Stress-responsive Brain Regions Research Articles

Evidence for a role of the basolateral amygdala in regulating regional metabolism in the stressed brain.

The brain regulates every physiological process in the body, including metabolism. Studies investigating brain metabolism have shown that stress can alter major metabolic processes, and that these processes can vary between regions. However, no study has investigated how metabolic pathways may be altered by stressor perception, or whether stress-responsive brain regions can also regulate metabolism. The basolateral amygdala (BLA), a region important for stress and fear, has reciprocal connections to regions responsible for metabolic regulation. In this study, we investigated how BLA influences regional metabolic profiles within the hippocampus (HPC) and medial prefrontal cortex (mPFC), regions involved in regulating the stress response and stress perception, using optogenetics in male C57BL/6 mice during footshock presentation in a yoked shuttlebox paradigm based on controllable (ES) and uncontrollable (IS) stress. RNA extracted from HPC and mPFC were loaded into NanoString® Mouse Neuroinflammation Panels, which also provides a broad view of metabolic processes, for compilation of gene expression profiles. Results showed differential regulation of carbohydrate and lipid metabolism, and insulin signaling gene expression pathways in HPC and mPFC following ES and IS, and that these differences were altered in response to optogenetic excitation or inhibition of the BLA. These findings demonstrate for the first time that individual brain regions can utilize metabolites in a way that are unique to their needs and function in response to a stressor, and that vary based on stressor controllability and influence by BLA.

Complement C5a Receptor Signaling Alters Stress Responsiveness and Modulates Microglia Following Chronic Stress Exposure

BackgroundAccumulating evidence underscores the pivotal role of heightened inflammation in the pathophysiology of stress-related diseases, but the underlying mechanisms remain elusive. The complement system, a key effector of the innate immune system, produces the C5–cleaved activation product C5a upon activation, initiating inflammatory responses through the canonical C5a receptor 1 (C5aR1). While C5aR1 is expressed in stress-responsive brain regions, its role in stress responsiveness remains unknown. MethodsTo investigate C5a-C5aR1 signaling in stress responses, mice underwent acute and chronic stress paradigms. Circulating C5a levels and messenger RNA expression of C5aR1 in the hippocampus and adrenal gland were measured. C5aR1-deficient mice were used to elucidate the effects of disrupted C5a-C5aR1 signaling across behavioral, hormonal, metabolic, and inflammation parameters. ResultsChronic restraint stress elevated circulating C5a levels while reducing C5aR1 messenger RNA expression in the hippocampus and adrenal gland. Notably, the absence of C5aR1 signaling enhanced adrenal sensitivity to adrenocorticotropic hormone, concurrently reducing pituitary adrenocorticotropic hormone production and enhancing the response to acute stress. C5aR1-deficient mice exhibited attenuated reductions in locomotor activity and body weight under chronic stress. Additionally, these mice displayed increased glucocorticoid receptor sensitivity and disrupted glucose and insulin homeostasis. Chronic stress induced an increase in C5aR1-expressing microglia in the hippocampus, a response mitigated in C5aR1-deficient mice. ConclusionsC5a-C5aR1 signaling emerges as a key metabolic regulator during stress, suggesting that complement activation and dysfunctional C5aR1 signaling may contribute to neuroinflammatory phenotypes in stress-related disorders. The results advocate for further exploration of complement C5aR1 as a potential therapeutic target for stress-related conditions.

Cortical-Hypothalamic Integration of Autonomic and Endocrine Stress Responses.

The prevalence and severity of cardiovascular disease (CVD) are exacerbated by chronic stress exposure. While stress-induced sympathetic activity and elevated glucocorticoid secretion impair cardiovascular health, the mechanisms by which stress-responsive brain regions integrate autonomic and endocrine stress responses remain unclear. This review covers emerging literature on how specific cortical and hypothalamic nuclei regulate cardiovascular and neuroendocrine stress responses. We will also discuss the current understanding of the cellular and circuit mechanisms mediating physiological stress responses. Altogether, the reviewed literature highlights the current state of stress integration research, as well unanswered questions about the brain basis of CVD risk.

