Post-weaning Isolation Research Articles

Transcriptomic analysis of rat prefrontal cortex following chronic stress induced by social isolation – Relevance to psychiatric and neurodevelopmental illness, and implications for treatment

Social isolation is an established risk factor for psychiatric illness, and became increasingly topical with the spread of SARS-CoV-2. We used RNA sequencing (RNA-Seq) to enable unbiased assessment of transcriptomic changes within the prefrontal cortex (PFC) of isolation-reared rats. To provide insight into the relevance of this manipulation for studying human illness, we compared differentially expressed genes (DEGs) and enriched biological functions against datasets involving post-mortem frontal cortical tissue from patients with psychiatric and neurodevelopmental illnesses. Sixteen male Sprague-Dawley rats were reared in groups of four or individually from weaning on postnatal day (PND) 22–24 until PFC tissue collection for RNA-Seq (PND64-66). We identified a total of 183 DEGs in isolates, of which 128 mirrored those in PFC tissue from patients with stress-related mental illnesses and/or neurodevelopmental conditions featuring social deficits. Seventy-one encode proteins classed as druggable by the gene-drug interaction database. Interestingly there are antagonists or inhibitors for the products of three of these up-regulated DEGs (Hrh3, Snca and Sod1) and agonists or activators for products of six of these down-regulated DEGs (Chrm4, Klf2, Lrrk2, Nr4a1, Nr4a3 and Prkca). Some have already undergone pre-clinical and clinical evaluation, and studies with the remainder may be warranted. Changes to Hrh3, Sod1, Chrm4, Lrrk2, Nr4a1 and Prkca were replicated in an independent cohort of sixteen male Sprague-Dawley rats via quantitative reverse transcription polymerase chain reaction (qRT-PCR). Our findings support the continued use of post-weaning isolation rearing to investigate the neurobiology of stress-related disorders and evaluate therapeutic targets.

Post-weaning social isolation alters sociability in a sex-specific manner.

Adolescence is a critical period for brain development in humans and stress exposure during this time can have lasting effects on behavior and brain development. Social isolation and loneliness are particularly salient stressors that lead to detrimental mental health outcomes particularly in females, although most of the preclinical work on social isolation has been done in male animals. Our lab has developed a model of post-weaning adolescent social isolation that leads to increased drug reward sensitivity and altered neuronal structure in limbic brain regions. The current study utilized this model to determine the impact of adolescent social isolation on a three-chamber social interaction task both during adolescence and adulthood. We found that while post-weaning isolation does not alter social interaction during adolescence (PND45), it has sex-specific effects on social interaction in adulthood (PND60), potentiating social interaction in male mice and decreasing it in female mice. As early life stress can activate microglia leading to alterations in neuronal pruning, we next examined the impact of inhibiting microglial activation with daily minocycline administration during the first three weeks of social isolation on these changes in social interaction. During adolescence, minocycline dampened social interaction in male mice, while having no effect in females. In contrast, during adulthood, minocycline did not alter the impact of adolescent social isolation in males, with socially isolated males exhibiting higher levels of social interaction compared to their group housed counterparts. In females, adolescent minocycline treatment reversed the effect of social isolation leading to increased social interaction in the social isolation group, mimicking what is seen in naïve males. Taken together, adolescent social isolation leads to sex-specific effects on social interaction in adulthood and adolescent minocycline treatment alters the effects of social isolation in females, but not males.

GABAergic and inflammatory changes in the frontal cortex following neonatal PCP plus isolation rearing, as a dual-hit neurodevelopmental model for schizophrenia

