochemical studies [Articolo su rivista]
Riva, M.; Brunello, N.; Rovescalli, A. C.; Galimberti, R.; Carfagna, N.; Carminati, P.; Pozzi, O.; Ricciardi, S.; Roncucci, R.; Rossi, A.; Racagni, G.
abstract

The behavioural and biochemical effects of reboxetine, an alpha-ariloxybenzyl derivative of morpholine which possesses potent antireserpine activity and selective inhibition of norepinephrine reuptake, has been studied in rats for its potential antidepressant activity. These extensive in vitro and in vivo studies have shown that reboxetine cominbes the inhibitory properties of classical tricyclic antidepressants on the reuptake of norepinephrine with an ability to desensitize beta adrenergic receptor function. It does not have appreciable interaction with muscarinic cholinergic and alpha1 adrenergic receptors. These neurochemical characteristics suggest that reboxetine may be expected to be therapeutically effective with a lower incidence of side effects than commonly seen with tricyclic antidepressants.


1989 - Effects of p-chlorophenylalanine (p-CPA) on brain serotonin (5-HT) and ultrasonic calling during sexual behavior in rats [Articolo su rivista]
Cagiano, R.; De Salvia, M. A.; Tortella, E.; Sales, G.; Brunello, N.; Racagni, G.; Cuomo, V.
abstract


1989 - Neurotransmitter systems and receptor plasticity in brain aging [Articolo su rivista]
Racagni, G.; Rovescalli, A. C.; Galimberti, R.; Brunello, N.
abstract


1989 - cAMP-dependent phosphorylation of soluble and crude microtubule fractions of rat cerebral cortex after prolonged desmethylimipramine treatment [Articolo su rivista]
Perez, J.; Tinelli, D.; Brunello, N.; Racagni, G.
abstract

We have analyzed the cAMP-dependent phosphorylation system in the cerebral cortex and hippocampus of rats after acute and chronic administration of desmethylimipramine. Prolonged desmethylimipramine administration modified the cAMP-dependent endogenous phosphorylation of a protein band with apparent molecular weight 280 kDa from the cerebrocortical-soluble fraction. The effect appeared to be specific and associated with the chronic but not the acute administration of desmethylimipramine since we did not obtain any modification in other endogenous cAMP phosphoproteins of either the particulate or soluble fraction of the cerebral cortex. 280 kDa was identified as the soluble microtubule associated protein 2 on the basis of molecular weight, endogenous phosphorylation and immunological recognition. Prolonged desmethylimipramine administration did not induce any modification in the soluble cAMP-dependent endogenous phosphorylation of 280 kDa in other brain areas such as hippocampus, striatum or cerebellum, suggesting a region-specific effect of chronic desmethylimipramine treatment. Microtubule-associated protein 2 is a neuronal protein highly enriched in the dendritic portion of neurons and represents one of the major substrates in the cell for the type II cAMP protein kinase. Since the type II cAMP protein kinase that catalyzes the phosphorylation of microtubule-associated protein 2 copurifies with microtubules, we performed endogenous phosphorylation using increasing concentrations of cAMP in a crude microtubule preparation where microtubule-associated protein 2 appeared to be more concentrated. Under our conditions the maximal effect occurred at 1 μM cAMP, revealing increased 32P incorporation in microtubule-associated protein 2 from a crude microtubule preparation obtained from the cerebral cortex of rats treated with desmethylimipramine. Photoaffinity labelling with 8-azido-[32P]cAMP of the various fractions obtained during the preparation of crude microtubules (S1, S2 and crude microtubules) revealed an increase in the labelling of a protein band with apparent molecular weight of 52 kDa after desmethylimipramine treatment. The labelling of a 47 kDa protein band, which is also present in S1 and S2 fractions was, however, not altered by drug treatment. In conclusion, our studies demonstrated that prolonged desmethylimipramine treatment elicited specific changes in the phosphorylation system associated with a crude microtubule fraction. © 1989.


1988 - Age‐Related Changes in Rat Serotonergic and Adrenergic Systems and in Receptor Responsiveness to Subchronic Desipramine Treatment [Articolo su rivista]
Brunello, N.; Riva, M.; Rovescalli, A. C.; Galimberti, R.; Racagni, G.
abstract

Abstract: Noradrenergic (NA) and serotonergic (5‐HT) receptor profiles were compared in the cerebral cortex of young adult (3 months old) and aged (24 months old) male Sprague Dawley rats. β and α‐1 receptors were significantly decreased in 24 month old rats, whereas α‐2 receptors remained unchanged. 5‐HT‐2 Postsynaptic receptors and 5‐HT high affinity uptake were reduced in aged animals; on the other hand the number of 3H‐imipramine (3H‐IMI) recognition sites located on serotonin nerve terminals and labelled also by 3H‐paroxetine (3H‐PAR), were significantly higher in the cerebral cortex of old rats. The ratio 5‐hydroxyindoleacetic acid (5‐HIAA)/5‐HT, an index of serotonin turn‐over, increased in some brain areas of senescent rats. There were no age‐related changes in the responsiveness of investigated binding sites to subchronic desipramine treatment; the density of β‐noradrenergic and 5‐HT‐2 serotonergic receptors and that of 3H‐IMI binding sites was reduced both in young adult and aged rats. 1988 Nordic Pharmacological Society


1988 - Interaction of 3,8-diazabicyclo (3.2.1) octanes with mu and delta opioid receptors [Articolo su rivista]
Cignarella, G.; Barlocco, D.; Tranquillini, M. E.; Volterra, A.; Brunello, N.; Racagni, G.
abstract

A series of 3,8-diazabicyclo (3.2.1) octanes (DBO) (1) substituted at the nitrogen atoms by acyl and aralkenyl groups, were tested in in vitro binding assays towards μ and δ opioid receptors. The most representative terms (1a, 1d, 1g, 1j,) were also evaluated for the analgesic potency in vivo by the hot plate method. Among the compounds tested the most potent was the p.nitrocinnamyl DBO (1d) which displayed a μ/δ selectivity and an analgesic activity respectively 25 and 17 fold those of morphine. On the contrary, the m.hydroxycinnamyl DBO (1g) was markedly less active as agonist than the parent 1a, thus suggesting that structure 1 interacts with opioid receptors in a different fashion than morphine. Compound 1j isomer of 1a which is provided with high μ affinity, but lower analgesic potency, was found to possess a mixed agonist-antagonist activity. © 1988 The Italian Pharmacological Society.