Unexpected Role of Physiological Estrogen in Acute Stress-Induced Memory Deficits.

Stress may promote emotional and cognitive disturbances, which differ by sex. Adverse outcomes, including memory disturbances, are typically observed following chronic stress, but are now being recognized also after short events, including mass shootings, assault, or natural disasters, events that consist of concurrent multiple acute stresses (MAS). Prior work has established profound and enduring effects of MAS on memory in males. Here we examined the effects of MAS on female mice and probed the role of hormonal fluctuations during the estrous cycle on MAS-induced memory problems and the underlying brain network and cellular mechanisms. Female mice were impacted by MAS in an estrous cycle-dependent manner: MAS impaired hippocampus-dependent spatial memory in early-proestrous mice, characterized by high levels of estradiol, whereas memory of mice stressed during estrus (low estradiol) was spared. As spatial memory requires an intact dorsal hippocampal CA1, we examined synaptic integrity in mice stressed at different cycle phases and found a congruence of dendritic spine density and spatial memory deficits, with reduced spine density only in mice stressed during high estradiol cycle phases. Assessing MAS-induced activation of brain networks interconnected with hippocampus, we identified differential estrous cycle-dependent activation of memory- and stress-related regions, including the amygdala. Network analyses of the cross-correlation of fos expression among these regions uncovered functional connectivity that differentiated impaired mice from those not impaired by MAS. In conclusion, the estrous cycle modulates the impact of MAS on spatial memory, and fluctuating physiological levels of sex hormones may contribute to this effect.SIGNIFICANCE STATEMENT: Effects of stress on brain functions, including memory, are profound and sex-dependent. Acute stressors occurring simultaneously result in spatial memory impairments in males, but effects on females are unknown. Here we identified estrous cycle-dependent effects of such stresses on memory in females. Surprisingly, females with higher physiological estradiol experienced stress-induced memory impairment and a loss of underlying synapses. Memory- and stress-responsive brain regions interconnected with hippocampus were differentially activated across high and low estradiol mice, and predicted memory impairment. Thus, at functional, network, and cellular levels, physiological estradiol influences the effects of stress on memory in females, providing insight into mechanisms of prominent sex differences in stress-related memory disorders, such as post-traumatic stress disorder.

5-HT2A receptor loss does not alter acute fluoxetine-induced anxiety and exhibit sex-dependent regulation of cortical immediate early gene expression.

Background: Acute treatment with the selective serotonin reuptake inhibitor (SSRI), fluoxetine (Flx), induces anxiety-like behavioral effects. The serotonin2A receptor (5-HT2A) is implicated in the modulation of anxiety-like behavior, however its contribution to the anxiogenic effects of acute Flx remains unclear. Here, we examined the role of the 5-HT2A receptor in the effects of acute Flx on anxiety-like behavior, serum corticosterone levels, neural activation and immediate early gene (IEG) expression in stress-responsive brain regions, using 5-HT2A receptor knockout (5-HT2A−/−) mice of both sexes. Methods: 5-HT2A−/− and wild-type (WT) male and female mice received a single administration of Flx or vehicle, and were examined for anxiety-like behavior, serum corticosterone levels, FBJ murine osteosarcoma viral oncogene homolog peptide (c-Fos) positive cell numbers in stress-responsive brain regions of the hypothalamus and prefrontal cortex (PFC), and PFC IEG expression. Results: The increased anxiety-like behavior and enhanced corticosterone levels evoked by acute Flx were unaltered in 5-HT2A−/− mice of both sexes. 5-HT2A−/− female mice exhibited a diminished neural activation in the hypothalamus in response to acute Flx. Further, 5-HT2A−/− male, but not female, mice displayed altered baseline expression of several IEGs (brain-derived neurotrophic factor (Bdnf), Egr2, Egr4, FBJ osteosarcoma gene (Fos), FBJ murine osteosarcoma viral oncogene homolog B (Fosb), Fos-like antigen 2 (Fosl2), Homer scaffolding protein (Homer) 1-3 (Homer1-3), Jun proto-oncogene (Jun)) in the PFC. Conclusion: Our results indicate that the increased anxiety and serum corticosterone levels evoked by acute Flx are not influenced by 5-HT2A receptor deficiency. However, the loss of function of the 5-HT2A receptor alters the degree of neural activation of the paraventricular nucleus (PVN) of the hypothalamus in response to acute Flx, and baseline expression of several IEGs in the PFC in a sexually dimorphic manner.

