Strain Differences in the Chronic Mild Stress Animal Model of Depression and Anxiety in Mice

Chronic mild stress (CMS) has been reported to induce an anhedonic-like state in mice that resembles some of the symptoms of human depression. In the present study, we used a chronic mild stress animal model of depression and anxiety to examine the responses of two strains of mice that have different behavioral responsiveness. An outbred ICR and an inbred C57BL/6 strain of mice were selected because they are widely used strains in behavioral tests. The results showed that the inbred C57BL/6 and outbred ICR mice were similarly responsive to CMS treatment in sucrose intake test (SIT) and open field test (OFT). However, the two strains showed quite different responses in forced swimming test (FST) and novelty-suppressed feeding (NSF) test after 3 weeks of CMS treatment. Only C57BL/6 mice displayed the depression- and anxiety-like behavioral effects in response to CMS treatment in FST and NSF test. Our results suggest that there are differences in responsiveness to CMS according to the different types of strain of mice and behavioral tests. Therefore, these results provide useful information for the selection of appropriate behavioral methods to test depression- and anxiety-like behaviors using CMS in ICR and C57BL/6 mice.     

Reduced anxiety and depression-like behaviors in mice lacking GABA transporter subtype 1.

Gamma-aminobutyric acid (GABA) transporter subtype 1 (GAT1), which transports extracellular GABA into presynaptic neurons, plays an important regulatory role in the function of GABAergic systems. However, the contributions of the GAT1 in regulating mental status are not fully understood. In this paper, we observed the behavioral alterations of GAT1 knockout (GAT1(-/-)) mice using several depression- and anxiety-related models (eg, the forced-swimming test and the tail-suspension test for testing depression-related behaviors; the open-field test, the dark-light exploration test, the emergence test, and the elevated plus maze (EPM) test for anxiety-related behaviors). Here we found that GAT1(-/-) mice showed a lower level of depression- and anxiety-like behaviors in comparison to wild-type mice. Furthermore, GAT1(-/-) mice exhibited measurable insensitivity to selected antidepressants and anxiolytics such as fluoxetine, amitriptyline, buspirone, diazepam, and tiagabine in the tail-suspension test and/or the EPM test. Moreover, the basal level of corticosterone was found to be significantly lower in GAT1(-/-) mice. These results showed that the absence of GAT1 affects mental status through enhancing the GABAergic system, as well as modifying the serotonergic system and the hypothalamic-pituitary-adrenal (HPA) activity in mice.     

Chronic stress alters neuropeptide Y signaling in the bed nucleus of the stria terminalis in DBA/2J but not C57BL/6J mice

Numerous rodent and human studies have demonstrated that neuropeptide Y (NPY) is involved in the regulation of anxiety-related behaviors. In this study, we examined whether there were differences in NPY signaling between two inbred mouse strains (C57BL/6J and DBA/2J) that exhibit divergent basal and stress-induced anxiety phenotypes. We focused on the bed nucleus of the stria terminals (BNST), a structure in the extended amygdala that is important for the regulation of anxiety-like behavior and contains NPY receptors. While results from whole-cell voltage-clamp recordings and immunofluorescence histochemistry revealed no significant basal differences in NPY signaling or NPY and NPY Y2 receptor (Y2R) expression in the BNST, these measures were differentially altered by chronic restraint stress. Ten days of chronic restraint stress increased basal GABAergic transmission and decreased NPY's ability to inhibit evoked GABAergic transmission in the dorsolateral BNST (dlBNST) via Y2R in DBA/2J, but not C57BL/6J, mice. Additionally, restraint stress increased NPY and Y2R expression across subregions of the BNST of DBA/2J mice 24 h after the last stress exposure, but no changes were observed in C57BL/6J mice. Together, these results suggest that chronic restraint stress engages the NPY system and alters NPY modulation of inhibitory transmission in the dlBNST of DBA/2J mice, but not C57BL/6J mice, which may be related to increased expression of anxiety-related behaviors in this strain.     