1988 - Receptor mediated responses and intracellular mechanisms after antidepressant treatment [Articolo su rivista]
Brunello, N.; Rovescalli, A. C.; Perez, J.; Tinelli, D.; Trotti, D.; Cuomo, V.; Racagni, G.
abstract


1987 - Absence of [3H]SCH 23390 specific binding sites in anterior pituitary: Dissociation from effects on prolactin secretion [Articolo su rivista]
Rovescalli, A. C.; Brunello, N.; Monopoli, A.; Ongini, E.; Racagni, G.
abstract

We extended a previous study that had shown the selective D1 receptor antagonist SCH 23390, at relatively high doses, to stimulate prolactin (PRL) secretion in the rat and weakly inhibit [3H]spiperone binding to striatum and anterior pituitary (AP) membranes. No specific [3H]SCH 23390 binding sites, up to the micromolar range, were detected in rat AP while specific, saturable [3H]SCH 23390 binding sites (low nanomolar range) were observed in the striatum. In vivo SCH 23390 (1 mg/kg s.c.) induced higher plasma PRL levels, not reversible by the D1 agonist SKF 38393. Similarly the postsynaptic serotonin (5-HT) antagonists metergoline and cyproheptadine did not influence the SCH 23390 effect on PRL. SCH 23390 was also unable to antagonize the decrease of PRL secretion induced by the selective D2 agonist LY 171555. However this latter compound prevented SCH 23390 as well as sulpiride from increasing the PRL concentrations above the control values. These data rule out the possibility that D1 or 5-HT receptors mediate the stimulation of PRL release by SCH 23390. This effect is more likely to be due to weak indirect interaction with AP-D2 receptors, as indicated by the non-competitive inhibition of [3H]spiperone binding to AP exerted by SCH 23390. Alternatively, non-specific mechanism triggered by the multiple behavioral changes elicited by such high doses of SCH 23390 may be involved. © 1987.


1987 - EFFECT OF SOME TRICYCLIC AND NONTRICYCLIC ANTIDEPRESSANTS ON [3H]IMIPRAMINE BINDING AND SEROTONIN UPTAKE IN RAT CEREBRAL CORTEX AFTER PROLONGED TREATMENT [Articolo su rivista]
Brunello, N.; Riva, M.; Volterra, A.; Racagni, G.
abstract

Summary— Chronic administration of different antidepressant drugs reduced the number of [3H]imipramine ([3H]IMI) binding sites in rat cerebral cortex. In the same experimental conditions, fluvoxamine and dothiepin, as well as desmethylimipramine, induced an increase in the maximal velocity of high affinity serotonin (5HT) uptake in cortical slices, whereas citalopram and viloxazine were ineffective in this regard. Our results indicate that even if 5HT uptake and [3H]IMI binding sites are located on the same nerve terminals, they are differently modulated. Increased Vmax of the 5HT uptake process could be due to a rebound phenomenon after withdrawal from drugs that acutely inhibit 5HT uptake. The effect on [3H]IMI sites might be explained through either the agonist properties of the drugs towards these sites or the involvement of mechanisms still unknown. 1987 Société Française de Pharmacologie et de Thérapeutique


1987 - Endocrine effects of 5-methoxytryptoline, 5-hydroxytryptoline and tryptoline, putative modulators of rat serotonergic system [Articolo su rivista]
Rovescalli, A. C.; Brunello, N.; Racagni, G.
abstract

The effects of tryptolines (THβCs), putative endogenous compounds acting on the serotonergic function, have been studied on endocrine parameters in rats. In particular, graded doses of 5-methoxytryptoline (5-MeOT), 5-hydroxytryptoline (5-OHT) and tryptoline (Tp) were ip or iv administered to the animals and the circulating titers of prolactin (PRL), growth hormone (GH) and luteinizing hormone (LH) were assayed either at 20 min or at various times after the injection of the compounds. The data herein reported show that THβCs exert endocrine effects, at least in a pharmacological condition. However, the three compounds unequally affect anterior pituitary function. In fact, while 5-MeOT, 5-OHT and Tp all enhance plasma PRL concentrations in a quick, short lasting and dose-related manner, 5-MeOT and Tp induce also decreases of serum LH levels in ovariectomized rats, and 5-MeOT only is able to diminish plasma GH titers. These findings underline an endocrine effect for THβCs in pharmacological conditions and may suggest a functional role for these compounds in the control of anterior pituitary function. © 1987, Italian Society of Endocrinology (SIE). All rights reserved.


