Behavioral and transcriptome alterations in male and female mice with postnatal deletion of TrkB in dorsal striatal medium spiny neurons.

BACKGROUND: The high affinity tyrosine kinase receptor, TrkB, is the primary receptor for brain derived neurotrophic factor (BDNF) and plays an important role in development, maintenance and plasticity of the striatal output medium size spiny neuron. The striatal BDNF/TrkB system is thereby implicated in many physiologic and pathophysiologic processes, the latter including mood disorders, addiction, and Huntington's disease. We crossed a mouse harboring a transgene directing cre-recombinase expression primarily to postnatal, dorsal striatal medium spiny neurons, to a mouse containing a floxed TrkB allele (fB) mouse designed for deletion of TrkB to determine its role in the adult striatum. RESULTS: We found that there were sexually dimorphic alterations in behaviors in response to stressful situations and drugs of abuse. Significant sex and/or genotype differences were found in the forced swim test of depression-like behaviors, anxiety-like behaviors on the elevated plus maze, and cocaine conditioned reward. Microarray analysis of dorsal striatum revealed significant dysregulation in individual and groups of genes that may contribute to the observed behavioral responses and in some cases, represent previously unidentified downstream targets of TrkB. CONCLUSIONS: The data point to a set of behaviors and changes in gene expression following postnatal deletion of TrkB in the dorsal striatum distinct from those in other brain regions.

Cell-autonomous alteration of dopaminergic transmission by wild type and mutant (DeltaE) TorsinA in transgenic mice.

Early onset torsion dystonia is an autosomal dominant movement disorder of variable penetrance caused by a glutamic acid, i.e. DeltaE, deletion in DYT1, encoding the protein TorsinA. Genetic and structural data implicate basal ganglia dysfunction in dystonia. TorsinA, however, is diffusely expressed, and therefore the primary source of dysfunction may be obscured in pan-neuronal transgenic mouse models. We utilized the tyrosine hydroxylase (TH) promoter to direct transgene expression specifically to dopaminergic neurons of the midbrain to identify cell-autonomous abnormalities. Expression of both the human wild type (hTorsinA) and mutant (DeltaE-hTorsinA) protein resulted in alterations of dopamine release as detected by microdialysis and fast cycle voltammetry. Motor abnormalities detected in these mice mimicked those noted in transgenic mice with pan-neuronal transgene expression. The locomotor response to cocaine in both TH-hTorsinA and TH-DeltaE-hTorsinA, in the face of abnormal extracellular DA levels relative to non-transgenic mice, suggests compensatory, post-synaptic alterations in striatal DA transmission. This is the first cell-subtype-specific DYT1 transgenic mouse that can serve to differentiate between primary and secondary changes in dystonia, thereby helping to target disease therapies.

Low dose naltrexone administration in morphine dependent rats attenuates withdrawal-induced norepinephrine efflux in forebrain.

The administration of low dose opioid antagonists has been explored as a potential means of detoxification in opiate dependence. Previous results from our laboratory have shown that concurrent administration of low dose naltrexone in the drinking water of rats implanted with subcutaneous morphine pellets attenuates behavioral and biochemical signs of withdrawal in brainstem noradrenergic nuclei. Noradrenergic projections originating from the nucleus tractus solitarius (NTS) and the locus coeruleus (LC) have previously been shown to be important neural substrates involved in the somatic expression of opiate withdrawal. The hypothesis that low dose naltrexone treatment attenuates noradrenergic hyperactivity typically associated with opiate withdrawal was examined in the present study by assessing norepinephrine tissue content and norepinephrine efflux using in vivo microdialysis coupled to high performance liquid chromatography (HPLC) with electrochemical detection (ED). The frontal cortex (FC), amygdala, bed nucleus of the stria terminalis (BNST) and cerebellum were analyzed for tissue content of norepinephrine following withdrawal in morphine dependent rats. Naltrexone-precipitated withdrawal elicited a significant decrease in tissue content of norepinephrine in the BNST and amygdala. This decrease was significantly attenuated in the BNST of rats that received low dose naltrexone pre-treatment compared to controls. No significant difference was observed in the other brain regions examined. In a separate group of rats, norepinephrine efflux was assessed with in vivo microdialysis in the BNST or the FC of morphine dependent rats or placebo treated rats subjected to naltrexone-precipitated withdrawal that received either naltrexone in their drinking water (5 mg/L) or unadulterated water. Following baseline dialysate collection, withdrawal was precipitated by injection of naltrexone and sample collection continued for an additional 4 h. At the end of the experiment, animals were transcardially perfused and the brains were removed for verification of probe placement. Low dose naltrexone pre-treatment significantly attenuated withdrawal-induced increases of extracellular norepinephrine in the BNST, with a smaller effect in the FC. These findings suggest that alterations in norepinephrine release associated with withdrawal may be attenuated in forebrain targets of noradrenergic brainstem neurons that may underlie reduced behavioral signs of withdrawal following low dose naltrexone administration.