The pathogenesis of schizophrenia begins in early neurodevelopment and leads to excitatory-inhibitory imbalance. It is therefore essential that preclinical models used to understand disease, select drug targets and evaluate novel therapeutics encompass similar neurochemical deficits. One approach to improved preclinical modelling incorporates dual-hit neurodevelopmental insults, like neonatal administration of phencyclidine (PCP, to disrupt development of glutamatergic circuitry) then post-weaning isolation (Iso, to mimic adolescent social stress). We recently showed that male Lister-hooded rats exposed to PCP-Iso exhibit reduced hippocampal expression of the GABA interneuron marker calbindin. The current study expanded on this by investigating changes to additional populations of GABAergic interneurons in frontal cortical and hippocampal tissue from the same animals (by immunohistochemistry) as well as levels of GABA itself (via ELISA). Because inflammatory changes are also implicated in schizophrenia, we performed additional immunohistochemical evaluations of Iba-1 positive microglia as well as ELISA analysis of IL-6 in the same brain regions. Single-hit isolation-reared and dual-hit PCP-Iso rats both showed reduced parvalbumin immunoreactivity in the prelimbic/infralimbic region of the frontal cortex. However, this was more widespread in PCP-Iso, extending to the medial/ventral and lateral/dorsolateral orbitofrontal cortices. Loss of GABAergic markers was accompanied by increased microglial activation in the medial/ventral orbitofrontal cortices of PCP-Iso, together with frontal cortical IL-6 elevations not seen following single-hit isolation rearing. These findings enhance the face validity of PCP-Iso, and we advocate the use of this preclinical model for future evaluation of novel therapeutics—especially those designed to normalise excitatory-inhibitory imbalance or reduce neuroinflammation.

Post-weaning social isolation alters sociability in a sex-specific manner.

Adolescence is a critical period for brain development in humans and stress exposure during this time can have lasting effects on behavior and brain development. Social isolation and loneliness are particularly salient stressors that lead to detrimental mental health outcomes particularly in females, although most of the preclinical work on social isolation has been done in male animals. Our lab has developed a model of post-weaning adolescent social isolation that leads to increased drug reward sensitivity and altered neuronal structure in limbic brain regions. The current study utilized this model to determine the impact of adolescent social isolation on a three-chamber social interaction task both during adolescence and adulthood. We found that while post-weaning isolation does not alter social interaction during adolescence (PND45), it has sex-specific effects on social interaction in young adulthood (PND60), potentiating social interaction in male mice and decreasing it in female mice. As early life stress can activate microglia leading to alterations in neuronal pruning, we next examined the impact of inhibiting microglial activation with daily minocycline administration during the first 3 weeks of social isolation on these changes in social interaction. During adolescence, minocycline dampened social interaction in male mice, while having no effect in females. In contrast, during young adulthood, minocycline did not alter the impact of adolescent social isolation in males, with socially isolated males exhibiting higher levels of social interaction compared to their group housed counterparts. In females, adolescent minocycline treatment reversed the effect of social isolation leading to increased social interaction in the social isolation group, mimicking what is seen in naïve males. Taken together, adolescent social isolation leads to sex-specific effects on social interaction in young adulthood and adolescent minocycline treatment alters the effects of social isolation in females, but not males.

Molecular and cellular mechanisms for differential effects of chronic social isolation stress in males and females.

Chronic social isolation stress during adolescence induces susceptibility for neuropsychiatric disorders. Here we show that 5-week post-weaning isolation stress induces sex-specific behavioral abnormalities and neuronal activity changes in the prefrontal cortex (PFC), basal lateral amygdala (BLA), and ventral tegmental area (VTA). Chemogenetic manipulation, optogenetic recording, and in vivo calcium imaging identify that the PFC to BLA pathway is causally linked to heightened aggression in stressed males, and the PFC to VTA pathway is causally linked to social withdrawal in stressed females. Isolation stress induces genome-wide transcriptional alterations in a region-specific manner. Particularly, the upregulated genes in BLA of stressed males are under the control of activated transcription factor CREB, and CREB inhibition in BLA normalizes gene expression and reverses aggressive behaviors. On the other hand, neuropeptide Hcrt (Hypocretin/Orexin) is among the top-ranking downregulated genes in VTA of stressed females, and Orexin-A treatment rescues social withdrawal. These results have revealed molecular mechanisms and potential therapeutic targets for stress-related mental illness.