1986 - Interaction of putative endogenous tryptolines with the hypothalamic serotonergic system and prolactin secretion in adult male rats [Articolo su rivista]
Rovescalli, A. C.; Brunello, N.; Franzetti, C.; Racagni, G.
abstract

The effect of 5-methoxytryptoline (5-MeOT), 5-hydroxytryptoline (5-OHT) and tryptoline (Tp), putative endogenous derivatives of the tryptamines, on plasma prolactin (PRL) concentrations has been investigated in the adult male rat. The possible involvement of the hypothalamic serotonergic system has been considered in the mediation of the hormonal effect of the tryptolines. Therefore, plasma PRL levels have been evaluated in rats receiving tryptolines after different pharmacological manipulations of central serotonergic function. Although the three compounds increased the plasma titers of PRL in a dose-dependent manner and enhanced the hypothalamic content of serotonin (5HT), they appear to affect the serotonergic system through different mechanisms. In particular, 5-OHT might act at a presynaptic level, since its hyperprolactinemic effect was antagonized both by the depletion of central 5HT content after p-chlorophenylalanine and by the degeneration of serotonergic terminals after 5,7-dihydroxytryptamine. In contrast, 5-MeOT behaved as if it had a postsynaptic site of action, being counteracted by the serotonergic postsynaptic antagonists metergoline and cyproheptadine. The unsubstituted tetrahydro-β-carboline, Tp, is probably active at both pre- and postsynaptic sites. The enhancing effect of Tp on PRL secretion was antagonized by chronic treatment with p-chlorophenylalanine, while it was also maintained in 5,7-dihydroxytryptamine-lesioned rats. These findings suggest that tryptolines may play a functional role in PRL secretion by interacting with central serotonergic systems through different biochemical mechanisms. © 1986 S. Karger AG, Basel.


1986 - Interaction of β-casomorphins with multiple opioid receptors: In vitro and in vivo studies in the newborn rat brain [Articolo su rivista]
Volterra, A.; Restani, P.; Brunello, N.; Galli, C. L.; Racagni, G.
abstract

β-Casomorphins (β-CMs) are peptidic fragments of bovine β-casein with potent opioid activity. In view of a possible physiological meaning of these milk-derived compounds, we have studied the affinity of some natural β-CMs and some semisynthetic analogues for μ-, δ- and κ-brain opioid receptors in newborn and adult rat. Moreover we have investigated whether a chronic treatment with the potent analogue d-Ala2-β-CM-4-NH2 during the suckling period could affect μ and δ opioid receptor function. Our findings demonstrate that β-CMs are μ-oriented compounds both in adult and in newborn rat brain. They display the same μ-affinity in newborn as well as in adult animals, however δ and κ-affinities appear different, probably due to the lower degree of maturation of these two receptors in the first days of life. A prolonged treatment with d-Ala2-β-CM-4-NH2 during the preweanling period is able to induce a delay in the ontogenetic increase of δ-receptor affinity, whereas it affects neither the affinity nor the density of μ-receptors. This effect could be related to a general action of opioids on cerebral maturative processes; moreover we point out that a β-CMs analogue, given orally to newborn animals, can induce modifications at central level, suggesting thus the hypothesis that β-CMs could be a biologically active peptide in the first stages of life. © 1986 Elsevier Science Publishers B.V. (Biomedical Division) All rights reserved.


1986 - Interaction of β-casomorphins with multiple opioid receptors: In vitro and in vivo studies in the newborn rat brain [Articolo su rivista]
Volterra, A.; Restani, P.; Brunello, N.; Galli, C. L.; Racagni, G.
abstract

β-Casomorphins (β-CMs) are peptidic fragments of bovine β-casein with potent opioid activity. In view of a possible physiological meaning of these milk-derived compounds, we have studied the affinity of some natural β-CMs and some semisynthetic analogues for μ-, δ- and χ-brain opioid receptors in newborn and adult rat. Moreover we have investigated whether a chronic treatment with the potent analogue d-Ala2-β-CM-4-NH2 during the suckling period could affect μ and δ opioid receptor function. Our findings demonstrate that β-CMs are μ-oriented compounds both in adult and in newborn rat brain. They display the same μ-affinity in newborn as well as in adult animals, however δ and χ-affinities appear different, probably due to the lower degree of maturation of these two receptors in the first days of life. A prolonged treatment with d-Ala2-β-CM-4-NH2 during the preweanling period is able to induce a delay in the ontogenetic increase of δ-receptor affinity, whereas it affects neither the affinity nor the density of μ-receptors. This effect could be related to a general action of opioids on cerebral maturative processes; moreover we point out that a β-CMs analogue, given orally to newborn animals, can induce modifications at central level, suggesting thus the hypothesis that β-CMs could be a biologically active peptide in the first stages of life. © 1986.


1986 - Lack of serotonergic modulation on 3H-imipramine binding sites in basal conditions and during prolonged treatment with desmethylimipramine [Articolo su rivista]
Brunello, N.; Rovescalli, A. C.; Riva, M.; Racagni, G.
abstract


1986 - Ontogenetic studies on MU, delta and kappa opioid receptors in rat brain [Articolo su rivista]
Volterra, A.; Brunello, N.; Restani, P.; Galli, C. L.; Racagni, G.
abstract

The ontogenetic pattern of multiple opioid binding sites in rat brain from birth until weaning has been investigated. [3H]-dihydromorphine ([3H]-DHM)3 [3H]-D-Ala2-D-Leu5-enkephalin ([3H]-DADLE) and [3H]-dynorphin A (1-8) ([3H]-DYN) as markers of mu (μ), delta (δ) and Kappa (K) sites were utilized respectively. The analysis of the kinetic parameters of [3H]-DHM binding shows that, at birth, μ sites possess an affinity similar to that of adult animals, and a density of 50%, which reaches 80% of the adult value at day 4. On the contrary, [3H]-DADLE binding in the first post-natal days shows low affinity and low density and δ-sites do not reach values comparable to the adult ones until the second week of life. The kinetic parameters of [3H]-DYN binding are almost undetectable during the preweanling period, due to the very low density of K sites at this stage of life. Displacement studies with μ-, δ- and K-selective ligands show that the Ki values on [3H]-DHM binding sites were similar in 4 day old and adult animals for all the tested compounds, whereas Ki values on [3H]-DADLE and [3H]-DYN binding sites reflected an immaturity of δ and K receptors. In conclusion, our data suggest that multiple opioid receptors follow different ontogenetic patterns. In the first stages of life only μ receptors are almost mature and possibly mediate endogenous opioid actions and exogenous opiate pharmaco-toxicological effects. © 1986.