Neuroinflammation Contributes to High Salt Intake-Augmented Neuronal Activation and Active Coping Responses to Acute Stress.

High dietary salt intake increases risk of stress-related neuropsychiatric disorders. Here, we explored the contribution of high dietary salt intake-induced neuroinflammation in key stress-responsive brain regions, the hypothalamic paraventricular nucleus and basolateral amygdala, in promoting exaggerated neuronal activation and coping behaviors in response to acute psychogenic stress. Mice that underwent high dietary salt intake exhibited increased active stress coping behaviors during and after an acute swim stress, and these were reduced by concurrent administration of minocycline, an inhibitor of microglial activation, without affecting body fluid hyperosmolality caused by high dietary salt intake. Moreover, minocycline attenuated high dietary salt intake-induced increases of paraventricular nucleus tumor necrosis factor-α, activated microglia (ionized calcium-binding adaptor molecule 1), and acute swim stress-induced neuronal activation (c-Fos). In the basolateral amygdala, similar effects were observed on ionized calcium-binding adaptor molecule 1+ and c-Fos+ counts, but not tumor necrosis factor-α levels. These data indicate that high dietary salt intake promotes neuroinflammation, increasing recruitment of neurons in key stress-associated brain regions and augmenting behavioral hyper-responsivity to acute psychological stress.

Genomic and epigenomic mechanisms of glucocorticoids in the brain.

Following the discovery of glucocorticoid receptors in the hippocampus and other brain regions, research has focused on understanding the effects of glucocorticoids in the brain and their role in regulating emotion and cognition. Glucocorticoids are essential for adaptation to stressors (allostasis) and in maladaptation resulting from allostatic load and overload. Allostatic overload, which can occur during chronic stress, can reshape the hypothalamic-pituitary-adrenal axis through epigenetic modification of genes in the hippocampus, hypothalamus and other stress-responsive brain regions. Glucocorticoids exert their effects on the brain through genomic mechanisms that involve both glucocorticoid receptors and mineralocorticoid receptors directly binding to DNA, as well as by non-genomic mechanisms. Furthermore, glucocorticoids synergize both genomically and non-genomically with neurotransmitters, neurotrophic factors, sex hormones and other stress mediators to shape an organism's present and future responses to a stressful environment. Here, we discuss the mechanisms of glucocorticoid action in the brain and review how glucocorticoids interact with stress mediators in the context of allostasis, allostatic load and stress-induced neuroplasticity.

Stress Increases Peripheral Axon Growth and Regeneration through Glucocorticoid Receptor-Dependent Transcriptional Programs.

Stress and glucocorticoid (GC) release are common behavioral and hormonal responses to injury or disease. In the brain, stress/GCs can alter neuron structure and function leading to cognitive impairment. Stress and GCs also exacerbate pain, but whether a corresponding change occurs in structural plasticity of sensory neurons is unknown. Here, we show that in female mice (Mus musculus) basal GC receptor (Nr3c1, also known as GR) expression in dorsal root ganglion (DRG) sensory neurons is 15-fold higher than in neurons in canonical stress-responsive brain regions (M. musculus). In response to stress or GCs, adult DRG neurite growth increases through mechanisms involving GR-dependent gene transcription. In vivo, prior exposure to an acute systemic stress increases peripheral nerve regeneration. These data have broad clinical implications and highlight the importance of stress and GCs as novel behavioral and circulating modifiers of neuronal plasticity.