Fluoxetine affects antioxidant system and promotes apoptotic signaling in Wistar rat liver

Selective serotonin reuptake inhibitors (SSRI) are a treatment of choice for stress related disorders including clinical depression and a range of anxiety-related disorders. In the experimental animals, chronic stress paradigms are considered as a model of depression, and in that context are used for examining the effects of different drug treatments. The present research was designed to investigate the effect of SSRI fluoxetine on antioxidant status and apoptotic signaling in Wistar rat liver, which is a central organ for activation and detoxification of many xenobiotics and reactive oxygen species. We also investigated whether chronic fluoxetine treatment exhibits the same effects in the liver of control animals vs. animals stressed by chronic psychosocial isolation. Our results revealed that fluoxetine downregulated the activity of superoxide dismutases and upregulated the activity of glutathione peroxidase in both rat groups, while elevating glutathione reductase activity and total antioxidant status only in stressed animals. These results suggested that fluoxetine interfered with stress-induced pathways of oxidative defense in the liver. In addition, in both experimental groups, fluoxetine induced several hallmarks of apoptosis in the liver, including a decrease in Bcl-2 expression and increased DNA fragmentation. However, apoptotic alterations were more pronounced in stressed animals, suggesting that stress related oxidative damage could have primed apoptotic effects of fluoxetine.     

Usefulness of antidepressants for improving the neuropathic pain-like state and pain-induced anxiety through actions at different brain sites.

Clinically, it is well known that chronic pain induces depression, anxiety, and a reduced quality of life. There have been many reports on the relationship between pain and emotion. We previously reported that chronic pain induced anxiety with changes in opioidergic function in the central nervous system. In this study, we evaluated the anxiolytic-like effects of several types of antidepressants under a chronic neuropathic pain-like state and searched for the brain site of action where antidepressants show anxiolytic or antinociceptive effects. Sciatic nerve-ligated mice exhibited thermal hyperalgesia and tactile allodynia from days 7 to 28 after nerve ligation. At 4 weeks after ligation, these mice showed a significant anxiety-related behavior in the light-dark test and the elevated plus-maze test. Under these conditions, repeated administration of antidepressants, including the tricyclic antidepressant (TCA) imipramine, the serotonin noradrenaline reuptake inhibitor (SNRI) milnacipran, and the selective serotonin reuptake inhibitor (SSRI) paroxetine, significantly prevented the anxiety-related behaviors induced by chronic neuropathic pain. These antidepressants also produced a significant reduction in thermal hyperalgesia and tactile allodynia. Moreover, the microinjection of paroxetine into the basolateral amygdala or cingulate cortex reduced anxiety-related behavior, and microinjection into the primary somatosensory cortex significantly attenuated thermal hyperalgesia. These findings suggest that serotonergic antidepressants are effective for treating anxiety associated with chronic neuropathic pain and may be useful for treating neuropathic pain with emotional dysfunction such as anxiety. Furthermore, SSRIs show anxiolytic and antinociceptive effects by acting on different brain regions.     

Long-lasting behavioural and molecular alterations induced by early postnatal fluoxetine exposure are restored by chronic fluoxetine treatment in adult mice

There is evidence that antidepressant drug treatment during a critical period of postnatal development renders mice susceptible to depression- and anxiety-related behaviour in adulthood. The mechanism of how early antidepressant treatment brings about long-term effects in emotional behaviour is not yet understood, but neurotrophins, particularly brain-derived neurotrophic factor (BDNF), have been implicated in this context. We examined the long-term effects of a transient early postnatal fluoxetine treatment on depression- and anxiety-related behaviours as well as gene expression of BDNF and its receptor TrkB in C57BL/6J mice. Treatment with fluoxetine between postnatal days P4 and P21 resulted in a significant loss of body weight and long-lasting behavioural inhibition in adult mice in response to stressful events such as the light–dark or open field tests. Postnatal fluoxetine exposure also decreased behavioural despair in the forced swim test. Both body weight and behavioural alterations were restored by chronic fluoxetine treatment in adulthood. The behavioral alterations were accompanied by changes in hippocampal BDNF mRNA. Specifically, we show that early-life fluoxetine exposure resulted in the long-term upregulation of BDNF expression in adult mice. However, chromatin immunoprecipitation studies did not reveal any changes in the acetylation or trimethylation of histone H3 at the BDNF promoters. Our experiments show that behavioural and molecular changes induced by early postnatal fluoxetine administration are reversed by chronic fluoxetine treatment of adult mice to control levels.     

Chronic administration of 13-cis-retinoic acid increases depression-related behavior in mice.