1986 - Variations of neuromediators in cerebral ageing. Effects of Ginkgo biloba extract [Articolo su rivista]
Racagni, G.; Brunello, N.; Paoletti, R.
abstract

Ginkgo biloba extract exerts a specific effect on the noradrenergic system and on beta-receptors. No variation was found in alpha 2-receptors and serotonin uptake. These findings provide the first evidence of central effects of a drug acting on cerebral ageing, connected specifically to reactivation of the noradrenergic system in the cerebral cortex.


1985 - Age-related changes in 5HT uptake and [3H]imipramine binding sites in rat cerebral cortex [Articolo su rivista]
Brunello, N.; Riva, M.; Volterra, A.; Racagni, G.
abstract


1985 - Biochemical and behavioral changes in rats after prolonged treatment with desipramine: interaction with p-chlorophenylalanine [Articolo su rivista]
Brunello, N.; Volterra, A.; Cagiano, R.; Ianieri, G. C.; Cuomo, V.; Racagni, G.
abstract

The decrease in the Bmax value of 3H-dihydroalprenolol (3H-DHA) binding to cortical membranes of rat brain induced by long-term administration of desipramine (DMI) was prevented by concomitant treatment with parachlorophenylalanine (pCPA). Acute administration of DMI significantly decreased locomotor activity in saline- and (pCPA)-pretreated rats. DMI-induced inhibition of locomotor activity was abolished in (pCPA)-pretreated rats chronically treated with DMI. Conversely, in pCPA-pretreated animals, acute DMI could still significantly decrease the locomotion of chronically DMI-treated rats. The data presented indicated that an intact serotoninergic system is required to enable antidepressant drugs to induce biochemical and behavioral changes following their chronic administration. © 1985 Springer-Verlag.


1985 - Comparative evaluation of the behavioural consequences of prenatal and early postnatal exposure to haloperidol in rats [Articolo su rivista]
Cuomo, V.; Cagiano, R.; Renna, G.; Serinelli, A.; Brunello, N.; Racagni, G.
abstract

The intensity of apomorphine (1-2 mg/kg)-induced stereotyped behaviour was significantly lower in 60 day old offspring of rats treated with haloperidol during pregnancy (0.5 mg/kg SC for 15 days, beginning on day 4 after conception) than in vehicle-exposed animals. Moreover, at 60 days of age, apomorphine (1-2 mg/kg)-induced inhibition of locomotor activity was much less marked in haloperidol-pretreated than in vehicle-pretreated rats. Conversely, our previous data showed that behavioural supersensitivity to apomorphine (1-2 mg/kg) was still present 40 days after the last administration of haloperidol, if the animals received the drug (0.5 mg/kg SC) from birth to 20 days of age. Finally, at 80 days of age, animals prenatally exposed to haloperidol were subjected to a differential reinforcement of low rates schedule (DRL-15 sec). The results indicate that the acquisition of the DRL task performance criterion (Rs/Rf≤2.5) was significantly more rapid in control animals than in haloperidol-pretreated rats. In this regard, we previously showed that early postnatal exposure to the neuroleptic also impairs the acquisition of the DRL schedule in adult rats. These data confirm and extend the differences in behavioural consequences of prenatal and early postnatal exposure to haloperidol, and further point to the usefulness of DRL task in the evaluation of subtle behavioural changes induced by psychotropic drugs in the absence of overt signs of neurotoxicity.


1985 - Effects of an extract of Ginkgo Biloba on noradrenergic systems of rat cerebral cortex [Articolo su rivista]
Brunello, N.; Racagni, G.; Clostre, F.; Drieu, K.; Braquet, P.
abstract

Oral treatment of rats withGinkgo biloba extract (Gb) elicited a biphasic effect on normetanephrine (NMN) content of cerebral cortex; an initial decrease was evident after 45 minutes, followed by a marked increase that was evident after 14 days. Chronic treatment with Gb led to decreases in the density of3H -dihydroalprenolol binding (after 27 days or 2 months) and in isoproterenol-stimulated adenylate cyclase activity (after 2 months) of the cerebral cortex. Taken together, these results indicate that the effects of Gb on the central β-adrenergic system might be involved in its therapeutic actions. © 1985 The Italian Pharmacological Society.


1985 - PRESYNAPTIC AND TRANSYNAPTIC MECHANISMS INVOLVED IN THE SUBSENSITIVITY OF RAT CORTICAL NORADRENERGIC SYSTEM AFTER LONG‐TERM ANTIDEPRESSANT TREATMENT [Articolo su rivista]
Racagni, G.; Brunello, N.; Mocchetti, I.; Cagiano, R.; Cuomo, V.
abstract