Anxiety and Epigenetics.

Anxiety disorders are highly prevalent psychiatric disorders often comorbid with depression and substance abuse. Twin studies have shown that anxiety disorders are moderately heritable. Yet, genome-wide association studies (GWASs) have failed to identify gene(s) significantly associated with diagnosis suggesting a strong role for environmental factors and the epigenome. A number of anxiety disorder subtypes are considered "stress related." A large focus of research has been on the epigenetic and anxiety-like behavioral consequences of stress. Animal models of anxiety-related disorders have provided strong evidence for the role of stress on the epigenetic control of the hypothalamic-pituitary-adrenal (HPA) axis and of stress-responsive brain regions. Neuroepigenetics may continue to explain individual variation in susceptibility to environmental perturbations and consequently anxious behavior. Behavioral and pharmacological interventions aimed at targeting epigenetic marks associated with anxiety may prove fruitful in developing treatments.

Abstract # 1804 Characterization of microglia in multiple brain regions of dominant and subordinate mice

Previous research demonstrates that microglia are activated by both chronic and acute stress, and that the type, intensity, and duration of the stressor may lead to variable alterations in morphology and markers of inflammation. To examine microglia differences in the brains of mice living in an ethologically relevant environment, 12 adult male CD1 mice were group housed for 21 days in an enriched enclosure measuring 150 × 150 × 80 cm. Mice were observed and social interactions were recorded for a minimum of two hours each day during the dark cycle. Hierarchy status was determined by analyzing the relative proportion of aggression given and received by each animal. Paraformaldehyde-fixed frozen brain tissue from the three most dominant and three most subordinate mice from seven cohorts was cryosectioned (50 microns), immunolabeled with Iba-1 antibody and stained with either fluorescence or DAB-peroxidase, and photographed using a 40X objective. Eight stress-responsive brain regions were analyzed, namely, CA1, medial amygdala, nucleus accumbens core and shell, ventral striatum, medial prefrontal cortex, ventral tegmental area, and substantia nigra. To assess differences in each brain region we considered cell number, size, and overall morphological state score on a scale of 1–4, with 1 being ramified and 4 being amoeboid. This study serves to characterize regional differences in microglia number and morphology with consideration to individual dominance ranking within a naturalistic environment.

The Pharmacology of Visceral Pain.

Visceral pain describes pain emanating from the internal thoracic, pelvic, or abdominal organs. Unlike somatic pain, visceral pain is generally vague, poorly localized, and characterized by hypersensitivity to a stimulus such as organ distension. While current therapeutics provides some relief from somatic pain, drugs used for treatment of chronic visceral pain are typically less efficacious and limited by multiple adverse side effects. Thus, the treatment of visceral pain represents a major unmet medical need. Further, more basic research into the physiology and pathophysiology of visceral pain is needed to provide novel targets for future drug development. In concert with chronic visceral pain, there is a high comorbidity with stress-related psychiatric disorders including anxiety and depression. The mechanisms linking visceral pain with these overlapping comorbidities remain to be elucidated. However, persistent stress facilitates pain perception and sensitizes pain pathways, leading to a feed-forward cycle promoting chronic visceral pain disorders. We will focus on stress-induced exacerbation of chronic visceral pain and provide supporting evidence that centrally acting drugs targeting the pain and stress-responsive brain regions may represent a valid target for the development of novel and effective therapeutics.

Differences in microglia morphology are associated with social hierarchy status and social stress

Microglia are resident immune cells of the central nervous system, are activated by neuronal damage due to injury, infection, or stress, and assume a spectrum of morphological states when stimulated. In their activated state microglia are responsible for perpetuating inflammation and neurotoxicity. In the present study, we housed six cohorts of 12 adult male CD-1 mice in an enriched, naturalistic environment for 21 days. Extensive observations of the socially dominant (fighting, chasing, mounting) and subordinate (fleeing, freezing) behaviors of animals were undertaken. We were able to identify the rank of each animal within each social hierarchy dependent upon its relative proportion of aggressive and subordinate behaviors expressed and received. After behavioral characterization, animals were perfused for further brain analysis. We examined microglia morphology using immunohistological techniques in three stress-responsive brain regions (striatum, CA1, medial amygdala) from the top three dominant and three most subordinate mice in each cohort. We describe the association between social status and chronic social stress experienced by mice living in a naturalistic context with individual variation in microglia morphological states.