Retinoid signaling plays a well-established role in neuronal differentiation, neurite outgrowth, and the patterning of the anteroposterior axis of the developing neural tube. However, there is increasing evidence that nutritional vitamin A status and retinoid signaling play an important role in the function of the adult brain. 13-Cis-retinoic acid (13-cis-RA) (isotretinoin or Accutane), a synthetic retinoid that is an effective oral treatment for severe nodular acne, has been linked with depression and suicide in patients. The purpose of this study was to test the hypothesis that chronic administration of 13-cis-RA would lead to depression-related behaviors in mice. Young, adult male mice received 13-cis-RA (1 mg/kg) by daily intraperitoneal injection for 6 weeks. This treatment paradigm produced plasma levels of 13-cis-RA that are comparable to those reported in human patients taking Accutane. In both the forced swim test and the tail suspension test, we found that 13-cis-RA-treated mice spent significantly more time immobile compared to vehicle-treated controls. In the open field test, there was no change in anxiety-related behavior in 13-cis-RA-treated mice. Furthermore, chronic administration of 13-cis-RA did not impair locomotion in either the open field or the rotarod test. Taken together, these results suggest that administration of 13-cis-RA increases depression-related behaviors in mice.     

Increased binding of cortical and hippocampal group II metabotropic glutamate receptors in isolation-reared mice

Post-weaning social isolation in rodents induces behavioral alterations, including hyperlocomotion, depression- and anxiety-like behaviors, aggression, and learning and memory deficits. These behavioral abnormalities may be related to the core symptoms in patients with neuropsychiatric disorders, such as schizophrenia and depression. In view of the recent studies that the group II metabotropic glutamate receptor (mGluR2/3) is involved in neuropsychiatric disorders, the present study examined the effect of isolation rearing on the binding of the mGluR2/3 antagonist [(3)H]LY341495 to mGluR2/3 in the mouse brain by in vitro autoradiography. The [(3)H]LY341495 binding in the prefrontal cortex, cerebral cortical layers I-III and hippocampus was significantly increased by rearing in social isolation while the binding in other brain regions was not altered. A saturation binding study of hippocampal membranes from isolation-reared mice revealed that the B(max) value increased significantly without any changes in the K(d) value. Moreover, the mGluR2/3 antagonist MGS0039 (1.0mg/kg, intraperitoneally) decreased the immobility time of isolation-reared mice in the forced swim test. These results suggest that isolation rearing causes an increase in mGluR2/3 densities in the prefrontal cortex and hippocampus and that the increased receptor function may contribute to pathogenic mechanisms for depression-like behavior of the isolation-reared mice.     

Deficiency in Endocannabinoid Signaling in the Nucleus Accumbens Induced by Chronic Unpredictable Stress

The nucleus accumbens (NAc) is a critical component of the reward circuitry, and dysfunction of the NAc may account for anhedonia and other symptoms of depression. Here, we investigated whether alterations in endocannabinoid (eCB) signaling in the NAc contribute to depression-like behaviors induced by chronic unpredictable stress (CUS) in mice. We compared three types of eCB/CB1 receptor-mediated synaptic plasticity in slices prepared from the NAc core of control and stress-exposed mice: depolarization-induced suppression of excitation, long-term depression, and the depression of field excitatory postsynaptic potentials (fEPSPs) induced by group I metabotropic glutamate receptor agonist DHPG. CUS (5–6-week exposure to stressors), but not sub-CUS (1 week exposure to stressors), induces depression-like behaviors and impairs these forms of eCB/CB1 receptor-mediated plasticity examined in the NAc core. Neither sub-CUS nor CUS altered the tissue contents of the eCBs, anandamide and 2-arachidonoylglycerol in the striatum. However, exposure to CUS, but not to sub-CUS, attenuated the depression of fEPSPs induced by the CB1 receptor agonist WIN 55 212-2. CUS exposure reduced the maximal effect without affecting the EC₅₀ of WIN 55 212-2 to induce fEPSP depression. Thus, impaired CB1 receptor function could account for CUS-induced deficiency in eCB signaling in the NAc. Both CUS-induced deficiency in eCB signaling and depression-like behaviors were reversed by in vivo administration of antidepressant fluoxetine. These results suggest that downregulation of eCB signaling in the NAc occurs after CUS and contributes to the pathophysiology of depression.     

