Structural and metabolic activity differences in serotonergic cell groups in a rat model of individual differences of emotionality and stress reactivity.

Serotonin regulates a diverse set of functions, including emotional behavior, cognition, sociability, appetite, and sleep. Serotonin is also a key trophic factor that shapes neurodevelopmental processes. Genetic and environmental factors that drive individual differences in the serotonergic system have the capacity to impact brain structure and behavior, and likely contribute to pathophysiological processes involved in neuropsychiatric disorders. Using adult rats selectively bred for low novelty exploration (Low Responders, LR), we previously demonstrated pronounced increases in the levels of their anxiety- and depression- relevant behaviors as compared to the selectively bred High Novelty Responder (HR) rats. These behavioral differences were accompanied by alterations in the expression of genes that regulate serotonin synthesis in the brainstem, and its signaling in the forebrain. The present study extends these observations with a focus on the organization and the metabolism of brainstem serotonin cell groups that provide serotonergic innervation of the hippocampus and other limbic regions of male HR/LR rats. Using design-based stereology, we found the median raphe (MnR) in adult male LR rats contains increased number of serotonergic neurons as compared to the HRs. This is preceded by an increase in the metabolic activity of the caudal dorsal raphe (DRC) and the intrafascicular DR (DRI) during early postnatal development. These findings suggest that structural and functional differences in the raphe-limbic projections shape behavioral inhibition throughout the lifespan.

Disrupted serotonin system development via early life antidepressant exposure impairs maternal care and increases serotonin receptor expression in adult female offspring.

Manipulating serotonin (5-HT) levels in the developing brain elicits a range of effects on brain function and behavior. For example, early-life exposure to selective 5-HT reuptake inhibitor (SSRI) antidepressants disrupts dorsal raphe function and triggers aberrant adult behaviors such as increased passive stress coping and anhedonia. However, much less is understood about how alterations in 5-HT signaling in early life impact outcomes in female offspring, including critical social functions such as maternal care. The present study shows that early-life SSRI exposure disrupts adult female offspring's maternal behavior. Pregnant/postpartum female Sprague-Dawley rats were treated with the SSRI citalopram in drinking water or provided regular tap water as control. Female offspring were raised to adulthood and mated with treatment-naïve males. Following parturition, we observed maternal behavior during portions of the light and dark phases of postnatal days (P)1-14. Relative to controls, dams with a history of early-life SSRI exposure exhibited decreased maternal care, including diminished arched-back nursing, reduced licking and grooming of pups, and increased behavioral inconsistency. Brains were collected from dams with and without a history of early-life SSRI exposure to measure relative mRNA expression of select 5-HT receptor transcripts (5HTR1A, -1B, -2A, -2C) in regions involved in maternal care. Early-life SSRI exposure augmented expression of 5-HTR1A in the medial preoptic area and 5-HTR1B, 5-HTR2A, and 5-HTR2C in the prefrontal cortex. These results demonstrate that early alterations to 5-HT signaling through SSRI exposure may disrupt nurturing parental behaviors and 5-HT receptor expression in affected female rat offspring.

Resilience to Stress: Lessons from Rodents about Nature versus Nurture.

Individual differences in human temperament influence how we respond to stress and can confer vulnerability (or resilience) to emotional disorders. For example, high levels of behavioral inhibition in children predict increased risk of mood and anxiety disorders in later life. The biological underpinnings of temperament are unknown, although improved understanding can offer insight into the pathogenesis of emotional disorders. Our laboratory has used a rat model of temperamental differences to study neurodevelopmental factors that lead to a highly inhibited, stress vulnerable phenotype. Selective breeding for high versus low behavioral response to novelty created two rat strains that exhibit dramatic behavior differences over multiple domains relevant to emotional disorders. Low novelty responder (bLR) rats exhibit high levels of behavioral inhibition, passive stress coping, anhedonia, decreased sociability and vulnerability to chronic stress compared to high novelty responders (bHRs). On the other hand, bHRs exhibit high levels of behavioral dis-inhibition, active coping, and aggression. This review article summarizes our work with the bHR/bLR model showing the developmental emergence of the bHR/bLR phenotypes, the role the environment plays in shaping it, and the involvement of epigenetic processes such as DNA methylation that mediate differences in emotionality and stress reactivity.

