Consequences of Post-Weaning Social Isolation on Anxiety Behavior and Related Neural Circuits in Rodents

Exposure to adverse experiences in early-life is implicated in the later vulnerability to development of psychiatric disorders, including anxiety and affective disorders in humans. Adverse early-life experiences likely impart their long-term consequences on mental health by disrupting the normal development of neural systems involved in stress responses, emotional behavior and emotional states. Neural systems utilizing the neurotransmitters serotonin, dopamine and the neuropeptide corticotropin-releasing factor (CRF) are implicated in mediating emotive behaviors, and dysfunction of these neurochemical systems is associated with mood/anxiety disorders. These neural systems continue maturing until early or mid-adolescence in humans, thus alterations to their development are likely to contribute to the long-term consequences of adverse early-life experiences. A large body of literature suggests that post-weaning isolation rearing of rodents models the behavioral consequences of adverse early-life experiences in humans. Overall, the majority findings suggest that post-weaning social isolation that encompasses pre-adolescence produces long-lasting alterations to anxiety behavior, while measures of monoaminergic activity in various limbic regions during social isolation suggest alterations to dopamine and serotonin systems. The goal of this review is to evaluate and integrate findings from post-weaning social isolation studies specifically related to altered fear and anxiety behaviors and associated changes in neuroendocrine function and the activity of monoaminergic systems

Listening to your partner: serotonin increases male responsiveness to female vocal signals in mice

The context surrounding vocal communication can have a strong influence on how vocal signals are perceived. The serotonergic system is well-positioned for modulating the perception of communication signals according to context, because serotonergic neurons are responsive to social context, influence social behavior, and innervate auditory regions. Animals like lab mice can be excellent models for exploring how serotonin affects the primary neural systems involved in vocal perception, including within central auditory regions like the inferior colliculus (IC). Within the IC, serotonergic activity reflects not only the presence of a conspecific, but also the valence of a given social interaction. To assess whether serotonin can influence the perception of vocal signals in male mice, we manipulated serotonin systemically with an injection of its precursor 5-HTP, and locally in the IC with an infusion of fenfluramine, a serotonin reuptake blocker. Mice then participated in a behavioral assay in which males suppress their ultrasonic vocalizations (USVs) in response to the playback of female broadband vocalizations (BBVs), used in defensive aggression by females when interacting with males. Both 5-HTP and fenfluramine increased the suppression of USVs during BBV playback relative to controls. 5-HTP additionally decreased the baseline production of a specific type of USV and male investigation, but neither drug treatment strongly affected male digging or grooming. These findings show that serotonin modifies behavioral responses to vocal signals in mice, in part by acting in auditory brain regions, and suggest that mouse vocal behavior can serve as a useful model for exploring the mechanisms of context in human communication

Early-age stress exposure and psychopathology development: vulnerability mechanisms and resilience