Reversible Gating of Endocannabinoid Plasticity in the Amygdala by Chronic Stress: A Potential Role for Monoacylglycerol Lipase Inhibition in the Prevention of Stress-Induced Behavioral Adaptation

Chronic stress is the primary environmental risk factor for the development and exacerbation of affective disorders, thus understanding the neuroadaptations that occur in response to stress is a critical step in the development of novel therapeutics for depressive and anxiety disorders. Brain endocannabinoid (eCB) signaling is known to modulate emotional behavior and stress responses, and levels of the eCB 2-arachidonoylglycerol (2-AG) are elevated in response to chronic homotypic stress exposure. However, the role of 2-AG in the synaptic and behavioral adaptations to chronic stress is poorly understood. Here, we show that stress-induced development of anxiety-like behavior is paralleled by a transient appearance of low-frequency stimulation-induced, 2-AG-mediated long-term depression at GABAergic synapses in the basolateral amygdala, a key region involved in motivation, affective regulation, and emotional learning. This enhancement of 2-AG signaling is mediated, in part, via downregulation of the primary 2-AG-degrading enzyme monoacylglycerol lipase (MAGL). Acute in vivo inhibition of MAGL had little effect on anxiety-related behaviors. However, chronic stress-induced anxiety-like behavior and emergence of long-term depression of GABAergic transmission was prevented by chronic MAGL inhibition, likely via an occlusive mechanism. These data indicate that chronic stress reversibly gates eCB synaptic plasticity at inhibitory synapses in the amygdala, and in vivo augmentation of 2-AG levels prevents both behavioral and synaptic adaptations to chronic stress.     

ROCK inhibition produces anxiety-related behaviors in mice

Rationale The role of Rho/Rho-associated kinase (ROCK) in regulating dendritic and axonal morphology during development has gained much attention. Very little is known, however, about the role of the Rho/ROCK pathway in emotional behavior.Objective To investigate the role of ROCK in emotional behaviors. We examined how the ROCK inhibitor Y27632 affects the performance of mice on three behavioral tests that measure anxiety-related behaviors.Results In the elevated plus-maze test, Y27632 (10 nmol, intracerebroventricular) induced a significant decrease in the percentage of time spent in the open arms and in the percentage of entries into open arms. In the fear conditioning test, Y27632-treated mice froze significantly more often and longer than did saline-treated mice. In the hole-board test, Y27632 significantly suppressed head-dipping behavior in Y27632-treated mice than in saline-treated mice. On the other hand, Y27632 did not produce on spontaneous alteration performance in the Y-maze test. These results indicate that ROCK inhibition increased anxiety-related behaviors.Conclusion Our findings suggest that the ROCK pathway is involved in the expression of anxiety- and fear-related behaviors. Furthermore, we propose that if the Rho/ROCK pathway plays an important role in mediating anxiety-related behaviors in humans, it may prove to be a novel system for anxiolytics to target.     

Magnesium-deficient diet alters depression- and anxiety-related behavior in mice--influence of desipramine and Hypericum perforatum extract

A relation between magnesium (Mg) status and mood disorders has been suggested, but evidence remains inconsistent. Therefore, we examined in mice whether Mg-depletion would alter behavior evaluated in established animal models of depression and anxiety and whether these effects would be sensitive to antidepressants. Compared to control mice fed with normal diet, mice receiving a low Mg diet (10% of daily requirement) for several weeks displayed increased immobility time in the forced swim test, indicating enhanced depression-like behavior. In addition, the partial Mg-depletion increased anxiety-related behavior in the light/dark and open field test, while locomotor activity or motor coordination was not influenced. Chronic oral administration of desipramine (30 mg/kg/day), or Hypericum extract LI160 (Hyp, 380 mg/kg/day) prevented the “pro-depression-like” forced swim behavior in Mg-depleted mice. Furthermore, the increase in anxiety-related behavior of Mg-depleted mice was abolished in both the open field and light dark test by Hyp. Taken together, we report that Mg-depletion leads to enhanced depression- and anxiety-related behavior in mice, which was further validated by the reversibility of the behavioral changes by known antidepressant and anxiolytic substances. Further, the utility of Mg-depletion as a new screening model for clinically active antidepressant and anxiolytic drugs is suggested.     

Behavioral effects of Bcl-2 deficiency: implications for affective disorders

New hypotheses regarding affective disorders suggest a critical role for cellular resilience and plasticity. Bcl-2 is a central protein in these processes and is elevated by mood stabilizers and antidepressants. In previous studies, mice with targeted mutations of Bcl-2 showed anxiety-related behavioral changes. The present study further explored the relationship between Bcl-2 and behavior using mice with a targeted mutation but with a different background strain than previously tested. Bcl-2 heterozygous mice (B6;129S2-Bcl-2/J) were tested in models of depression, mania and anxiety. Compared to Wild Type (WT) controls, mutant mice showed behaviors modeling two facets of mania: increased reward seeking and amphetamine sensitization. Moreover, the sensitization was attenuated by chronic pretreatment with lithium. In contrast to previous data, the mutation did not affect measures of anxiety. Although data are still minimal, it supports additional studies of the role of Bcl-2 in affective and anxiety disorders. The importance of background strain in behavioral phenotypes of mutant mice is known and the current lack of effect on anxiety measures may be related to high baseline anxiety of WTanimals. More precise studies of Bcl-2 in affective and anxiety disorders will be possible when specific pharmacological modulators of Bcl-2 become available.     