Modeling heritability of temperamental differences, stress reactivity, and risk for anxiety and depression: Relevance to research domain criteria (RDoC).

Animal models provide important tools to study biological and environmental factors that shape brain function and behavior. These models can be effectively leveraged by drawing on concepts from the National Institute of Mental Health Research Domain Criteria (RDoC) Initiative, which aims to delineate molecular pathways and neural circuits that underpin behavioral anomalies that transcend psychiatric conditions. To study factors that contribute to individual differences in emotionality and stress reactivity, our laboratory utilized Sprague-Dawley rats that were selectively bred for differences in novelty exploration. Selective breeding for low versus high locomotor response to novelty produced rat lines that differ in behavioral domains relevant to anxiety and depression, particularly the RDoC Negative Valence domains, including acute threat, potential threat, and loss. Bred Low Novelty Responder (LR) rats, relative to their High Responder (HR) counterparts, display high levels of behavioral inhibition, conditioned and unconditioned fear, avoidance, passive stress coping, anhedonia, and psychomotor retardation. The HR/LR traits are heritable, emerge in the first weeks of life, and appear to be driven by alterations in the developing amygdala and hippocampus. Epigenomic and transcriptomic profiling in the developing and adult HR/LR brain suggest that DNA methylation and microRNAs, as well as differences in monoaminergic transmission (dopamine and serotonin in particular), contribute to their distinct behavioral phenotypes. This work exemplifies ways that animal models such as the HR/LR rats can be effectively used to study neural and molecular factors driving emotional behavior, which may pave the way toward improved understanding the neurobiological mechanisms involved in emotional disorders.

Perinatal SSRI Exposure Disrupts G Protein-coupled Receptor BAI3 in Developing Dentate Gyrus and Adult Emotional Behavior: Relevance to Psychiatric Disorders.

Selective serotonin reuptake inhibitor (SSRI) antidepressants are widely prescribed to pregnant women suffering with depression, although the long-term impact of these medications on exposed offspring are poorly understood. Perinatal SSRI exposure alters human offspring's neurodevelopment and increases risk for psychiatric illness in later life. Rodent studies suggest that perinatal SSRI-induced behavioral abnormalities are driven by changes in the serotonin system as well as epigenetic and transcriptomic changes in the developing hippocampus. A major gene altered by perinatal SSRI exposure is the G-protein coupled receptor Brain Angiogenesis Inhibitor 3 (BAI3). Our present study shows that perinatal exposure to the SSRI citalopram increases mRNA expression of Bai3 and related molecules (including its C1ql ligands) in the early postnatal dentate gyrus of male and female offspring. Transient Bai3 mRNA knockdown in perinatal SSRI-exposed dentate gyrus lessened behavioral consequences of perinatal SSRI exposure, leading to increased active stress coping. To determine translational implications of this work, we examined expression of BAI3 and related molecules in hippocampus and prefrontal cortex from patients that suffered with depression or schizophrenia relative to healthy control subjects. We found sex- and region-specific changes in mRNA expression of BAI3 and its ligands C1QL2 and C1QL3 in men and women with a history of psychiatric disorders compared to healthy controls. Together these results suggest that abnormal BAI3 signaling may contribute to molecular mechanisms that drive adverse effects of perinatal SSRI exposure, and show evidence for alterations of BAI3 signaling in the hippocampus of patients that suffer depression and schizophrenia.

Inborn differences in emotional behavior coincide with alterations in hypothalamic paraventricular motor projections.

Integrated behavioral responses to emotionally salient stimuli require the concomitant activation of descending neural circuits that integrate physiological, affective, and motor responses to stress. Our previous work interrogated descending circuits in the brainstem and spinal cord that project to motor and sympathetic targets. The hypothalamic paraventricular nucleus (PVN), a key node of this circuitry, integrates multiple motor and sympathetic responses activated by stress. The present study sought to determine whether descending projections from the PVN to targets in muscle and adrenal gland are differentially organized in rats with inborn differences in emotionality and stress responsivity. We utilized retrograde transsynaptic tract-tracing with unique pseudorabies virus (PRV) recombinants that were injected into sympathectomized gastrocnemius muscle and adrenal gland in two rat models featuring inborn differences in emotional behavior. Our tract-tracing results revealed a significant decrease in the number of PVN neurons with poly-synaptic projections to the gastrocnemius in male Wistar Kyoto [WKY] rats (versus Sprague Dawley rats) and selectively bred Low Novelty Responder [bLR] rats (versus selectively bred High Novelty Responder [bHR] rats). These neuroanatomical differences mirrored behavioral observations showing that both WKY and bLR rats display marked inhibition of emotional motor responses in a variety of settings relative to their respective controls. Our findings suggest that, in male rodents, PVN poly-synaptic projections to skeletal muscle may regulate emotional motor and coping responses to stress. More broadly, perturbations in PVN motor circuitry may play a role in mediating psychomotor disturbances observed in depression or anxiety-related disorders.

