Tyra Zetterström

Fluoxetine-induced change in rat brain expression of brain-derived neurotrophic factor varies depending on length of treatment

Recent studies indicate that brain-derived neurotrophic factor (BDNF) may be implicated in the clinical action of antidepressant drugs. Repeated (2-3 weeks) administration of antidepressant drugs increases BDNF gene expression. The onset of this response as well as concomitant effects on the corresponding BDNF protein is however, unclear. The present study investigated the effects of acute and chronic administration of the selective serotonin reuptake inhibitor, fluoxetine (10mg/kg p.o.), upon regional rat brain levels of BDNF mRNA and protein expression. To improve the clinical significance of the study, fluoxetine was administered orally and mRNA and protein levels were determined ex vivo using the techniques of in situ hybridisation histochemistry and immunocytochemistry respectively. Direct measurement of BDNF protein was also carried out using enzyme-linked immunosorbent assay (ELISA). Four days of once daily oral administration of fluoxetine induced decreases in BDNF mRNA (hippocampus, medial habenular and paraventricular thalamic nuclei). Whilst 7 days of treatment showed a non-significant increase in BDNF mRNA, there were marked and region-specific increases following 14 days of treatment. BDNF protein levels remained unaltered until 21 days of fluoxetine treatment, when the numbers of BDNF immunoreactive cells were increased, reaching significance in the pyramidal cell layer of CA1 and CA3 regions of Ammons horn (CA1 and CA3) but not in the other sub-regions of the hippocampus. Indicative of the highly regional change within the hippocampus, the ELISA method failed to demonstrate significant up-regulation at 21 days, measuring levels of BDNF protein in the whole hippocampus. In contrast to the detected time dependent and biphasic response of the BDNF gene, activity-regulated, cytoskeletal-associated protein (Arc) mRNA showed a gradual increase during the 14-day course of treatment. The results presented here show that BDNF is expressed differentially depending on length of fluoxetine administration, which could contribute in explaining the slow onset of antidepressant activity observed with selective serotonin reuptake inhibitors.

Antidepressant drug treatment induces Arc gene expression in the rat brain.

The mechanism underlying the therapeutic effect of antidepressants is not known but neuroadaptive processes akin to long-term potentiation have been postulated. Arc (Activity-regulated, cytoskeletal-associated protein) is an effector immediate early gene implicated in LTP and other forms of neuroplasticity. Recent data show that Arc expression is regulated by brain 5-hydroxytryptamine neurones, a target of many antidepressants. Here in situ hybridisation and immunohistochemistry were used to examine whether Arc expression in rat brain is altered by antidepressant drug treatment. Repeated administration of the monoamine reuptake inhibitors paroxetine, venlafaxine or desipramine induced region-specific increases in Arc mRNA. These increases were greatest in regions of the cortex (frontal and parietal cortex) and hippocampus (CA1 layer) and absent in the caudate putamen. Repeated treatment with the monoamine oxidase inhibitor, tranylcypromine, increased Arc mRNA in a similar fashion to the monoamine reuptake inhibitors. The antidepressant drugs also increased the number of Arc-immunoreactive cells in the parietal cortex. Acute antidepressant injection, and repeated administration of the antipsychotic drug chlorpromazine, produced either limited or no changes in Arc mRNA. The data suggest that chronic treatment with antidepressant drugs induces Arc gene expression in specific regions across the rat forebrain. Up-regulation of Arc expression may be part of the process by which antidepressant drugs achieve long-term changes in synaptic function in the brain.

Acute onset by 5-HT6-receptor activation on rat brain brain-derived neurotrophic factor and activity-regulated cytoskeletal-associated protein mRNA expression