Abstract: Chronic treatment with antidepressants has been shown to produce a subsensitivity of noradrenergic neurons, both at presynaptic and postsynaptic sites. Important mechanisms, whereby the activity of noradrenergic neurons is regulated, could be the sensitivity of presynaptic alpha2‐adrenoceptors and the participation of transynaptic mechanisms involving other neurons. In this report we demonstrate that transynaptic factors involving the serotonergic system may be relevant to the regulation of the function of alpha2‐receptors in antidepressant chronically treated animals. In fact, we provide evidence of a markedly deminished responsiveness of noradrenergic neurons to an alpha2‐agonist (clonidine) or antagonist (mianserin) in biochemical and behavioral studies following serotonergic denervation with 5,7‐dihydroxytryptamine. These results indicate that a functional interrelationship between serotonergic and noradrenergic systems might play an important role in the adaptive changes which bring the noradrenergic neurons to a lower level of activity after chronic antidepressant administration. 1985 Nordic Pharmacological Society


1985 - Serotonergic modulation of cortical rat noradrenergic system in the mechanism of action of antidepressant drugs [Articolo su rivista]
Brunello, N.; Mocchetti, I.; Volterra, A.; Cuomo, V.; Racagni, G.
abstract


1984 - Behavioural and biochemical effects in C57BL/6J mice after a prolonged treatment with the δ‐opiate antagonist ICI 154129 [Articolo su rivista]
Volterra, A.; Brunello, N.; Cagiano, R.; Cuomo, V.; Racagni, G.
abstract

A long term treatment with the δ‐selective opiate antagonist NN‐bisallyl‐Tyr‐Gly‐Gly‐ψ‐(CH2S)‐Phe‐Leu‐OH (ICI 154129) produces an increase in the number of δ‐opiate binding sites, whereas the same treatment with the non selective opiate antagonist naloxone results in an enhancement of both μ‐ and δ‐binding sites. This biochemical effect in naloxone‐pretreated mice is paralleled by a more pronounced increase in locomotor activity induced by a challenge dose of morphine. In contrast, no difference in the effect of morphine was seen in ICI 154129‐pretreated mice with respect to control. These data suggest that the locomotor response to morphine in C57 mice is not mediated through δ‐opiate receptors. 1984 Royal Pharmaceutical Society of Great Britain


1984 - Behavioural and neurochemical changes in offspring of rats exposed to methyl mercury during gestation [Articolo su rivista]
Cuomo, V.; Ambrosi, L.; Annau, Z.; Cagiano, R.; Brunello, N.; Racagni, G.
abstract


1984 - Modulation of opioid system in C57 mice after repeated treatment with morphine and naloxone: Biochemical and behavioral correlates [Articolo su rivista]
Brunello, N.; Volterra, A.; Di Giulio, A. M.; Cuomo, V.; Racagni, G.
abstract

C57 BL/6J (C57) mice display a particular pattern of responses following morphine administration, such as a rapid development of tolerance to the pharmacological action of the opiate and an increase in locomotor activity after a single injection of the drug. We have measured met-enkephalin content and the responsiveness of different opiate receptors after repeated administration of morphine and naloxone. Prolonged morphine administration changed neither met-enkephalin levels, nor the density of opiate receptors in mice brain. In contrast repeated administration of the opiate antagonist naloxone, produced a marked increase in the number of 3H-DHM and 3H-DADLE binding sites in striatum and brainstem without modifying met-enkephalin concentrations. Behavioral studies have indicated that the morphine-induced increase in locomotor activity is enhanced in naloxone pretreated mice, thus suggesting a possible correlation between the behavioral response to morphine in C57 mice and the higher number of opiate receptors in the striatum. © 1984.


1984 - Transynaptic mechanisms in the action of antidepressant drugs [Articolo su rivista]
Racagni, G.; Brunello, N.
abstract

Antidepressant drugs act on different neuronal system and pre- and postsynaptic sites. Integrated transynaptic events are considered to be involved in those adaptive changes which seem to be operative after a prolonged administration. The authors explain how these long-term effects, rather than the acute pharmacological actions, are most likely to represent the biochemical mechanism underlying the delayed onset of antidepressant therapeutic efficacy. Among the possible mechanisms responsible for the adaptation of central aminergic neurons, interactions between serotonergic and noradrenergic systems, chemico-physical properties of the membranes and the modulatory actions of hormones and cotransmitters are considered. © 1984.


1983 - Behavioural changes after acute and chronic administration of typical and atypical antidepressants in rats: Interactions with reserpine [Articolo su rivista]
Cuomo, V.; Cagiano, R.; Brunello, N.; Fumagalli, R.; Racagni, G.
abstract

Acute administration of desipramine, mianserin and viloxazine produces a significant decrease of locomotor activity in rats. Conversely, the locomotion of rats receiving the antidepressants for a prolonged period of time is not modified with respect to control animals. Moreover, the reserpine-induced decrement of locomotion is not influenced by chronic treatment with any of the antidepressants used in the present study. Since tolerance develops rapidly to these behavioural effects they may be indicative of compensatory changes in neuronal systems which may occur during prolonged antidepressant treatment. © 1983.


1983 - On the mode of action of imipramine: Relationship between serotonergic axon terminal function and down-regulation of β-adrenergic receptors [Articolo su rivista]
Barbaccia, M. L.; Brunello, N.; Chuang, D. M.; Costa, E.
abstract

Recognition sites for [3H]imipramine and I3H]mianserin are located in different structures and regulate different neuronal functions. Recognition sites for [3H]imipramine are located on serotonergic terminals, are part of the supramolecular organization of the uptake mechanisms and can be down-regulated by prolonged administration of the drug. When the number of recognition sites for imipramine is down-regulated, uptake of 5-hydroxytryptamine (5HT) in rat brain hippocampal slices is increased. The presence of the binding sites for imipramine in 5HT terminals is essential to mediate the down-regulation of recognition sites for norepinephrine (NE) and NE-mediated stimulation of adenylate cyclase. Mianserin binds on a site that is modulated by 5HT, the number of its binding sites is not down-regulated by repeated treatment and, like imipramine, decreases the NE-dependent cyclase but nbt the number of β-adrenergic receptor recognition sites. Repeated treatment with imipramine and mianserin down-regulated the number of 5HT2 recognition sites. Several lines of evidence indicate that binding site for mianserin is related but not identical to the 5HT2 receptor binding site. © 1983.