The effect of corticotropin-releasing factor on prepulse inhibition is independent of serotonin in Brown Norway and Wistar-Kyoto rats.

Prepulse inhibition (PPI), a form of sensorimotor gating, is reduced in a number of psychiatric disorders. Two experiments were conducted to determine whether corticotropin-releasing factor (CRF), which decreases PPI, does so via effects on serotonin (5-HT). Wistar-Kyoto (WKY) and Brown Norway (BN) rats were used in both experiments in order to examine whether strain-dependent differences would be apparent in response to manipulations of the CRF and 5-HT systems. In the first experiment, WKY and BN rats received a subcutaneous injection of the 5-HT(2A/C) receptor antagonist, ketanserin (2.0 mg/kg). Ten minutes later, rats received an intracerebroventricular (ICV) infusion of either 6.0 microl saline or CRF (0.3 microg or 3.0 microg). CRF decreased PPI despite blockade of 5-HT(2A/C) receptors with ketanserin. In the second experiment, WKY and BN rats received an intraperitoneal injection of the 5-HT synthesis inhibitor, p-chlorophenylalanine (PCPA, 150 mg/kg), 48 and 24 h prior to testing. On testing day, rats received an ICV infusion of either 6.0 microl saline or CRF (0.3 microg or 3.0 microg). CRF decreased PPI despite 5-HT depletion. These findings suggest that CRF does not decrease PPI via effects on 5-HT, since neither blockade of 5-HT(2A/C) receptors nor 5-HT depletion attenuated this decrease.

Depletion of serotonin and catecholamines block the acute behavioral response to different classes of antidepressant drugs in the mouse tail suspension test.

RATIONALE: Few studies have investigated whether the behavioral effects elicited by different types of antidepressant drugs are mediated by either serotonin (5-HT) or the catecholamines norepinephrine (NE) and dopamine (DA). OBJECTIVES: By depleting 5-HT, or NE and DA, the present study investigated the contributions of these monoamines to the acute behavioral effects of selective serotonin reuptake inhibitors (SSRIs; fluoxetine and citalopram) and norepinephrine reuptake inhibitors (NRIs; desipramine and reboxetine) in the mouse tail suspension test (TST). RESULTS: Depletion of 5-HT tissue content by para-chlorophenylalanine (PCPA), an inhibitor of tryptophan hydroxylase, completely blocked reductions of immobility by the SSRIs in the TST. In contrast, PCPA did not alter the behavioral effects of the NRIs. Inhibition of catecholamine synthesis by alpha-methyl-para-tyrosine (AMPT) reduced brain NE and DA tissue content, whereas disruption of vesicular storage with reserpine decreased brain NE, DA and 5-HT tissue content. However, neither treatment completely prevented responses to desipramine, fluoxetine, or citalopram in the TST. Depleting both newly synthesized and vesicular components of NE and DA transmission with a combination of reserpine and AMPT completely prevented the behavioral effects of desipramine, reboxetine, and fluoxetine and attenuated those of citalopram. Although PCPA did not alter baseline immobility, AMPT and reserpine increased baseline values in the TST. CONCLUSIONS: These studies demonstrated that endogenous 5-HT synthesis mediates the behavioral effects of SSRIs, but not NRIs, in the TST. In contrast, disruption of the behavioral effects of NRI and SSRI antidepressants required disruption of both catecholamine synthesis and vesicular storage and release mechanisms.