Stress-induced pain: a target for the development of novel therapeutics.

Although current therapeutics provide relief from acute pain, drugs used for treatment of chronic pain are typically less efficacious and limited by adverse side effects, including tolerance, addiction, and gastrointestinal upset. Thus, there is a significant need for novel therapies for the treatment of chronic pain. In concert with chronic pain, persistent stress facilitates pain perception and sensitizes pain pathways, leading to a feed-forward cycle promoting chronic pain disorders. Stress exacerbation of chronic pain suggests that centrally acting drugs targeting the pain- and stress-responsive brain regions represent a valid target for the development of novel therapeutics. This review provides an overview of how stress modulates spinal and central pain pathways, identifies key neurotransmitters and receptors within these pathways, and highlights their potential as novel targets for therapeutics to treat chronic pain.

Differential changes in amygdala and frontal cortex Pde10a expression during acute and protracted withdrawal.

Alcohol use disorders are persistent problems with high recidivism rates despite repeated efforts to quit drinking. Neuroadaptations that result from alcohol exposure and that persist during periods of abstinence represent putative molecular determinants of the propensity to relapse. Previously we demonstrated a positive association between phosphodiesterase 10A (PDE10A) gene expression and elevations in relapse-like alcohol self-administration in rats with a history of stress exposure. Because alcohol withdrawal is characterized by heightened anxiety-like behavior, activation of stress-responsive brain regions and an elevated propensity to self-administer alcohol, we hypothesized that Pde10a expression also would be upregulated in reward- and stress-responsive brain regions during periods of acute (8–10 h) and protracted (6 weeks) alcohol withdrawal. During acute withdrawal, elevated Pde10a mRNA expression was found in the medial and basolateral amygdala (BLA), as well as the infralimbic and anterior cingulate subdivisions of the medial prefrontal cortex, relative to alcohol-naïve controls. The BLA was the only region with elevated Pde10a mRNA expression during both acute and protracted withdrawal. In contrast to the elevations, Pde10a mRNA levels tended to be reduced during protracted withdrawal in the dorsal striatum, prelimbic prefrontal cortex, and medial amygdala. Together these results implicate heightened PDE10A expression in the BLA as a lasting neuroadaptation associated with alcohol dependence.

Sexually dimorphic patterns of neural activity in response to juvenile social subjugation

After experiencing juvenile social subjugation (JSS), adult female rats display more severe depression- and anxiety-like behaviors than adult males, suggesting that JSS is encoded in a sex-specific manner. To test this hypothesis, prepubertal rats (P28–33) were subjected to 10 aggressive acts in ≤10min from an aggressive adult male, a 10min encounter with a non-aggressive adult male, or to 10min in an empty, clean cage (handled control) and were sacrificed one hour later. We then used unbiased stereology to estimate the total number and proportion of neurons immunoreactive for the immediate early gene product Fos bilaterally in the basolateral amygdala (BLA), the anterior and posterior subdivisions of the bed nucleus of the stria terminalis, and the paraventricular nucleus of the hypothalamus (PVN). Overall, females’ Fos responses were less selective than males’. The BLA in males displayed a selective Fos response to the non-aggressive male, whereas no such selectivity occurred in the BLA of females. Additionally, there were more neurons overall in the left BLA than the right and this lateralization was specific to males. The principal subdivision of the BST (BSTpr) in males responded selectively to JSS, whereas the BSTpr in females responded to both the non-aggressive and aggressive males. We also found that the regional volume and neuron number of the BSTpr is greater in males than in females. Finally, the PVN in males was, like the BLA, selective for the non-aggressive male, whereas none of the experiences elicited a selective response in females. The greater selectivity for non-threatening stimuli in males in three stress-responsive brain regions may be a clue as to why males are less susceptible to the anxiogenic effects of JSS.