1983 - Serotonin‐Elicited Amplification of Adenylate Cyclase Activity in Hippocampal Membranes from Adult Rat [Articolo su rivista]
Barbaccia, M. L.; Brunello, N.; Chuang, D. -M.; Costa, E.
abstract

The activity of the adenylate cyclase located in membranes prepared from hippocampus of adult rat can be stimulated by serotonin (5‐HT) (Ka= 4 x 10‐7M). The maximal effect is obtained with 10 μM 5‐HT. Freezing of the tissue decreases the 5‐HT stimulation; this stimulation is optimal in the presence of 82.5 mM Tris‐maleate buffer (pH 7.4) and 50 μM GTP. The adenylate cyclase activity of membranes prepared from cortex, hypothalamus, and colliculi of adult rats is not significantly stimulated by 5‐HT. Dopamine (DA) also stimulates adenylate cyclase located in hippocampal membranes; its effect can be blocked by haloperidol (10‐6 M), which fails to inhibit 5‐HT stimulation. Moreover, p‐chlorophenylalanine treatment for 2 weeks or selective lesion of 5‐HT axons afferent to the hippocampus increases the Vmax of 5‐HT stimulation, but fails to change that of DA stimulation. The 5‐HT stimulation can be inhibited by metergoline, spiroperidol, and pizotyline (10‐6 M), but not by the same concentrations of mianserin, ketanserine, alprenolol, phenoxybenzamine, and mepyramine. The 5‐HT stimulation of adenylate cyclase of hippocampal membranes can be mimicked by tryptamine, 5‐methoxytryptamine, bufotenine, and to a lesser extent by LSD; N‐methyltryptamine, N‐methyltryptophan, and 5‐hydroxytryptophan are inactive. Studies with kainic acid suggest that the 5‐HT recognition site (5‐HT1) linked to adenylate cyclase is located on the membrane of intrinsic hippocampal neurons. Copyright © 1983, Wiley Blackwell. All rights reserved


1983 - Temporal sequence of changes in central noradrenergic system of rat after prolonged antidepressant treatment: Receptor desensitization and neurotransmitter interactions [Articolo su rivista]
Racagni, G.; Mocchetti, I.; Calderini, G.; Battistella, A.; Brunello, N.
abstract

It has been shown that different receptor components may be involved in the adaptive changes occurring in noradrenergic (NE) neurones after prolonged periods of exposure to antidepressant drugs. In this report the desensitization of NE-coupled adenylate cyclase (NE-AC), β-adrenergic receptors and [3H]imipramine ([3H]-IMI or [3H]desipramine ([3H]DMI) binding sites have been temporally correlated with in vivo changes of NE utilization. Normetanephrine (NMN) was measured as indicator of NE synaptic events involved in antidepressant action. Concentrations of normetanephrine were increased after acute desipramine (DMI), viloxazine and mianserin administration. Following 3 days of treatment, the antidepressant-induced increase of normetanephrine became tolerant and NE neurones were resistant to the antidepressant effect until the 15th day of treatment. After two weeks, DMI elicited a significant decrease in the content of normetanephrine. A different pattern of changes has been found in the temporal modification of [3H]IMI recognition sites, β-adrenoceptors and NE-AC activity after chronic DMI treatment. Binding sites and receptors were down regulated after 10 days of treatment preceding the decrease in normetanephrine content. No down-regulation was observed in [3H]-DMI binding sites. Studies on the effects of antidepressants during brain maturation revealed that the mechanisms which cause desensitization of β-receptors and [3H]-IMI binding sites appear in the early stages of postnatal life. Since [3H]-IMI and [3H]-DMI recognition sites have been shown to be located on serotonergic (5-HT) and noradrenergic neurones respectively, the interactions between NE and 5-HT neurones could represent possible mechanisms implicated in receptor desensitization. The experiments presented involving lesions of 5-HT neurones have clearly demonstrated that NE release in rat cerebral cortex is under a tonic serotonergic influence. Alterations in the chemico-physical properties of the synaptic membranes might be also taken in consideration for the mechanisms underlying receptor modulation. In fact, evidence is provided that in neural tissue phospholipid methylation can be affected. In conclusion, the temporal sequence of changes in cortical noradrenergic neurones, after chronic antidepressant treatment, has demonstrated that integrated mechanisms are operative for the function of the overall system. © 1983.


1982 - Characterization of typical and atypical antidepressant recognition sites in rat brain [Articolo su rivista]
Brunello, N.; Chuang, D. M.; Costa, E.
abstract


1982 - Different synaptic location of mianserin and imipramine binding sites [Articolo su rivista]
Brunello, N.; Chuang, D. M.; Costa, E.
abstract

The high-affinity binding sites for mianserin and imipramine appear to be located in different neurons of rat brain. Studies in which lesions were produced with 5,7-dihydroxytryptamine and other studies in which the 5-hydroxytryptamine content was decreased with p-chlorophenylalanine indicate that some of the imipramine binding sites are on serotonin axon terminals and others are on nonserotonergic synapses. The sites that bind mianserin are on postsynaptic serotonin sites as well as on synapses of other neuronal systems. Copyright © 1982 AAAS.