Post-weaning social isolation causes sex-specific alterations to dendritic spine density in subregions of the prefrontal cortex and nucleus accumbens of adult mice

Post-weaning social isolation stress has been shown to increase addiction-like behavior in adulthood. These long-term behavioral alterations may be due to long lasting isolation-induced structural changes to neurons in brain regions involved in reward processing. Previous studies have shown that various stressors alter dendritic spine density in the prefrontal cortex (PFC) and the nucleus accumbens, though many of these studies examine the short-term effects of stress, and are primarily conducted in males. There is mounting evidence that males and females exhibit differences in their stress responses, with some studies showing sex differences in stress-induced plasticity. To determine the long-lasting, sex-specific alterations in spine density following post-weaning social isolation, male and female mice were either isolated or group housed at weaning and spine density was measured once they reached adulthood. Post-weaning isolation increased spine density in the PFC of both the males and females, although the effects in the infralimbic cortex were more pronounced in the females. In the nucleus accumbens, adolescent isolation increased spine density in males only in the core and shell. Females also had higher baseline spine density than males in the nucleus accumbens core. Together these data suggest that adolescent social isolation causes long-term, sex-specific alterations to the prefrontal cortex and the nucleus accumbens.

Hippocampal chloride transporter KCC2 contributes to excitatory GABA dysregulation in the developmental rat model of schizophrenia

Recent studies have revealed an altered expression of NKCC1 and KCC2 in prefrontal cortex (PFC) and hippocampus of schizophrenic patients. Despite extensive considerations, the alteration of NKCC1 and KCC2 co-transporters at different stages of development has not been fully studied. Therefore, we evaluated the expression of these transporters in PFC and hippocampus at time points of four, eight, and twelve weeks in post-weaning social isolation rearing rat model. For this purpose, 23–25 days-old rats were classified into social- or isolation-reared groups. The levels of NKCC1 and KCC2 mRNA expression were evaluated at hippocampus or PFC regions at the time-points of four, eight, and twelve weeks following housing. Post-weaning isolation rearing decreased the hippocampal KCC2 mRNA expression level, but does not affect the NKCC1 mRNA expression. However, no significant difference was observed in the PFC mRNA levels of NKCC1 and KCC2 in the isolation-reared group compared to the socially-reared group during the course of modeling. Further, we assessed the therapeutic effect of selective NKCC1 inhibitor bumetanide (10 mg/kg), on improvement of prepulse inhibition (PPI) test on twelve weeks isolation-reared rats. Intraperitoneal administration of bumetanide (10 mg/kg) did not exert beneficial effects on PPI deficit. Our findings show that isolation rearing reduces hippocampal KCC2 expression level and may underlie hippocampal GABA excitatory. In addition, 10 mg/kg bumetanide is not effective in improving the reduced PPI of twelve weeks isolation-reared rats.Collectively, our findings show that hippocampal chloride transporter KCC2 contributes to excitatory GABA dysregulation in the developmental rat model of schizophrenia.

Duration of Social Isolation Affects Production of Nitric Oxide in the Rat Brain.

Social isolation deprives rodents of social interactions that are critical for normal development of brain and behavior. Several studies have indicated that postweaning isolation rearing may affect nitric oxide (NO) production. The aim of this study was to compare selected behavioral and biochemical changes related to NO production in the brain of rats reared in social isolation for different duration. At the age of 21 days, male Sprague Dawley rats were randomly assigned into four groups reared in isolation or socially for 10 or 29 weeks. At the end of the rearing, open-field and prepulse inhibition (PPI) tests were carried out. Furthermore, in several brain areas we assessed NO synthase (NOS) activity, protein expression of nNOS and iNOS isoforms and the concentration of conjugated dienes (CD), a marker of oxidative damage and lipid peroxidation. Social isolation for 10 weeks resulted in a significant decrease in PPI, which was accompanied by a decrease in NOS activity in the cerebral cortex and the cerebellum, an increase in iNOS in the hippocampus and an increase in CD concentration in cortex homogenate. On the other hand, a 29 week isolation had an opposite effect on NOS activity, which increased in the cerebral cortex and the cerebellum in animals reared in social isolation, accompanied by a decrease in CD concentration. The decrease in NOS activity after 10 weeks of isolation might have been caused by chronic stress induced by social isolation, which has been documented in previous studies. The increased oxidative state might result in the depleted NO bioavailability, as NO reacts with superoxide radical creating peroxynitrite. After 29 weeks of isolation, this loss of NO might be compensated by the subsequent increase in NOS activity.