Rats bred for high anxiety exhibit distinct fear-related coping behavior, hippocampal physiology, and synaptic plasticity-related gene expression.

The hippocampus is essential for learning and memory but also regulates emotional behavior. We previously identified the hippocampus as a major brain region that differs in rats bred for emotionality differences. Rats bred for low novelty response (LRs) exhibit high levels of anxiety- and depression-like behavior compared to high novelty responder (HR) rats. Manipulating the hippocampus of high-anxiety LR rats improves their behavior, although no work to date has examined possible HR/LR differences in hippocampal synaptic physiology. Thus, the current study examined hippocampal slice electrophysiology, dendritic spine density, and transcriptome profiling in HR/LR hippocampus, and compared performance on three hippocampus-dependent tasks: The Morris water maze, contextual fear conditioning, and active avoidance. Our physiology experiments revealed increased long-term potentiation (LTP) at CA3-CA1 synapses in HR versus LR hippocampus, and Golgi analysis found an increased number of dendritic spines in basal layer of CA1 pyramidal cells in HR versus LR rats. Transcriptome data revealed glutamate neurotransmission as the top functional pathway differing in the HR/LR hippocampus. Our behavioral experiments showed that HR/LR rats exhibit similar learning and memory capability in the Morris water maze, although the groups differed in fear-related tasks. LR rats displayed greater freezing behavior in the fear-conditioning task, and HR/LR rats adopted distinct behavioral strategies in the active avoidance task. In the active avoidance task, HRs avoided footshock stress by pressing a lever when presented with a warning cue; LR rats, on the other hand, waited until footshocks began before pressing the lever to stop them. Taken together, these findings concur with prior observations of HR rats generally exhibiting active stress coping behavior while LRs exhibit reactive coping. Overall, our current findings coupled with previous work suggest that HR/LR differences in stress reactivity and stress coping may derive, at least in part, from differences in the developing and adult hippocampus.

Perinatal exposure to the SSRI paroxetine alters the methylome landscape of the developing dentate gyrus.

Evidence in humans and rodents suggests that perinatal exposure to selective serotonin reuptake inhibitor (SSRI) antidepressants can have serious long-term consequences in offspring exposed in utero or infancy via breast milk. In spite of this, there is limited knowledge of how perinatal SSRI exposure impacts brain development and adult behaviour. Children exposed to SSRIs in utero exhibit increased internalizing behaviour and abnormal social behaviour between the ages of 3 and 6, and increased risk of depression in adolescence; however, the neurobiological changes underlying this behaviour are poorly understood. In rodents, perinatal SSRI exposure perturbs hippocampal gene expression and alters adult emotional behaviour (including increased depression-like behaviour). The present study demonstrates that perinatal exposure to the SSRI paroxetine leads to DNA hypomethylation and reduces DNA methyltransferase 3a (Dnmt3a) mRNA expression in the hippocampus during the second and third weeks of life. Next-generation sequencing identified numerous differentially methylated genomic regions, including altered methylation and transcription of several dendritogenesis-related genes. We then tested the hypothesis that transiently decreasing Dnmt3a expression in the early postnatal hippocampus would mimic the behavioural effects of perinatal SSRI exposure. We found that siRNA-mediated knockdown of Dnmt3a in the dentate gyrus during the second to third week of life produced greater depression-like behaviour in adult female (but not male) offspring, akin to the behavioural consequences of perinatal SSRI exposure. Overall, these data suggest that perinatal SSRI exposure may increase depression-like behaviours, at least in part, through reduced Dnmt3a expression in the developing hippocampus.