1982 - Down-regulation of β-adrenergic receptors following repeated injections of desmethylimipramine: Permissive role of serotonergic axons [Articolo su rivista]
Brunello, N.; Barbaccia, M. L.; Chuang, D. -M.; Costa, E.
abstract

The injection of desmethylimipramine (DMI) twice daily for 3 weeks reduced the density of β-adrenergic receptor recognition sites located in crude synaptic membranes prepared from the cortex and hippocampus and attenuated the stimulation of the membrane-bound adenylate cyclase by isoproterenol. Both actions were abolished if prior to treatment with desmethylimipramine the serotonergic axons were destroyed by an intraventricular injection of 5,7-dihydroxytryptamine. These results show that the down-regulation of β-adrenergic receptors elicited by repeated injections of desmethylimipramine occurs only if the serotonergic axons are intact. © 1982.


1982 - Increase in Exogenous Choline Fails to Elevate the Content or Turnover Rate of Cortical, Striatal, or Hippocampal Acetylcholine [Articolo su rivista]
Brunello, N.; Cheney, D. L.; Costa, E.
abstract

Abstract: The present experiments were designed to test whether increasing the availability of choline to rat brain increases the rate of acetylcholine synthesis in that organ. The content of choline and acetylcholine and the turnover rate of acetylcholine in striatum, hippocampus, and cerebral cortex were measured following changes in dietary choline, intraperitoneal choline, or intravenous infusion of choline. Increasing plasma choline caused some increase in tissue choline but did not increase acetylcholine levels nor acetylcholine turn‐over rate in any of the areas of brain studied. Indeed, in hippocampus, choline decreased the turnover rate of acetylcholine. Copyright © 1982, Wiley Blackwell. All rights reserved


1982 - Neurochemical and pharmacogenetic aspects in the control of pain [Articolo su rivista]
Brunello, N.; Volterra, A.; Racagni, G.
abstract


1982 - Ontogenetic study of [3H]imipramine binding sites and serotonin uptake system: Indication of possible interdependence [Articolo su rivista]
Mocchetti, I.; Brunello, N.; Racagni, G.
abstract


1982 - Regulation of the high affinity binding sites for typical and atypical antidepressants in rat brain [Relazione in Atti di Convegno]
Ghuang, D. M.; Brunello, N.; Kinnier, W. J.; Costa, E.
abstract

We have characterized and studied the regulation of the high affinity binding sites in rat brain for a typical antidepressant (3H-imipramine) and an atypical antide-pressant (3H-mianserin). Repeated injections of imipramine for 10 to 14 days decreased the number of the binding sites for 3H-imipramine in the hippocampus but not in the cortex or cerebellum. Conversely, in the same rats the number of β-adrenergic receptor binding sites was down regulated in the cortex and cerebellum but not in the hippocampus. Available data indicates that down regulation of β-adrenergic receptors can be dissociated from the decrease in the number of imipramine binding sites. The binding sites for imipramine in hippocampal membranes were rapidly and irreversibly inactivated when the incubation temperature was raised from 0 to 37 °. Experiments using leupeptin and EGTA suggest that this temperature-induced reduction in the number of binding sites are due to an action of a Ca2+ dependent protease. Studies of the subcellular distribution indicates that 3H-imipramine binding sites are located in neurons. A destruction of serotonergic neurons by intracerebral injection of 5,7 dihydroxytryptamine led to a reduction in the number of 3H-imipramine binding sites in several brain areas examined; in contrast the number of 3H-mianserin binding sites was increased in some brain regions by the same treatment. Chronic treatment with p-Cl-phenylalanine increased the number of sites for mianserin but not for imipramine. Thus in serotonergic synapses, imipramine binding sites are in part located pre-synaptically whereas mianserin binding sites are in part located post-synaptically. © 1982.


1982 - Specific binding of [3H]mianserin and [3H]imipramine to structures of rat hippocampus [Articolo su rivista]
Brunello, N.; Chuang, D. -M.; Costa, E.
abstract


1982 - Studies on a neuronal-like transport system for serotonin in two cell lines [Articolo su rivista]
Vaghi, M.; Curatolo, L.; Algeri, S.; Brunello, N.; Mennini, T.; Morasca, L.; Salmona, A. L.; Mella, L.; Locati, M.; Dolfini, E.
abstract

The active transport of serotonin (5-HT) was studied in two cultured cell lines; FET (a finite fibroblast-like cell line derived from the osteomuscular structures of C3H embryo mice) and EUE (a continuous cell line derived from a human embryonal epithelium). These transport mechanisms are characterized by a high affinity carrier mediated system (km approx 1.3 uM and 1.0 uM respectively; Vmax approx 8 pmol/20 min × 106 cells and 2 pmol/20 min × 106 cells respectively) and temperature dependence. Fluoxetine (Lilly 110140) and Chlorimipramine preferentially block FET but not EUE serotonin uptake. Our results suggest that a serotonin-like transport system is present in unspecialized cultured cells, but it differs kinetically from the neuronal serotonin transport mechanism. © 1982 The Italian Pharmacological Society.


1982 - Use of specific brain lesions to study the site of action of antidepressants [Relazione in Atti di Convegno]
Brunello, N.; Chuang, D. M.; Costa, E.
abstract

We have shown that the binding sites for typical antidepressants such as imipramine and atypical antidepressants such as mianserin are not located exclusively in serotonergic synapses. To study the mechanisms involved we have decided to use the hippocampus as a model because of the possibility to elicit surgical lesion of specific pathways. Repeated treatment of rats with desipramine decreased the density of β-adrenergic receptors in the hippocampus and cortex. This receptor down regulation was prevented by lesioning the serotonin neurons with an intracerebral injection of 5,7 dihydroxytryptamine. Unilateral transection of the fimbria-fornix led to a reduction in the number of imipramine binding sites in the deafferented side of the hippocampus. Conversely, the number of the binding sites for mianserin was increased by this lesion. Intrahippocampal injection of kainic acid brought about a reduction in the number of mianserin recognition sites leaving intact the number of imipramine recognition sites. Therefore our present results are consistent with the hypothesis that imipramine and mianserin recognition sites are located in different synapses. However these synapses are functionally related because they appear to be located in a neuronal system that operates in connecting and functionally integrating serotonergic and adrenergic nerve terminals. © 1982.