Behavioral and neurochemical changes induced by post-weaning female rats isolation

Rodents post-weaning isolation could be considered a significant early-life model of chronic stress. The aim of our study was to investigate the effects of female rats post-weaning isolation on different behavioral tests and neurochemical parameters. On postnatal day 21, female pups were housed either isolated or socially in groups (4/cage) for 6 weeks. After the 6th week adult female rats were behaviorally examined using open field test, three chamber social interaction test, hole board test and condition avoidance test. In addition, hippocampal content of neurotransmitters, oxidative stress and inflammatory markers were measured. Isolated female rats exhibited behavioral changes, female rats shows a significant increase in anxiety and a significant decrease in locomotor activity in comparison to the control group using open field test . Moreover, female rats isolation promoted a significant decrease in sociality index as compared to control groups. Finally, female rats post-weaning isolation of invoked a significant impairment in long-term and short-term memory evaluated by both conditioned avoidance and hole board tests, in comparison to the control group. Furthermore, female rats post- weaning isolation induced oxidative stress, inflammation, and neurotransmitter alterations. Collectively female rats post weaning isolation had significantly prompted both behavioral and neurochemical changes which could represent the essential important symptoms of schizophrenia.

Interplay of Prenatal and Postnatal Risk Factors in the Behavioral and Histological Features of a "Two-Hit" Non-Genetic Mouse Model of Schizophrenia.

Schizophrenia is a multifactorial developmental neuropsychiatric disorder. This study examined the interplay of maternal infection and postweaning social isolation, which are prenatal and postnatal risk factors, respectively. Pregnant mice received poly I:C or saline injection on gestation day 9 and the pups were weaned at postnatal day 28. After weaning, male offspring were randomly assigned into group-rearing and isolation-rearing groups. In their adulthood, we performed behavioral tests and characterized the histochemical features of their mesocorticolimbic structures. The sociability and anxiety levels were not affected by either manipulation, but synergistic effects of the two hits on stress-coping behavior was observed. Either of the single manipulations caused defects in sensorimotor gating, novel object recognition and spatial memory tests, but the combination of the two hits did not further exacerbate the disabilities. Prenatal infection increased the number of dopaminergic neurons in midbrain, whereas postweaning isolation decreased the GABAergic neurons in cortex. Single manipulation reduced the dendritic complexity and spine densities of neurons in the medial prefrontal cortex (mPFC) and dentate gyrus. Our results support the current perspective that disturbances in brain development during the prenatal or postnatal period influence the structure and function of the brain and together augment the susceptibility to mental disorders, such as schizophrenia.

Gestational poly(I:C) attenuates, not exacerbates, the behavioral, cytokine and mTOR changes caused by isolation rearing in a rat ‘dual-hit’ model for neurodevelopmental disorders