Locus ceruleus regulates sensory encoding by neurons and networks in waking animals.

Substantial evidence indicates that the locus ceruleus (LC)-norepinephrine (NE) projection system regulates behavioral state and state-dependent processing of sensory information. Tonic LC discharge (0.1-5.0 Hz) is correlated with levels of arousal and demonstrates an optimal firing rate during good performance in a sustained attention task. In addition, studies have shown that locally applied NE or LC stimulation can modulate the responsiveness of neurons, including those in the thalamus, to nonmonoaminergic synaptic inputs. Many recent investigations further indicate that within sensory relay circuits of the thalamus both general and specific features of sensory information are represented within the collective firing patterns of like-modality neurons. However, no studies have examined the impact of NE or LC output on the discharge properties of ensembles of functionally related cells in intact, conscious animals. Here, we provide evidence linking LC neuronal discharge and NE efflux with LC-mediated modulation of single-neuron and neuronal ensemble representations of sensory stimuli in the ventral posteriomedial thalamus of waking rats. As such, the current study provides evidence that output from the LC across a physiologic range modulates single thalamic neuron responsiveness to synaptic input and representation of sensory information across ensembles of thalamic neurons in a manner that is consistent with the well documented actions of LC output on cognition.

Differential behavioral effects of the antidepressants reboxetine, fluoxetine, and moclobemide in a modified forced swim test following chronic treatment.

RATIONALE: The forced swim test (FST) is the most widely used model for assessing potential antidepressant activity in rodents following acute or short-term treatment. However, few studies have compared the effects of short- and long-term antidepressant treatment on behaviors in the test, despite the need to treat patients chronically to produce clinical effects. OBJECTIVES: The current studies examined whether antidepressants from different classes produce different behavioral effects following short-term treatment and whether such effects change following administration for a longer duration. METHODS: The effects of administering short-term (3 days) and long-term (14 days) treatments of antidepressants from three different chemical classes with distinct mechanisms of action via osmotic minipump were examined: the selective norepinephrine reuptake inhibitor reboxetine (10 and 60 mg kg(-1) day(-1)), the selective serotonin reuptake inhibitor fluoxetine (2.5 and 15 mg kg(-1) day(-1)), and the reversible inhibitor of monoamine oxidase moclobemide (2.5 and 15 mg kg(-1) day(-1)). All testing was carried out in a 15-min test with no preswim session in order to negate any confounding aspect of an induction procedure. RESULTS: The majority of antidepressant-sensitive behavioral changes were observed in the first 5 min of the test. The low dose of reboxetine failed to alter behavior in the test after 3 days but significantly decreased immobility and increased climbing behavior following administration for 14 days, whereas the high dose of reboxetine was equally effective following 3 and 14 days of treatment. In a similar fashion, the low dose of fluoxetine failed to alter behavior in the test following 3 days, but showed an augmented response on immobility and increased swimming following administration for 14 days. The high dose of fluoxetine was slightly more effective at reducing immobility following administration for 14 days than 3 days. The low dose of moclobemide decreased immobility and increased climbing behavior following treatment for 3 days, but increases in both swimming and climbing behaviors were measured following treatment for 14 days. Treatment with the high dose of moclobemide for 3 days decreased immobility and increased swimming, whereas treatment for 14 days significantly increased both active behaviors (swimming and climbing). CONCLUSIONS: Antidepressants from three different classes produce different effects on active behaviors in the FST. The effects of antidepressants were augmented following chronic administration for 14 days, especially when given at low doses. This suggests that modifications of the FST can be used to examine the onset of action of antidepressant agents produced by long-term administration.

Blockade of the mGlu5 receptor decreases basal and stress-induced cortical norepinephrine in rodents.