A number of previous studies have shown that chronic but not acute treatment with antidepressant drugs targeting the central 5-HT system, enhances mRNA expression for a number of genes including, brain-derived neurotrophic factor (BDNF) and the effector immediate early gene (IEG), activity-regulated, cytoskeletal-associated protein (Arc). The present study investigated the effects of 5-HT(6)-receptor activation on hippocampal and cortical levels of mRNA expression of BDNF and Arc in the rat. The selective 5-HT(6)-receptor agonist LY-586713 was administered acutely (0.1-10 mg/kg, s.c.) and mRNA levels of BDNF and Arc were measured 18 h later. Administration of LY-586713 caused a bell-shaped dose response on hippocampal BDNF mRNA expression, having no effect at 0.1 mg/kg, a significant up-regulation at 1 mg/kg and no effect at 10 mg/kg. The up-regulation in BDNF expression observed at 1 mg/kg was completely blocked by pre-treatment with the selective 5-HT(6)-receptor antagonist SB-271046 (10 mg/kg, s.c.). The effective dose (1 mg/kg) of LY-586713 on the induction of BDNF expression was also tested on Arc expression. Acute administration of LY-586713 at this dose caused marked increases of the Arc mRNA levels in cortical and hippocampal regions. These increases were also attenuated by SB-271046 (10 mg/kg) in all regions of the hippocampus, as well as the parietal cortex. However, in frontal cortical regions there was no attenuation by the antagonist. Moreover, SB-271046 alone increased Arc expression in these regions. The results presented here provide the first evidence for the involvement of the 5-HT(6) receptor in regulating BDNF and Arc mRNA expression, suggesting that LY-586713 has potential effects on neuronal plasticity. Overall, these findings suggest that, as opposed to more general 5-HT receptor activation by, for example, antidepressants, direct 5-HT(6)-receptor activation results in a more rapid rise in BDNF and Arc mRNA expression which does not require repeated administration.

Biphasic change in BDNF gene expression following antidepressant drug treatment explained by differential transcript regulation.

Brain-derived neurotrophic factor (BDNF) has been suggested as a possible target for the treatment of depression. The effect by antidepressant drugs on BDNF mRNA expression is, however, strictly dependent on both treatment duration and time after the last administration. The rat BDNF gene itself is complex and expresses four different mRNA isoforms which can be regulated by different signaling cascades. The aim of the present study was to test the hypothesis that the previously shown biphasic action by the antidepressant drugs on total BDNF expression is explained by differential BDNF transcript regulation. For this purpose, we used in situ hybridization with exon-specific oligo nucleotides for exon V (total BDNF mRNA), exon I (protein synthesis-dependent transcripts), exon III and exon IV (immediate early-gene like-transcripts). Following an acute injection, all three drugs tested: fluoxetine, desipramine and TCP decreased total BDNF mRNA (exon V) as well as exon IV mRNA, while no significant effect was recorded for exons I and III mRNAs. In contrast chronic administration of all three drugs resulted in increased expression of exon V- and exon I-containing transcripts (fluoxetine and TCP only) but no significant changes were recorded for exon III and IV mRNAs. Electroconvulsive shock administration showed up-regulation of all four BDNF mRNAs following a single shock, but after repeated administration increases were restricted to exons I- and V-containing transcripts. In summary, this study shows clear evidence of differential BDNF transcript regulation following acute and chronic antidepressant drug treatment.

Differential regulation of psychostimulant-induced gene expression of brain derived neurotrophic factor and the immediate-early gene Arc in the juvenile and adult brain

Psychostimulant drugs are widely used in children for the treatment of attention‐deficit/hyperactivity disorder. Recent animal studies have suggested that exposure to these agents in early life could be detrimental to brain development. Here, for the first time, the effect of methylphenidate (MPH) and d‐amphetamine (AMPH) on the expression of two key genes for neuronal development and plasticity, brain‐derived neurotrophic factor (bdnf) and the effector immediate early gene activity‐regulated, cytoskeletal‐associated protein (Arc), was examined in both juvenile and adult rats. Both MPH [2 mg/kg, intraperitoneal (i.p.)] and AMPH (0.5 mg/kg, i.p.) induced marked decreases of bdnf mRNA in hippocampal and cortical brain regions of juveniles, whereas effects in adults were significantly less (hippocampus) or opposite (frontal cortex). In comparison, Arc mRNA was decreased (hippocampus and parietal cortex), largely unaffected (frontal cortex) or increased (striatum) in juveniles, whereas in adults, Arc mRNA increased in most brain regions. MPH‐induced locomotion was also measured, and showed a much smaller increase in juveniles than in adults. In summary, our data show that the effects of MPH and AMPH on expression of the neurodevelopmentally important genes, bdnf and Arc, differ markedly in juvenile and adult rats, with juveniles showing evidence of brain region‐specific decreases in both genes. These age‐dependent effects on gene expression may be linked with the reported long‐term harmful effects of psychostimulants in animal models.