Many psychiatric illnesses have a multifactorial etiology involving genetic and environmental risk factors that trigger persistent neurodevelopmental impairments. Several risk factors have been individually replicated in rodents, to understand disease mechanisms and evaluate novel treatments, particularly for poorly-managed negative and cognitive symptoms. However, the complex interplay between various factors remains unclear. Rodent dual-hit neurodevelopmental models offer vital opportunities to examine this and explore new strategies for early therapeutic intervention. This study combined gestational administration of polyinosinic:polycytidylic acid (poly(I:C); PIC, to mimic viral infection during pregnancy) with post-weaning isolation of resulting offspring (to mirror adolescent social adversity). After in vitro and in vivo studies required for laboratory-specific PIC characterization and optimization, we administered 10 mg/kg i.p. PIC potassium salt to time-mated Lister hooded dams on gestational day 15. This induced transient hypothermia, sickness behavior and weight loss in the dams, and led to locomotor hyperactivity, elevated striatal cytokine levels, and increased frontal cortical JNK phosphorylation in the offspring at adulthood. Remarkably, instead of exacerbating the well-characterized isolation syndrome, gestational PIC exposure actually protected against a spectrum of isolation-induced behavioral and brain regional changes. Thus isolation reared rats exhibited locomotor hyperactivity, impaired associative memory and reversal learning, elevated hippocampal and frontal cortical cytokine levels, and increased mammalian target of rapamycin (mTOR) activation in the frontal cortex – which were not evident in isolates previously exposed to gestational PIC. Brains from adolescent littermates suggest little contribution of cytokines, mTOR or JNK to early development of the isolation syndrome, or resilience conferred by PIC. But notably hippocampal oxytocin, which can protect against stress, was higher in adolescent PIC-exposed isolates so might contribute to a more favorable outcome. These findings have implications for identifying individuals at risk for disorders like schizophrenia who may benefit from early therapeutic intervention, and justify preclinical assessment of whether adolescent oxytocin manipulations can modulate disease onset or progression.

Rosuvastatin improves olanzapine’s effects on behavioral impairment and hippocampal, hepatic and metabolic damages in isolated reared male rats

Background & aimSchizophrenia is a chronic, disabling neurological illness. This study investigated the effect of rosuvastatin (RSU) addition to the antipsychotic drug: olanzapine (OLZ) in treatment of post-weaning isolation rearing (IR) damaging effect and assessed behavioral impairment, metabolic and hepatic abnormalities, oxidative stress, and inflammatory markers. MethodsTreatment with OLZ (6 mg/kg, P.O.) and/or RSU (10 mg/kg, I.P.) have been started 6 weeks after isolation. We assessed behavioral tests, serum cortisol level, and hippocampal content of neurotransmitters. In addition, we assessed histopathology, inflammatory and oxidative stress markers of hippocampus, liver and adipose tissue ResultsTreatment of IR animals with OLZ, and/or RSU significantly counteracted the changes in hippocampus, liver and adipose tissue induced by post-weaning IR. Co-treatment of IR rats with both OLZ and RSU showed additive effects in some areas like improving both tumor necrosis factor alpha (TNFα) in both hippocampus and liver, histopathology of liver, oxidative stress markers of adipose tissue, β3 adrenergic receptors (ADRβ3), serum cortisol and total cholesterol. In addition, RSU alone alleviated the damage of IR rats by the same efficacy as OLZ with more benefit in cognition and exploration. Conclusionpost-weaning IR as a model has behavioral, hippocampal, hepatic and marked metabolic changes more relevant to schizophrenia than drug-induced models. These effects were ameliorated by RSU and/or OLZ that are explained by their antioxidant, anti-inflammatory, anti-stress and anti-hyperlipidemic properties. Interestingly, co-treatment with both drugs showed a better effect.

Behavioral alterations induced by post-weaning isolation rearing of rats are accompanied by reduced VGF/BDNF/TrkB signaling in the hippocampus

Post-weaning social isolation has been shown to be a relevant animal model for studying the mechanisms underlying psychopathological states induced by early-life stressful experiences. Besides extensively studied brain-derived neurotrophic factor (BDNF) and tyrosine receptor kinase B (TrkB) receptor, increasing attention is being given to a neuropeptide precursor VGF (non-acronymic). Several lines of evidence indicate an interplay between the neurotrophins and nitric oxide signaling. This study investigated the long-term consequences of post-weaning social isolation on behavior, VGF/BDNF/TrkB pathway and two isoforms of nitric oxide synthase (NOS) in the hippocampus and examined whether these effects were sex-specific. Male and female Sprague-Dawley rats were reared either in social isolation or social groups from postnatal day 21 for 9 weeks (n = 12–15/group and sex). Post-weaning social isolation induced impairments in sensorimotor gating and increased anxiety-like behavior in rats of both sexes. These behavioral alterations were accompanied by attenuated gene expression of VGF and TrkB receptor in the hippocampus. Isolation-induced reduction in VGF gene expression was more evident in male isolates. Similar changes were found in neuronal NOS (nNOS) gene expression with reduced mRNA levels in male isolates. Gene expression of BDNF and inducible NOS was not influenced by isolation rearing or sex. In addition, sex-specific patterns of VGF and nNOS gene expression in the hippocampus with higher mRNA levels in males than in females were revealed. The present study demonstrates a relationship between nNOS, VGF, BDNF, and TrkB confirming a link between nitric oxide and neurotrophins signaling pathways. Our findings indicate that long-term post-weaning social isolation alters signaling via VGF/BDNF/TrkB and nNOS that could interfere with neurodevelopmental processes which may contribute to pathological behavioral symptoms in adulthood. Future studies are needed to support this suggestion since the direct mechanistic link has not been approached in this study.