An unexpected increase in restraint duration alters the expression of stress response habituation

While habituation develops to a repeated psychological stressor, manipulating certain parameters of the stress challenge experience may lead to dishabituation of the stress response. In this experiment, we investigated whether the behavioral, endocrine, and neural responses (c-fos mRNA immediate early gene expression) to a psychological stressor (restraint) differ when the duration of the stressor given on the test day violates expectations based on prior stress experience. Rats experienced 10min of daily restraint on Days 1–4 followed by a challenge with either the same duration (10min) or a longer duration (30min) of restraint on Day 5. Rats' behavior was video recorded during the Day 5 restraint episode, and trunk blood and brain tissue were collected 30min following restraint onset. Struggling behavior was manually scored as active attempts to escape the restraint device. Rats who experienced the same duration of repeated restraint showed a significant decrease of plasma corticosterone (CORT) compared to the 10min acute restraint group (habituation). In addition, these rats showed decreased active struggling over repeated restraint trials. Conversely, the rats showed an increased CORT response (dishabituation) when they experienced a longer duration of restraint on Day 5 than they had previously. These rats showed a habituated behavioral response during the first 10min of restraint, however struggling behavior increased once the duration of restraint exceeded the expected duration (with a peak at 12min). This peak in struggling behavior did not occur during 30min acute restraint, indicating that the effect was related to the memory of previous restraint experience and not due to a longer duration of restraint. In contrast, these animals showed habituated c-fos mRNA expression in the paraventricular nucleus (PVN), lateral septum (LS), and medial prefrontal cortex (mPFC) in response to the increased stressor duration. Thus, there was a dissociation between c-fos mRNA expression in key stress responsive brain regions and the behavioral and endocrine response to increased stressor duration. This dissociation may have been due to a greater lag time for c-fos mRNA responses to reflect the impact of a dishabituation response. In conclusion, habituation of the endocrine and behavioral stress response occurred when the duration of the stressor matches the previous experience, while dishabituation of the stress response was triggered (with remarkable temporal precision) by an unexpected increase in stress duration.

Corticotropin-releasing factor and urocortin I activate CREB through functionally selective Gβγ signaling in hippocampal pyramidal neurons

Stress is a perceived perturbation in the environment of the organism that affects numerous extrahypothalamic brain regions including the hippocampus, a limbic structure critical for learning, spatial memory and the regulation of stress hormones. Though many effects of stress on the hippocampus are mediated via local glucocorticoid action, there is now ample evidence for the contributions of the stress peptides corticotropin-releasing factor (CRF) and urocortin I (UCN). Thus, understanding the intracellular signaling pathways activated by stress peptides is required to fully understand the mechanisms by which stress influences the hippocampus. Here we elucidate molecular mechanisms by which CRF and UCN induce phosphorylation of the activity-dependent transcription factor CREB, a molecule critical for numerous forms of neuronal plasticity. We report that nanomolar concentrations of both CRF and UCN lead to a rapid, CRF receptor 1 (CRFR1)- and Gβγ-dependent increase in CREB phosphorylation in rat hippocampal pyramidal neurons. Interestingly, CRF- and UCN-induced signaling pathways diverge downstream of Gβγ, with UCN, but not CRF, signaling to CREB via a MEK/MAPK-dependent pathway. These data suggest novel molecular mechanisms by which stress can directly impact hippocampal neurons, as well as highlight an emerging role for Gβγ signaling in mediating the effects of stress peptides in extrahypothalamic stress-responsive brain regions.