Age-dependent effects of methylphenidate in the prefrontal cortex: evidence from electrophysiological and Arc gene expression measurements

Methylphenidate, a drug widely used for attention deficit hyperactivity disorder in children, may affect neuronal function differently in young and adult subjects, particularly in the prefrontal cortex, a brain structure that does not fully develop until adulthood. We compared the impact of development on the effects of methylphenidate on single unit electrical activity and mRNA expression of the effector immediate early gene activity-regulated cytoskeletal-associated protein (Arc) following methylphenidate in the prefrontal cortex in adult (more than 60 days old) and juvenile (25—35 days old) rats. Methylphenidate, administered under urethane anaesthesia to adult rats, at doses ranging from 1 mg/kg to 3 mg/kg intravenously, exerts a progressive activation of firing of prefrontal cortex neurones (30% to 84% from baseline). This activation was significantly lower in the juvenile rats, reaching only 37% of baseline levels at the highest dose (3 mg/kg, intravenous). In adults, methylphenidate (4 mg/kg intraperitoneal) produced marked increases in Arc mRNA levels compared with saline controls by 123% and 164% in cingulated and orbital cortex, respectively. Corresponding values for the juvenile rats were significantly lower (42% and 79%). In summary, this multi-approach investigation showed that the reactivity of prefrontal cortex neurones to methylphenidate differs markedly in juvenile and adult rats.

MIF (Pro-Leu-Gly-NH2): failure to affect oxotremorine effects in mice and rats as well as fluphenazine catalepsy or amphetamine hyperactivity in rats.

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Chronic methylphenidate regulates genes and proteins mediating neuroplasticity in the juvenile rat brain

Methylphenidate (MPH) is the front-line psychostimulant medication prescribed for alleviating the symptoms associated with attention deficit hyperactivity disorder (ADHD) in children. Here, we investigated the effects of chronic MPH (2.0mg/kg, twice daily for 15days) exposure to young rats (20-25days old at start of treatment) on the expression of genes and proteins associated with neuroplasticity, such as activity regulated cytoskeleton-associated protein (Arc), insulin receptor substrate protein 53 (IRSp53), cell division control protein 42 (Cdc42), and actin-related protein 2 (Arp2). Chronic MPH increased Arc expression in areas of the cerebrum including, the striatum, nucleus accumbens and hippocampus. In addition, chronic MPH also increased the expression of IRSp53 in the striatum, while Cdc42 and Arp2 were specifically increased in the nucleus accumbens. Conversely, chronic MPH decreased Arc and IRSp53 protein expression in the cerebellum, indicating differential effects of the drug in cerebral areas relative to the cerebellum. Overall, our results indicate that chronic MPH treatment increases expression of genes and proteins associated with dendritic spine formation and neuronal plasticity in target areas of the cerebrum while it decreases the expression in the cerebellum.

The role of 5-hydroxytryptamine receptor subtypes in the regulation of brain-derived neurotrophic factor gene expression

The study aims to investigate the role of 5‐hydroxytryptamine receptor subtypes in mediating the inhibitory effect of the selective serotonin reuptake inhibitor (fluoxetine on brain‐derived neurotrophic factor gene (bdnf) expression in rat hippocampus.

Chapter 1.1 What did we learn from microdialysis

Abstract At the beginning of 1980s, microdialysis emerged as a novel method for monitoring brain neurochemistry in vivo. This was followed by a rapid, worldwide uptake of microdialysis and an explosion of papers reporting the application of the technique across a broad range of neuroscience research areas. This article discusses the historical background to in vivo neurochemical monitoring, and sets out the techniques available prior to the first microdialysis studies. Detail is then given of some of the events and people involved in one of the laboratories that played a central role in the development of microdialysis and the dissemination of this technique to the wider neuroscience community. Finally, some of the early experiments are outlined and used to illustrate a few of the many contributions that microdialysis has made to present-day neuroscience.