Beyond the Activity-Based Anorexia Model: Reinforcing Values of Exercise and Feeding Examined in Stressed Adolescent Male and Female Mice.

Anorexia nervosa (AN), mostly observed in female adolescents, is the most fatal mental illness. Its core is a motivational imbalance between exercise and feeding in favor of the former. The most privileged animal model of AN is the “activity-based anorexia” (ABA) model wherein partly starved rodents housed with running wheels exercise at the expense of feeding. However, the ABA model bears face and construct validity limits, including its inability to specifically assess running motivation and feeding motivation. As infant/adolescent trauma is a precipitating factor in AN, this study first analyzed post-weaning isolation rearing (PWIR) impacts on body weights and wheel-running performances in female mice exposed to an ABA protocol. Next, we studied through operant conditioning protocols i) whether food restriction affects in a sex-dependent manner running motivation before ii) investigating how PWIR and sex affect running and feeding drives under ad libitum fed conditions and food restriction. Besides amplifying ABA-elicited body weight reductions, PWIR stimulated wheel-running activities in anticipation of feeding in female mice, suggesting increased running motivation. To confirm this hypothesis, we used a cued-reward motivated instrumental task wherein wheel-running was conditioned by prior nose poke responses. It was first observed that food restriction increased running motivation in male, but not female, mice. When fed grouped and PWIR mice were tested for their running and palatable feeding drives, all mice, excepted PWIR males, displayed increased nose poke responses for running over feeding. This was true when rewards were proposed alone or within a concurrent test. The increased preference for running over feeding in fed females did not extend to running performances (time, distance) during each rewarded sequence, confirming that motivation for, and performance during, running are independent entities. With food restriction, mice displayed a sex-independent increase in their preference for feeding over running in both group-housed and PWIR conditions. This study shows that the ABA model does not specifically capture running and feeding drives, i.e. components known to be affected in AN.

Postweaning Isolation Rearing Alters the Adult Social, Sexual Preference and Mating Behaviors of Male CD-1 Mice.

Objective: No study has comprehensively evaluated the effect of postweaning isolation on the social and sexual behaviors of a certain strain of rodents in ethology. The present study aims to explore how and to what extent isolation rearing after postweaning affects the social and sexual behaviors of male CD-1 mice in adulthood systematically.Methods: Male CD-1 mice were randomly assigned to two groups: isolation reared (IS, one mouse per cage, n = 30) and group housed (GH, five mice per cage, n = 15). The mice underwent isolation rearing from postnatal day 23 to day 93. Then, social affiliation, recognition and memory were measured through selection task experiments. Social interaction under a home cage and novel environment were measured via resident–intruder and social interaction test, respectively. Furthermore, sexual preference, homosexual and heterosexual behaviors were measured.Results: Our study found that postweaning isolation increased the social affiliation for conspecifics (p = 0.001), reduced social recognition (p = 0.042) and impaired social memory. As for social interaction, isolated mice showed a remarkably increased aggression toward the intruder male in a home cage or novelty environment. For instance, isolated mice presented a short attack latency (p < 0.001), high attack frequency (p < 0.001) and long attack duration (p < 0.001). In addition, isolated mice exhibited further social avoidance. Contrastingly, isolated mice displayed a reduced sexual preference for female (IS: 61.47 ± 13.80%, GH: 70.33 ± 10.06%, p = 0.038). As for heterosexual behavior, isolated mice have a short mating duration (p = 0.002), long mounting latency (p = 0.002), and long intromission latency (p = 0.015). However, no association was observed between postweaning isolation and homosexual behavior in male CD-1 mouse.Conclusion: Postweaning isolation increased the social affiliation, impaired the social cognition and considerably increased the aggression in social interaction of adult male CD-1 mice. Postweaning isolation induced a decreased sexual preference for female in adulthood. Postweaning isolation extended the latency to mate, thereby reducing mating behavior. No association was observed between isolation and homosexual behavior.