1981 - Effects of immobilization and cold exposure on the turnover rate of acetylcholine in rat brain areas [Articolo su rivista]
Brunello, N.; Tagliamonte, A.; Chenky, D. L.; Costa, E.
abstract

The effect of two 'Stressors', immobilization and cold exposure, on the turnover rate of acetylcholine in various brain areas has been determined. Neither short term (10 min) nor long term (1 24 h) immobilization stress modifies the acetylcholine or choline concentrations or the turnover rate of acetylcholine in various rat brain areas. Cold exposure reduces the turnover rate of acetylcholine in the hypothalamus and modifies some of the other cholinergic parameters (e.g. choline content or fractional rate constant for acetylcholine efflux) in all areas studied. These results suggest that cholinergic neurons may be involved in thermoregulation but may not necessarily be involved in the phenomenon of immobilization stress. © 1981.


1981 - The septal-hippocampal cholinergic pathway: Role in antagonism of pentobarbital anesthesia and regulation by various afferents [Articolo su rivista]
Brunello, N.; Cheney, D. L.
abstract

Earlier studies have demonstrated that pentobarbital reduces the turnover rate of acetylcholine (ACh) in hippocampus and that this effect may be mediated via the septal-hippocampal cholinergic pathway. Moreover, the narcosis associated with the administration of pentobarbital may be reversed by intraseptal injection of such chemically diverse compounds as bicuculline, a potent γ-aminobutyric acid antagonist; thyrotropin-releasing hormone, a neuroactive tripeptide; and kainic acid, a rigid analog of glutamate. To determine whether or not these three compounds modulate the metabolism of ACh in hippocampus, they have been injected intraseptally in pentobarbital-pretreated rats and the turnover rate of ACh has been determined by gas chromatography-mass fragmentography. Pentobarbital produces a dose-dependent decrease in the turnover rate of ACh in cortex and hippocampus but not in striatum. The effect appears to be maximum at 30 min and returns to normal within 15 min of recovering the righting reflex. Slow local infusion of either bicuculline or thyrotropin-releasing hormone into the septum reverses the pentobarbital-induced narcosis and antagonizes the pentobarbital-induced decrease in the hippocampal turnover rate of ACh. Administration of kainic acid into the lateral, but not the medial, septum reduces specifically the glutamic acid decarboxylase activity in the ipsilateral septum without altering the choline acetyltransferase activity or the turnover rate of ACh in the hippocampus. Moreover, kainic acid injected into the lateral septum antagonizes the pentobarbital narcosis and reverses the pentobarbital-induced decrease in the ACh turnover rate in the ipsilateral hippocampus, but not in the contralateral hippocampus. It appears that all three compounds antagonize the pentobarbital-induced decrease in hippocampal ACh turnover rate and the pentobarbital narcosis by modulating neurons in the lateral septum, presumably through an action on the GABAergic interneurons.


1980 - Different adaptive responses by rat striatal dopamine synthetic and receptor mechanisms after repeated treatment with d-amphetamine, methylphenidate and nomifensine [Articolo su rivista]
Algeri, S.; Brunello, N.; Vantini, G.
abstract

The effect of repeated administration of d-amphetamine (AMP), methylphenidate (MP) and nomifensine (NOM) on dopamine receptors, striatal adenylate cyclase and tyrosine hydroxylase was studied. No one of the three stimulating drugs changed the receptor number or their affinity to haloperidol specific binding. Adenylate cyclase from rats treated with NOM showed a subsensitivity to the stimulation by dopamine. AMP treatment decreased the activity of striatal tyrosine hydroxylase. MP was ineffective in all the parameters studied. These results show that drugs believed to belong to the same class of stimulants are different in their effect on the CNS adaptive mechanisms.


1980 - Increased choline availability fails to increase acetylcholine turnover rate in rat brain areas [Articolo su rivista]
Cheney, D. L.; Brunello, N.; Costa, E.
abstract


1978 - Catecholamine synthesis in brain of ageing rat [Articolo su rivista]
Ponzio, F.; Brunello, N.; Algeri, S.
abstract


1978 - Effect of enkephalins on catecholamine metabolism in rat CNS [Articolo su rivista]
Algeri, S.; Brunello, N.; Calderini, G.; Consolazione, A.
abstract


1978 - The activity of some neurotransmitter-synthetizing enzymes in experimental cobalt epilepsy [Articolo su rivista]
Altamura, A. C.; Bonati, M.; Brunello, N.; Giordano, P. L.; Algeri, S.
abstract

The activities of tyrosine hydroxylase (TH), dihydropteridine reductase (DHPR), aromatic aminoacid decarboxylase (AAD) and glutamic acid decarboxylase (GAD), enzymes involved in the synthesis of neurotransmitters possibly implicated in epilepsy, were measured in some brain areas of rats with cobalt-induced epilepsy. Although TH, AAD and GAD were significantly decreased in the cortex of treated animals, the reduction was not correlated with the development of the electroclinical patterns of epilepsy. The reduction observed might be better explained by nerve degeneration caused by the experimental lesion. © 1978.


1977 - Dihydropteridine reductase and tyrosine hydroxylase activities in rat brain during development and senescence: a comparative study [Articolo su rivista]
Algeri, S.; Bonati, M.; Brunello, N.; Ponzio, F.
abstract