RATIONALE: Glutamate, the major excitatory neurotransmitter in the brain mediates its effects by both ionotropic and metabotropic receptor subtypes. Recently, the search for selective ligands for glutamate receptor subtypes has led to the discovery of 2-methyl-6-(phenylethynyl)pyridine (MPEP), an antagonist specific for metabotropic glutamate receptor 5 (mGlu5). This receptor is highly expressed in limbic forebrain regions and is thought to modulate anxiety-related processes. The noradrenergic nucleus locus coeruleus (LC) is an important mediator of stress responses and dysfunction of this system is implicated in affective disorders such as anxiety and depression. OBJECTIVES: We sought to assess the effects of mGlu5 receptor antagonists, MPEP and 3-[(2-methyl-1,3-thiazol-4-yl)ethynyl]pyridine (MTEP) on cortical norepinephrine (NE) levels. METHODS: In vivo microdialysis and high-pressure liquid chromatography with electrochemical detection (HPLC-ED) were used to assess the effects of mGlu5 antagonism on extracellular NE in the frontal cortex, a major terminal field of the LC. RESULTS: Blockade of the mGlu5 receptor elicited significant reductions in extracellular NE in the frontal cortex. The benzodiazepine diazepam also reduced cortical NE. Furthermore, MPEP administration attenuated stress-induced increases in extracellular NE. CONCLUSIONS: Taken together, these data show that MPEP and MTEP, through their blockade of the mGlu5, reduce extracellular norepinephrine, the impact of which may contribute to their anxiolytic actions.

Norepinephrine-deficient mice lack responses to antidepressant drugs, including selective serotonin reuptake inhibitors.

Mice unable to synthesize norepinephrine (NE) and epinephrine due to targeted disruption of the dopamine beta-hydroxylase gene, Dbh, were used to critically test roles for NE in mediating acute behavioral changes elicited by different classes of antidepressants. To this end, we used the tail suspension test, one of the most widely used paradigms for assessing antidepressant activity and depression-related behaviors in normal and genetically modified mice. Dbh(-/-) mice failed to respond to the behavioral effects of various antidepressants, including the NE reuptake inhibitors desipramine and reboxetine, the monoamine oxidase inhibitor pargyline, and the atypical antidepressant bupropion, even though they did not differ in baseline immobility from Dbh(+/-) mice, which have normal levels of NE. Surprisingly, the effects of the selective serotonin reuptake inhibitors (SSRIs) fluoxetine, sertraline, and paroxetine were also absent or severely attenuated in the Dbh(-/-) mice. In contrast, citalopram (the most selective SSRI) was equally effective at reducing immobility in mice with and without NE. Restoration of NE by using L-threo-3,4-dihydroxyphenylserine reinstated the behavioral effects of both desipramine and paroxetine in Dbh(-/-) mice, thus demonstrating that the reduced sensitivity to antidepressants is related to NE function, as opposed to developmental abnormalities resulting from chronic NE deficiency. Microdialysis studies demonstrated that the ability of fluoxetine to increase hippocampal serotonin was blocked in Dbh(-/-) mice, whereas citalopram's effect was only partially attenuated. These data show that NE plays an important role in mediating acute behavioral and neurochemical actions of many antidepressants, including most SSRIs.

The promises and pitfalls of reboxetine.

The antidepressant compound, morpholine, 2-[(2-ethoxyphenoxy)phenylmethyl]-,methanesulfonate, or reboxetine, is a selective noradrenergic reuptake inhibitor that acts by binding to the norepinephrine (NE) transporter and blocking reuptake of extracellular NE back into terminals. This compound has low affinity for other transporters and receptors. The development of reboxetine as a potential antidepressant stems from the prior demonstration that blockade of the NE transporter imparts antidepressant activity. Desipramine, lofepramine, and nortryptiline are examples of tricyclic antidepressant (TCA) compounds from the first generation of antidepressants that exert their effects by blockade of NE reuptake. Maprotiline, a non-tricyclic compound, is also a NE selective reuptake inhibitor. Unfortunately, these antidepressants are also associated with interactions with muscarinic, histaminergic, and adrenergic receptors, which are known to contribute to a variety of untoward side effects. Despite the positive pharmacological profile of reboxetine, i.e., selectivity and specificity, with relatively fewer side effects, its use as an antidepressant is currently limited to Europe. Reboxetine is marketed as Edronax in the UK, Norebox in Italy, and as Irenor in Spain. It is registered in Germany, Sweden, Denmark, Ireland, Austria and Finland. Based on studies conducted primarily outside the US, the FDA granted a preliminary letter of approval in 1999. However, more recent clinical studies conducted in the US and Canada, prompted by the FDA, resulted in a letter of non-approval. To date, it is unclear why the further development of reboxetine as an antidepressant in the US has been halted. Despite this setback, reboxetine has been a valuable pharmacological tool to assess the role of the noradrenergic system in preclinical studies of depressive disorder.