Post-weaning social isolation of rats induces reduction in the gene expression of vascular endothelial growth factor (VEGF) in the hippocampus.

The role of vascular endothelial growth factor (VEGF) in chronic stress and neurodevelopmental disorders is of growing research interest. Here we show that post-weaning isolation rearing of rats decreased gene expression of VEGF in the hippocampus. Gene expression of VEGF upstream regulator fibroblast growth factor-2 (FGF-2) or its downstream mediator endothelial nitric oxide synthase (eNOS) was unchanged. Other signaling pathways appear to be involved in isolation-induced reduction in VEGF gene expression. Sex differences in VEGF and eNOS gene expression with significantly higher mRNA levels in females than males were revealed.

Post-weaning social isolation of rats leads to long-term disruption of the gut microbiota-immune-brain axis

Early-life stress is an established risk for the development of psychiatric disorders. Post-weaning isolation rearing of rats produces lasting developmental changes in behavior and brain function that may have translational pathophysiological relevance to alterations seen in schizophrenia, but the underlying mechanisms are unclear. Accumulating evidence supports the premise that gut microbiota influence brain development and function by affecting inflammatory mediators, the hypothalamic–pituitaryadrenal axis and neurotransmission, but there is little knowledge of whether the microbiota-gut-brain axis might contribute to the development of schizophrenia-related behaviors. To this end the effects of social isolation (SI; a well-validated animal model for schizophrenia)-induced changes in rat behavior were correlated with alterations in gut microbiota, hippocampal neurogenesis and brain cytokine levels. Twenty-four male Lister hooded rats were housed in social groups (group-housed, GH, 3 littermates per cage) or alone (SI) from weaning (post-natal day 24) for four weeks before recording open field exploration, locomotor activity/novel object discrimination (NOD), elevated plus maze, conditioned freezing response (CFR) and restraint stress at one week intervals. Post-mortem caecal microbiota composition, cortical and hippocampal cytokines and neurogenesis were correlated to indices of behavioral changes. SI rats were hyperactive in the open field and locomotor activity chambers traveling further than GH controls in the less aversive peripheral zone. While SI rats showed few alterations in plus maze or NOD they froze for significantly less time than GH following conditioning in the CFR paradigm, consistent with impaired associative learning and memory. SI rats had significantly fewer BrdU/NeuN positive cells in the dentate gyrus than GH controls. SI rats had altered microbiota composition with increases in Actinobacteria and decreases in the class Clostridia compared to GH controls. Differences were also noted at genus level. Positive correlations were seen between microbiota, hippocampal IL-6 and IL-10, conditioned freezing and open field exploration. Adverse early-life stress resulting from continuous SI increased several indices of ‘anxiety-like’ behavior and impaired associative learning and memory accompanied by changes to gut microbiota, reduced hippocampal IL-6, IL-10 and neurogenesis. This study suggests that early-life stress may produce long-lasting changes in gut microbiota contributing to development of abnormal neuronal and endocrine function and behavior which could play a pivotal role in the aetiology of psychiatric illness.

Effect of post-weaning isolation on anxiety- and depressive-like behaviors of C57BL/6J mice.