Stress is a general term describing the altered physiological response of humans or animals to any physical, emotional or mental request. Stress could be “acute”, if it occurs for a short time period, or “chronic” if it lasts for a long time, finally resulting in a maladaptive response with harmful effects on the body. In this context, several lines of evidence have shown that exposure to chronic stress, especially during early phases of life in which key neurodevelopmental steps occur, is significantly involved in the onset and progression of different psychiatric disorders, including psychosis. Moreover, it has been reported that psychotic patients frequently experience other mental disturbances, such as anxiety and depression. However, so far, mechanisms underlying psychosis comorbidities have not been clarified. Furthermore, while the detrimental effects of the exposure to stressful events during pregnancy have been extensively investigated, the impact of preconceptional chronic stress exposure on the offspring still remains unexplored. In this context, animal models of chronic stress- induced psychiatric disorders are very useful research tools. Here, we used an environmental animal model of chronic stress exposure, i.e. the rat social isolation model which has been widely described to induce behavioural, neurochemical, neuroendocrine, metabolic and biomolecular alterations reminiscent of what observed in psychotic patients. In particular, in the first part of this study we investigated whether exposure to social isolation from weaning (postnatal day-PND21) to young adulthood (PND70) could induce depressive- and anxiety-like behaviours, as well as neurochemical and neuroendocrine alterations in rats. In the second part of the study, we evaluated whether female exposure to social isolation before pregnancy could determine cognitive, neurochemical and redox-related dysfunctions in the offspring, reared, in its turn, in group or in isolation condition from PND21 to PND70. Results of the first part of the study showed that socially isolated rats developed a depressive-like behaviour, as demonstrated by the significant increase in the immobility frequency and the reduction of the swimming frequency in the forced swimming test. Moreover, exposure to social isolation also induced an anxious-like state, as evidenced in the elevated zero maze test by a significant decrease in the time spent in the open corridors and an enhanced time spent in the closed ones, and in the open field test by the elevation of the grooming time. Those behavioural alterations were accompanied by a significant decrease of noradrenaline (NA) and serotonin (5- HT) levels, as well as an increase of 5-HT turnover, in amygdala. Seven weeks of isolation also led to a reduction of gamma aminobutyric acid (GABA) amount and an elevation of glutamate (GLU) amount in amygdala and hippocampus. Following social isolation exposure, central neurochemical alterations also reflected at peripheral levels, with a significant increase of kynurenine and a reduction of 5-HT concentrations. Neurochemical alterations were accompanied by a decrease of plasmatic oxytocin, prolactin, ghrelin and melatonin amounts, while plasmatic levels of Neuropeptide S were not modified. Results of the second part of the study showed that the offspring of socially isolated females developed cognitive alterations, as evidenced in the novel object recognition test, where it was not able to discriminate between a familiar and a novel object, and in the passive avoidance task, in which it showed a significant reduction of the latency time. Behavioural dysfunctions were accompanied by increased NA levels and reduced 5-HT amounts in the prefrontal cortex. In this same area, a significant decrease of GABA levels was found, whereas no significant differences in GLU content were observed. Furthermore, chronic social stress exposure led to redox-related alterations in terms of cortical increase of reactive oxygen species amount, reduced expression of Nf-kB, NOX2 and NOX1, as well as altered expression of antioxidant enzymes, such as SOD1 and CoQ10. In conclusion, our findings provide novel insights in the understanding of the neurochemical and neuroendocrine alterations underlying the comorbidity among different stress-induced neuropsychiatric disorders, such as psychosis, anxiety and depression. In addition to this, preconceptional exposure to chronic social stress might have a profound influence on the offspring neurodevelopment in terms of cognitive, neurochemical and redox-related alterations. Therefore, our results open new perspectives for targeted therapies acting on neurobiological pathways, also identifying specific time points for possible preventive and therapeutic strategies against chronic stress-induced neuropsychiatric disorders

Stress in critical period of early life influences glutamate excitatory neurotransmission: implication for neuropsychiatric disorders

Early adverse events occurring during critical periods of brain development may shape the individual’s developmental trajectory and increase vulnerability to stress-related disorders across lifespan. The aim of my PhD thesis was to investigate the role of glutamatergic neurotransmission in the early-life stress induced impairments of emotional, cognitive, and social scaffolding in functionally involved brain regions. The study was realized by using the model of postweaning social isolation (PWSI) in mice and the model of perinatal stress (PRS) in rat. Firstly, I compared the behavioral and biochemical profiles of PWSI-induced inbred C57BL/6 N mice to those of BTBR mice, a rodent model used to study autism spectrum disorders in humans. Male C57BL/6 N mice were socially housed at weaning (postnatal day 21) or isolated for four weeks before being subjected to experimental analysis at 48 days of age. Male BTBR mice were socially housed at weaning and subjected to analysis at the same age. Metabotropic glutamate receptor of type 2 (mGluR2), glucocorticoid and mineralocorticoid receptors levels were all decreased in the hippocampus of PWSI and BTBR animals. Moreover, both PWSI mice and BTBR mice displayed decreased social behavior (social investigation and ultrasonic vocalizations), demonstrating that the lack of social stimuli throughout adolescence causes an endophenotype that mirrors some behavioral features of autism spectrum disorders. Secondly, I investigated the possible corrective role exerted by resocialization on the PWSI-induced hippocampal glutamatergic disequilibrium. Results showed that PWSI-induced reduction of hippocampal mGluR2 was not recovered by one week of resocialization. I have also investigated the effect of PRS in adult and aged rats of both sexes on the expression of AMPA and GABAA receptor subunits in the hippocampus (ventral and dorsal) and prefrontal cortex in relation to emotional and cognitive behaviors. PRS induced sex-dimorphic and age-dependent effects on some receptor subunits and behavior. Particularly, in both adulthood and ageing, PRS reduced open-arm exploration and recognition score in males, while it improved them in females. Interestingly, in the dorsal hippocampus, PRS reduced the expression of the GluA2 subunit in adult male rats and increased the expression in adult female rats. Moreover, PRS reduced the expression of the GluA3 AMPA receptor subunit in the prefrontal cortex and in dorsal hippocampus of adult male rats, an effect which was limited to the prefrontal cortex of adult female rats. Remarkably, changes in GluA2/GluA3 subunits and behavior induced by PRS persisted in aged male rats, but not females. The α1 subunit of the pentameric GABAA receptor was also studied in adult and aged rats of both sexes. PRS enhanced the expression of the α1 subunit of GABAA in the dorsal hippocampus of both sexes and reduced it in the prefrontal cortex exclusively in females. In aged PRS subjects a reduction of GABAA-α1 subunit protein levels in the prefrontal cortex was observed in the male gender. Extending the analysis to synaptic vesicle proteins we found reduction of synaptophysin, syntaxin and rab3a levels in the ventral hippocampus of PRS males, but not females, at adulthood and ageing. Interestingly, control male rats are characterized by a greater density of hippocampal levels of syntaxin, munc-18, synapsin IIa, VAMP, synaptophysin and rab3a with respect to female rats. This difference was abolished by PRS, suggesting that PRS caused dysmasculinization across lifespan. These findings suggest that changes in the expression levels of AMPA and GABAA receptors contribute to the sex-divergent behavioral phenotype induced by PRS in adult and aged rats. Collectively, my research shows that early-life stress-induced alterations converge to severe impairment of glutamatergic neurotransmission, underscoring its key role in determining and maintaining the proper developmental trajectory across the lifespan