Simultaneous analyses of the neurochemical and behavioral effects of the norepinephrine reuptake inhibitor reboxetine in a rat model of antidepressant action.

RATIONALE: The forced swimming test (FST) is a rodent behavioral assay widely used to predict clinical efficacy of putative antidepressants. Few studies have examined the effects of the FST on neurotransmitter levels and how antidepressant drug treatment may alter neurotransmitter levels and behavior simultaneously during the performance of a stressful task. OBJECTIVES: The present study examined the role of norepinephrine in mediating active behaviors in the FST after treatment with reboxetine, a selective norepinephrine reuptake inhibitor. METHODS: High-pressure liquid chromatography was used to analyze microdialysis samples collected from awake, freely moving rats before, during and after exposure to the FST. Reboxetine (10 mg/kg) was given three times over a 24-h period prior to the test swim. Behavioral responses, including immobility, swimming and climbing, were counted during the 5-min test on day 1 and day 2. RESULTS: The first exposure to swim stress elicited a 65% increase in extracellular norepinephrine (NE). A second exposure on day 2 elicited a 52% increase of NE and a behavioral profile characterized by increased immobility and a reduction of active behaviors. A subchronic course (three injections over 24 h) of treatment with reboxetine between the two swim exposures resulted in antidepressant-like activity, i.e., decreased immobility and increased climbing behavior on day 2. A significantly greater increase in extracellular NE (112%) was observed in the group of animals that received reboxetine injections. CONCLUSIONS: Treatment with reboxetine in a schedule commonly used in the FST resulted in a potentiated noradrenergic response to the swim challenge concomitant with behavioral alterations consistent with antidepressant-like activity.

Behavioral and neurochemical effects of 5-(4-[4-(5-Cyano-3-indolyl)-butyl)-butyl]-1-piperazinyl)-benzofuran-2-carboxamide (EMD 68843): a combined selective inhibitor of serotonin reuptake and 5-hydroxytryptamine(1A) receptor partial agonist.

5-(4-[4-(5-Cyano-3-indolyl)-butyl)-butyl]-1-piperazinyl)-benzofuran-2-carboxamide (EMD 68843; vilazodone) is a novel compound with combined high affinity and selectivity for the 5-hydroxytryptamine (5-HT) transporter and 5-HT(1A) receptors. EMD 68843 was tested as a prototype compound, which benefits from dual pharmacological effects that could increase extracellular 5-HT to levels higher than those produced by conventional selective serotonin reuptake inhibitors (SSRIs). In Sf9 cells, EMD 68843 increased guanosine 5'-O-(3-[(35)S]thiotriphosphate) binding to 69% of the magnitude of the full 5-HT(1A) receptor agonist R-(1)-trans-8-hydroxy-2-[N-n-propyl-N-(39-iodo-29-propenyl)] aminotetralin (8-OH-PIPAT), indicating that it is a partial agonist at 5-HT(1A) receptors. Acute, systemic administration of EMD 68843 produced a larger maximal increase of extracellular 5-HT than the SSRI fluoxetine in both the ventral hippocampus (HPv) (558 versus 274%) and the frontal cortex (FC) (527 versus 165%). Regional differences in the response to the two drugs were also observed. These effects may be attributed to the differential regulation of 5-HT release in the HPv and FC by 5-HT(1A) autoreceptors. When challenged with the 5-HT(1A) receptor agonist 8-hydroxy-2-(di-n-propylamino) tetralin (8-OH-DPAT), EMD 68843-induced increases in extracellular 5-HT were greatly reduced in the HPv but to a lesser extent in the FC. In behavioral studies, EMD 68843 produced antidepressant-like effects in the forced swimming test in both rats and mice but only within a narrow dosage range. Like fluoxetine, EMD 68843 did not produce the symptoms of the 5-HT behavioral syndrome in rats but, unlike fluoxetine, pretreatment with EMD 68843 blocked expression of the 5-HT behavioral syndrome induced by 8-OH-DPAT. Taken together, the results show that EMD 68843 augments extracellular 5-HT levels in forebrain regions to a greater extent than fluoxetine. At higher doses, however, weak efficacy of EMD 68843 at postsynaptic 5-HT(1A) receptors may inhibit the expression of rodent antidepressant-like behaviors.