Effects of post-weaning isolation on depressive- and anxiety-like behaviors in rodents have been well studied in the past. However, few studies included both sexes in a single experiment to study the sex difference in this animal model. The present study investigated the effect of post-weaning isolation on anxiety- and depressive-like behaviors in both male and female C57BL/6J mice. Mice were individually or grouped housed from postnatal day 21 for 5weeks until behavioral tests began. The results showed that social isolation resulted in increased anxiety in the open field. Isolated-reared female, but not male mice showed an increased transition between two compartments in the light-dark box and a decreased immobile time in the forced swim test. We conclude that post-weaning isolation has a sex-specific effect on emotional behaviors.

Effects of social deprivation on social and depressive-like behaviors and the numbers of oxytocin expressing neurons in rats

Social isolation is a known stressor that negatively impacts the well-being of social species. In rodents, social deprivation experienced either before or after weaning profoundly impacts adult behavioral and neuroendocrine profiles. This study compared the effects of post-natal and post-weaning social deprivation on behavioral profiles and hypothalamic oxytocin (OT) neurons. Male and female Sprague–Dawley rats were assigned to two post-natal groups, maternally separated (MS) or non-MS. MS pups were separated from their mothers for 4h daily during post-natal days 2–21 while non-MS litters remained undisturbed. Animals were then weaned and assigned to single or group housing conditions (SH/GH). Social behaviors were evaluated two weeks later and at 2–3 months of age, depressive-like behavioral profiles were assessed using the forced swim and sucrose preference tests. Animals were euthanized, and hypothalamic OT neurons were quantified. Post-weaning isolation significantly impacted behavioral profiles, with SH animals displaying more social behaviors than GH animals. SH animals also exhibited more immobility behavior in the forced swim test and a decreased sucrose preference. Effects of sex and MS were relatively limited. Correlation analyses revealed an inverse relationship between the display of antagonistic social behaviors and the numbers of OT cells in the anterior parvicellular division of the paraventricular nucleus (PVNap). There were no correlations between numbers of OT neurons and prosocial or depressive-like behaviors. Our results demonstrate a rapid and persistent disruption of behaviors in SH animals and suggest that some of these effects may be associated with numbers of OT neurons in the PVNap.

Social isolation mediated anxiety like behavior is associated with enhanced expression and regulation of BDNF in the female mouse brain

Adverse early life experience is prominent risk factors for numerous psychiatric illnesses, including mood and anxiety disorders. It imposes serious long-term costs on the individual as well as health and social systems. Hence, developing therapies that prevent the long-term consequences of early life stress is of utmost importance, and necessitates a better understanding of the mechanisms by which early life stress triggers long-lasting alterations in gene expression and behavior. Post-weaning isolation rearing of rodents models the behavioral consequences of adverse early life experiences in humans and it is reported to cause anxiety like behavior which is more common in case of females. Therefore, in the present study, we have studied the impact of social isolation of young female mice for 8weeks on the anxiety like behavior and the underlying molecular mechanism. Elevated plus maze and open field test revealed that social isolation caused anxiety like behavior. BDNF, a well-known molecule implicated in the anxiety like behavior, was up-regulated both at the message and protein level in cerebral cortex by social isolation. CREB-1 and CBP, which play a crucial role in BDNF transcription, were up-regulated at mRNA level in cerebral cortex by social isolation. HDAC-2, which negatively regulates BDNF expression, was down-regulated at mRNA and protein level in cerebral cortex by social isolation. Furthermore, BDNF acts in concert with Limk-1, miRNA-132 and miRNA-134 for the regulation of structural and morphological plasticity. Social isolation resulted in up-regulation of Limk-1 mRNA and miRNA-132 expression in the cerebral cortex. MiRNA-134, which inhibits the translation of Limk-1, was decreased in cerebral cortex by social isolation. Taken together, our study suggests that social isolation mediated anxiety like behavior is associated with up-regulation of BDNF expression and concomitant increase in the expression of CBP, CREB-1, Limk-1 and miRNA-132, and decrease in the expression of HDAC-2 and miRNA-134 in the cerebral cortex.