Effects of long-term individual housing of middle-aged female Octodon degus on spatial learning and memory in the Barnes maze task

©2023 Popović, Baño-Otalora, Rol, Venero, Madrid and Popović. This manuscript version is made available under the CC-BY 4.0 license http://creativecommons.org/licenses/by/4.0/. This document is the Published version of a Published Work that appeared in final form in Frontiers in Behavioral Neuroscience. To access the final edited and published work see https://doi.org/10.3389/fnbeh.2023.1221090Prolonged social isolation is a form of passive chronic stress that has consequences on human and animal behavior. The present study was undertaken to elucidate whether the long-term isolation would precipitate age-related changes in anxiety and spatial learning and memory in degus. Methods: We investigated the effects of long-term social isolation on anxiety levels in the light-dark test, and spatial orientation abilities in the Barnes maze. Middle-aged female Octodon degus were allocated to either group-housed (3 animals per cage) or individually-housed for 5 months. Results: Under this experimental condition, there were no significant group differences in the anxiety level tested in the light-dark test and in the motivation to escape from the Barnes maze. There were no significant differences in cortisol levels between individually- and group-housed animals. On the last acquisition training day of spatial learning, individually- housed animals had a significantly higher number of correct responses and a smaller number of reference and working memory errors than the group-housed animals. In addition, isolated animals showed a tendency for reference and working memory impairment on the retention trial, while group-housed degus showed improvement in these parameters. Discussion and conclusion: The present study indicates that prolonged social isolation during adulthood in female degus has a dual effect on spatial orientation. Specifically, it results in a significant improvement in acquisition skills but a slight impairment in memory retention. The obtained cognitive changes were not accompanied by modification in anxiety and cortisol levels. Keywords: Barnes maze; Octodon degus; anxiety; learning and memory; light-dark test; social isolation

THE EFFECTS OF EARLY LIFE STRESS ON LONG-TERM POTENTIATION IN PATHWAY FROM THE MEDIAL PREFRONTAL CORTEX TO THE BASOLATERAL AMYGDALA

A leading neurocircuitry model of emotional regulation points to the pathway from the medial prefrontal cortex (mPFC) to the basolateral amygdala (BLA). This pathway has been implicated in fear conditioning and extinction studies and its malfunction is hypothesized to underlie affective disorders such as PTSD and anxiety. Interestingly, the mPFC-BLA pathway shows delayed maturation in both humans and rats, rendering it vulnerable to early life stress (ELS). Indeed, several studies have linked ELS to emotional dysregulation as well as changes in the amygdala and PFC. However, no study has ever been done on the effect of ELS on long-term potentiation (LTP) in this pathway. In fact, very few studies on LTP in the mPFC-BLA pathway have been conducted at all which is surprising given LTP’s role in learning and memory and given the mPFC-BLA pathway’s proposed role in fear conditioning/extinction. Therefore, using electrophysiological methods in awake, freely behaving rats, the current study examined whether ELS in the form of neonatal isolation (ISO) affects LTP in the mPFC-BLA pathway. Results indicate that the mPFC-BLA pathway is resistant to LTP in both control and ISO rats following both sustained and theta burst high frequency stimulation (HFS). In fact, rats showed a tendency toward long-term depression (LTD) especially following sustained stimulation at 200 Hz. Small sample sizes prevented a meaningful comparison of LTD across ISO and control groups

Calbindin Deficits May Underlie Dissociable Effects of 5-HT6 and mGlu7 Antagonists on Glutamate and Cognition in a Dual-Hit Neurodevelopmental Model for Schizophrenia