Stress-induced hyperlocomotion as a confounding factor in anxiety and depression models in mice

Chronic stress is broadly used to model anxiety and depression. However, in chronic stress models, anxiety- and depression-like behaviors might be masked by unspecific effects of stress. We tested whether chronic stress in mice can induce unspecific changes in locomotion, and whether these changes interfere with the measurement of anxiety and forced-swimming behaviors. Also, we studied these latter behaviors in relation to the duration of stress, the lighting conditions during testing, and after the injection of diazepam. We employed a 4-week chronic stress paradigm, adopted from a model of stress-induced anhedonia and a 1-week subchronic stress, both consisting of rat exposure, restraint stress and tail suspension. Chronically stressed mice, tested under bright and moderate illumination, exhibited 'anxiolytic-like' behavior along with prolonged swimming and hyperactivity. These behaviors were not detectable under weak illumination or after the injection of diazepam (0.25 mg/kg). Instead, normal locomotion, increased anxiety and inhibited swimming were revealed under these conditions. Thus, chronic stress can induce hyperlocomotion in mice, which is triggered by acute stressors such as light, and interferes with the evaluation of anxiety and forced swimming. One week of stress did not change locomotion and forced swimming, and increased anxiety irrespective of illumination applied during testing. Our data can possibly explain previously reported contradictions in the behavioral testing of mice with chronic stress models of anxiety and depression.     

Antidepressant and Proneurogenic Influence of Environmental Enrichment in Mice: Protective Effects vs Recovery

Physical–cognitive activity has long-lasting beneficial effects on the brain and on behavior. Environmental enrichment (EE) induces brain activity known to influence the behavior of mice, as measured in learned helplessness paradigms (forced swim test), and neurogenic cell populations in the hippocampal dentate gyrus. However, it is not completely clear whether the antidepressant and proneurogenic effects of EE are different in animals that are naive or pre-exposed to the stress inducing helplessness, and if this depends on the type of stressor. It also remains unclear whether differential effects are exerted on distinct neurogenic subpopulations. We found that EE has a protective effect in adult female mice (C57BL/6J) when exposed twice to the same stressor (forced swim test) but it has no influence on recovery. The repeated exposure to this stressor was analyzed together with the effects of EE on different neurogenic populations distinguished by age and differentiation state. Younger cells are more sensitive and responsive to the conditions, both the positive and negative effects. These results are relevant to identify the cell populations that are the targets of stress, depression, and enrichment, and that form part of the mechanism responsible for mood dysfunctions.     

The effects of social isolation on neuropeptide Y levels, exploratory and anxiety-related behaviors in rats

NPY is one of the most abundantly expressed peptides within the CNS, and has been previously demonstrated to be altered in several animal models of depression, as well as to be differentially regulated by acute and repeated stress. The effect of social deprivation, through isolation housing, on brain NPY concentrations in adult rats has not been previously investigated. The effects of 12 weeks of social isolation, in adult rats, on anxiety-related behaviors and central concentrations of NPY in: hypothalamus, amygdala, caudate-putamen, hippocampus, and frontal cortex were evaluated. Single housed animals spent significantly more time on the open arms of the elevated plus maze and in the central area of the open field as compared to pair housed controls. These data are most likely indicative of enhanced exploration and novelty seeking. Concentrations of neuropeptide Y were increased in the caudate-putamen of the single housed subjects. NPY levels in caudate/putamen and hypothalamus were also significantly correlated with time spent in the open arms of the elevated plus maze. These data suggest that chronic social isolation, in these adult Wistar rats, did not increase anxiety but produced enhanced exploration in tests of anxiety, an effect that was associated with NPY concentrations in the striatum and hypothalamus.     