Effects of acute and chronic reboxetine treatment on stress-induced monoamine efflux in the rat frontal cortex.

Reboxetine is a selective noradrenergic reuptake inhibitor that displays an antidepressant profile in both animal tests and in clinical trials. The present study examined the ability of reboxetine to alter stress-induced increases in norepinephrine, serotonin and dopamine efflux in the frontal cortex in awake behaving rats. Acute systemic administration of reboxetine (0.3-20.0 mg/kg) dose-dependently increased extracellular norepinephrine in the frontal cortex while having no effect on extracellular serotonin. At 20 mg/kg, reboxetine also increased extracellular dopamine. Application of a 20-min tailpinch stress increased extracellular norepinephrine. This effect was greatly potentiated in rats pretreated with reboxetine. Tailpinch did not elicit increases in dopamine in saline treated animals but this stimulus increased dopamine levels following reboxetine pretreatment. Furthermore, chronic administration of reboxetine for 14 days resulted in elevated basal concentrations of extracellular norepinephrine and dopamine and a greater net increase of extracellular norepinephrine and dopamine, but not serotonin, in response to tailpinch compared with vehicle control animals. Taken together, these data support the view that the noradrenergic and dopaminergic systems are modified by reboxetine treatment and may be important factors in the mechanism of action of antidepressant compounds.

Noradrenergic lesions differentially alter the antidepressant-like effects of reboxetine in a modified forced swim test.

The novel antidepressant reboxetine is a selective norepinephrine reuptake inhibitor. In this study, the antidepressant-like effects of reboxetine were characterized in a modified rat forced swim test. Further, in order to investigate the role of the locus coeruleus and lateral tegmental noradrenergic systems in the mediation of reboxetine's effects, the impact of different chemical lesions of these two pathways was examined on the behavioral responses induced by reboxetine in the forced swim test. Reboxetine (5-20 mg/kg, s.c.) dose-dependently decreased immobility and swimming behavior in the forced swim test while it simultaneously increased climbing behavior. These effects were similar to those previously demonstrated with tricyclic antidepressants and are indicative of reboxetine's effects on the noradrenergic system. Discrete local injections of the neurotoxin 6-hydroxydopamine were employed to lesion the ventral noradrenergic bundle arising from cells located in the lateral tegmentum. This resulting lesion completely prevented reboxetine (10 mg/kg, s.c.)-induced decreases in immobility and increases in climbing behavior, demonstrating that an intact ventral noradrenergic bundle is required for the manifestation of reboxetine-induced antidepressant-like behavior in the test. In contrast, lesions of the dorsal noradrenergic bundle which consists of neurons arising from the nucleus locus coereleus, were achieved by systemic pretreatment with the selective noradrenergic neurotoxin N-(2-chloroethyl)-N-2-bromobenzylamine (DSP-4; 50 mg/kg, i.p.). The ability of reboxetine (10 mg/kg, s.c.) to increase climbing and decrease immobility was augmented by DSP-4 pretreatment. Furthermore, neither lesions of the dorsal noradrenergic bundle nor the ventral noradrenergic bundle altered baseline immobility scores in the forced swim test. Taken together, these data suggest that forebrain regions innervated by these two distinct noradrenergic pathways exert opposing influences on the behavioral response to reboxetine in the rat forced swim test.