© 2020, The Author(s). Despite several compounds entering clinical trials for the negative and cognitive symptoms of schizophrenia, few have progressed beyond phase III. This is partly attributed to a need for improved preclinical models, to understand disease and enable predictive evaluation of novel therapeutics. To this end, one recent approach incorporates “dual-hit” neurodevelopmental insults like neonatal phencyclidine plus isolation rearing (PCP-Iso). Glutamatergic dysfunction contributes to schizophrenia pathophysiology and may represent a treatment target, so we used enzyme-based microsensors to evaluate basal- and drug-evoked glutamate release in hippocampal slices from rats that received neonatal PCP and/or isolation rearing. 5-HT6 antagonist-evoked glutamate release (thought to be mediated indirectly via GABAergic disinhibition) was reduced in PCP-Iso, as were cognitive effects of a 5-HT6 antagonist in a hippocampal glutamate-dependent novel object discrimination task. Yet mGlu7 antagonist-evoked glutamatergic and cognitive responses were spared. Immunohistochemical analyses suggest these findings (which mirror the apparent lack of clinical response to 5-HT6 antagonists in schizophrenia) are not due to reduced hippocampal 5-HT input in PCP-Iso, but may be explained by reduced calbindin expression. This calcium-binding protein is present in a subset of GABAergic interneurons receiving preferential 5-HT innervation and expressing 5-HT6 receptors. Its loss (in schizophrenia and PCP-Iso) would be expected to reduce interneuron firing and potentially prevent further 5-HT6 antagonist-mediated disinhibition, without impacting on responses of VIP-expressing interneurons to mGlu7 antagonism. This research highlights the importance of improved understanding for selection of appropriate preclinical models, especially where disease neurobiology impacts on cells mediating the effects of potential therapeutics

Inhibition of prandial and waterspray-induced rat grooming by 8-OH-DPAT

The effects of 8-OH-DPAT treatment on rat grooming behaviour, elicited either prandially or in response to spraying with water were investigated. Dose (≤0.1 mg/kg s.c.) response studies employed momentary time sampling over 30 or 60 min with behaviour being scored in one of 6 or 7 (depending on food availability) mutually exclusive categories (feeding, active, scratching, face-grooming, body grooming, genital-grooming and resting) at 15 s intervals. In non-deprived rats, tested with wet mash available, feeding and activity frequencies were increased, but resting and total grooming were inhibited by 8-OH-DPAT. Face-, body- and genital-grooming occurred at higher levels than scratching, but all categories were reduced with reductions in scratching occurring at a lower dose (0.01 mg/kg). Misting rats with a fine water spray selectively increased body grooming and decreased activity without altering feeding, while 8-OH-DPAT increased feeding and reduced face-, body- and genital-grooming, without affecting already low levels of scratching. In misted rats, tested without food, 8-OH-DPAT reduced face-, body- and genital-grooming and increased resting. These results confirm i) that the water spray technique is a useful method for increasing grooming and ii) that 8-OH-DPAT has a suppressant effect on grooming independent of response competition from enhanced feeding

Adolescent Alcohol Exposure Persistently Impacts Adult Neurobiology and Behavior

Adolescence is a developmental period when physical and cognitive abilities are optimized, when social skills are consolidated, and when sexuality, adolescent behaviors, and frontal cortical functions mature to adult levels. Adolescents also have unique responses to alcohol compared with adults, being less sensitive to ethanol sedative–motor responses that most likely contribute to binge drinking and blackouts. Population studies find that an early age of drinking onset correlates with increased lifetime risks for the development of alcohol dependence, violence, and injuries. Brain synapses, myelination, and neural circuits mature in adolescence to adult levels in parallel with increased reflection on the consequence of actions and reduced impulsivity and thrill seeking. Alcohol binge drinking could alter human development, but variations in genetics, peer groups, family structure, early life experiences, and the emergence of psychopathology in humans confound studies. As adolescence is common to mammalian species, preclinical models of binge drinking provide insight into the direct impact of alcohol on adolescent development. This review relates human findings to basic science studies, particularly the preclinical studies of the Neurobiology of Adolescent Drinking in Adulthood (NADIA) Consortium. These studies focus on persistent adult changes in neurobiology and behavior following adolescent intermittent ethanol (AIE), a model of underage drinking. NADIA studies and others find that AIE results in the following: increases in adult alcohol drinking, disinhibition, and social anxiety; altered adult synapses, cognition, and sleep; reduced adult neurogenesis, cholinergic, and serotonergic neurons; and increased neuroimmune gene expression and epigenetic modifiers of gene expression. Many of these effects are specific to adolescents and not found in parallel adult studies. AIE can cause a persistence of adolescent-like synaptic physiology, behavior, and sensitivity to alcohol into adulthood. Together, these findings support the hypothesis that adolescent binge drinking leads to long-lasting changes in the adult brain that increase risks of adult psychopathology, particularly for alcohol dependence