Inconsistent effects of photoperiod manipulations in tests for affective-like changes in mice: implications for the selection of appropriate model animals

Deficiencies in appropriate animal models are a significant factor hindering the research of affective disorders. Significant data suggest that systems related to circadian rhythms are strongly linked to affective changes, but study with animal models in this context had unclear and inconsistent results. Circadian physiology is significantly different in diurnal and nocturnal animals and a recent project showed that in diurnal rodents, short photoperiods induce depression and anxiety-like phenotypes. This study was designed to evaluate the possibility that using a similar methodology would also result in behavioral changes in nocturnal mice. Mice from two strains were maintained in either short photoperiod, neutral photoperiod or long photoperiod for 3 weeks and tested for depression or anxiety-related behaviors, as done earlier with the diurnal rodents. Tests included activity levels, sweet solution preference, elevated plus-maze, resident-intruder aggression, and forced swim test. Tests were conducted either during the light phase or during the dark phase of the mice. In contrast to the clear phenotype in diurnal rodents, the effects of photoperiod manipulations in nocturnal mice were inconsistent. These results suggest that diurnal rodents may be advantageous compared with nocturnal species for studies exploring the relationship between circadian rhythms and affective disorders.     

Antidepressant and anxiolytic effects of the proprietary Chinese medicine Shexiang Baoxin pill in mice with chronic unpredictable mild stress

Depression and anxiety often co-occur with cardiac diseases. The Shexiang Baoxin pill (SBP) is a proprietary Chinese medicine initially used to treat cardiac conditions. This study explored whether SBP has antidepressant and anxiolytic effects in addition to hormonal and psychotropic mechanisms. Mice underwent 6 weeks of chronic unpredictable mild stress (CUMS) to induce depression- and anxiety-like behavior. During the 6-week experiment, mice received SBP at intragastric doses of 20.25 mg/kg or 40.5 mg/kg daily. Animals were then tested for depression in sucrose preference, forced-swimming, and tail suspension paradigms, and for anxiety in open field and elevated plus maze tests. Both SBP doses significantly reduced anhedonic behavior in the sucrose preference test; the high SBP dose also increased the number of entries into the central zone of the open field. SBP-treated mice had markedly lower blood levels of corticotrophin-releasing hormone (CRH) and adrenocorticotropic hormone (ACTH) than stressed mice treated with vehicle. Either low- or high-dose SBP reversed stress-induced reductions of norepinephrine (NE) and dopamine (DA) metabolites and the expression levels of brain-derived neurotrophic factor (BDNF), nerve growth factor (NGF), and glial cell-derived neurotrophic factor (GDNF) in related brain regions. These results suggest that SBP could prevent and alleviate prolonged stress-induced anhedonia and anxiety in association with its suppression of the hypothalamic-pituitary-adrenal (HPA) axis hyperactivity, modulation of brain monoamine neurotransmitter metabolism and neurotrophins. SBP may be particularly suitable for the management of depressive and anxiety disorders in patients with cardiac conditions.     

Involvement of α-MSH in the social isolation induced anxiety- and depression-like behaviors in rat

Although physical isolation of rats is known to cause anxiety- and depression-like symptoms, the underlying mechanisms are not fully understood. We have attempted to define the role of endogenous melanocortins (MC) in the manifestation of these symptoms. Weaning rats were socially isolated for 6 weeks and subjected to behavioral paradigms like elevated plus maze (EPM), social interaction, and forced swim test (FST). While socially isolated rats spent less time in social interaction, and showed significantly decreased activity in the open arms of the EPM, the immobility time in FST was significantly increased thus reflecting anxiety- and depression-like phenotypes. Intracerebroventricular injection of HS014 (5 or 10 nmol/rat), selective antagonist of MC4 receptors, attenuated these symptoms. This suggested the involvement of endogenous alpha-melanocyte stimulating hormone (α-MSH) in anxiety and depression. With a view to determining the neuroanatomical substrates in which the endogenous α-MSH may process the related information, profile of the peptide in paraventricular (PVN), arcuate (ARC), dorsomedial hypothalamic-dorsal (DMNd) and -ventral (DMNv) nuclei, and central nucleus of amygdala (CeA) was investigated with immunohistochemistry. While social isolation significantly reduced α-MSH-immunoreactivity profile in all these components, re-socialization of the socially isolated rats, over a period of 72 h, resulted in full recovery of the α-MSH-immunoreactivity profile; the symptoms of anxiety- and depression-like behaviors were also fully attenuated. We suggest that α-MSH in the PVN, ARC, DMNd, DMNv and CeA, acting via MC4 receptors, are involved in manifestation of affective disorders like anxiety and depression.