Sex differences in synaptic plasticity in stress-responsive brain regions following chronic variable stress

Increased stress responsiveness is implicated in the etiology of mood and anxiety disorders, including depression and post-traumatic stress disorder. Additionally, stress-related affective disorders have a higher incidence in women than men. Chronic stress in rodents produces numerous neuromorphological changes in a variety of limbic brain regions. Here, we examined the sex-dependent differences in presynaptic innervation of the paraventricular nucleus of the hypothalamus (PVN), prefrontal cortex (PFC), bed nucleus of the stria terminalis (BST), and amygdala in response to chronic variable stress (CVS). Following 14 days of CVS, the presynaptic protein synaptophysin was assessed in male and female rats. Our results demonstrate that synaptophysin staining density was higher in females than males in all brain areas evaluated, indicating sex differences in the organization of presynaptic innervation. After CVS, the PVN, principal nucleus of the BST (BSTpr), and basolateral nucleus of the amygdala (BLA) displayed significantly reduced synaptophysin density in females but not males. Furthermore, males showed an increase in synaptophysin in the PVN after CVS, suggesting a sex difference in the modulation of presynaptic inputs to the PVN following chronic stress. Overall, these data suggest marked sex differences in PVN, BSTpr, and BLA presynaptic innervation as a consequence of chronic stress, which may be associated with differential stress responsivity and perhaps susceptibility to pathologies in males and females.

Anxiety‐related behavioral inhibition in rats: a model to examine mechanisms underlying the risk to develop stress‐related psychopathology

Behavioral inhibition (BI) is an adaptive defensive response to threat; however, children who display extreme BI as a stable trait are at risk for development of anxiety disorders and depression. The present study validates a rodent model of BI based on an ethologically relevant predator exposure paradigm. We show that individual differences in rat BI are stable and trait-like from adolescence into adulthood. Using in situ hybridization to quantify expression of the immediate early genes homer1a and fos as measures of neuronal activation, we show that individual differences in BI are correlated with the activation of various stress-responsive brain regions that include the paraventricular nucleus of the hypothalamus and CA3 region of the hippocampus. Further supporting the concept that threat-induced BI in rodents reflects levels of anxiety, we also show that BI is decreased by administration of the anxiolytic, diazepam. Finally, we developed criteria for identifying extreme BI animals that are stable in their expression of high levels of BI and also show that high BI (HBI) individuals exhibit maladaptive appetitive responses following stress exposure. These findings support the use of predator threat as a stimulus and HBI rats as a model to study mechanisms underlying extreme and stable BI in humans.

Chronic stress alters the density and morphology of microglia in a subset of stress-responsive brain regions

The current study, in parallel experiments, evaluated the impact of chronic psychological stress on physiological and behavioural measures, and on the activation status of microglia in 15 stress-responsive brain regions. Rats were subjected, for 14 days, to two 30 min sessions of restraint per day, applied at random times each day. In one experiment the effects of stress on sucrose preference, weight gain, core body temperature, and struggling behaviour during restraint, were determined. In the second experiment we used immunohistochemistry to investigate stress-induced changes in ionized calcium-binding adaptor molecule-1 (Iba1), a marker constitutively expressed by microglia, and major histocompatibility complex-II (MHC-II), a marker often expressed on activated microglia, in a total of 15 stress-responsive nuclei. We also investigated cellular proliferation in these regions using Ki67 immunolabelling, to check for the possibility of microglial proliferation. Collectively, the results we obtained showed that chronic stress induced a significant increase in anhedonia, a decrease in weight gain across the entire observation period, a significant elevation in core body temperature during restraint, and a progressive decrease in struggling behaviour within and over sessions. With regard to microglial activation, chronic stress induced a significant increase in the density of Iba1 immunolabelling (nine of 15 regions) and the number of Iba1-positive cells (eight of 15 regions). Within the regions that exhibited an increased number of Iba1-positive cells after chronic stress, we found no evidence of a between group difference in the number of MHC-II or Ki67 positive cells. In summary, these results clearly demonstrate that chronic stress selectively increases the number of microglia in certain stress-sensitive brain regions, and also causes a marked transition of microglia from a ramified-resting state to a non-resting state. These findings are consistent with the view that microglial activation could play an important role in controlling and/or adapting to stress.