Rebecca M. Shansky, PhD - US grants

DESCRIPTION (provided by applicant): A majority of Americans will be exposed to a trauma in their lifetimes, but only 10% of those people will develop Post-Traumatic Stress Disorder (PTSD) as a result. Women, however, are twice as likely as men to develop PTSD after a trauma. Identification of the neurobiological factors that contribute to PTSD susceptibility and resilience in women is critical to developing more effective treatments, but basic science research into this problem is generally lacking. PTSD is characterized by a strong association between the trauma and its associated cues, a behavioral phenotype that may be the result of disrupted connectivity between the medial prefrontal cortex (mPFC) and amygdala. This pathway has been shown in animal models to mediate extinction from conditioned fear, and animals that fail to extinguish may be a good model of PTSD vulnerability. This proposal will explore neuroanatomical markers of vulnerability and resilience in male and female rats. Animals will be behaviorally characterized in classic cued fear conditioning and extinction protocols, identifying sex differences in variability in these behavioral tests. Using a combinatio of retrograde tracer and iontophoretic fluorescent microinjections, the dendritic morphology of mPFC neurons that project to the amygdala will be analyzed in animals that show high (vulnerable) and low (resilient) levels of freezing after extinction. These experiments will establish baseline sex differences in variability in fear behavior, which will be useful for interpreting future sex differences studies. Moreover, morphology analysis will provide a comprehensive neuroanatomical profile of vulnerability and resilience in both males and females, thus identifying areas that signify vulnerability uniquely in females. PUBLIC HEALTH RELEVANCE: Post-Traumatic Stress Disorder (PTSD) afflicts only 10% of people exposed to a trauma, but occurs twice as frequently in women as in men. Alterations in the structure and function of neurons that project from the medial prefrontal cortex (mPFC) to the amygdala may underlie the symptoms of PTSD, but sex differences in this pathway have not been thoroughly explored. This project will morphologically characterize neurons in the mPFC-amygdala pathway in male and female rats and correlate these measures with behavior in a model of PTSD, thus identifying sex-specific neuroanatomical markers of vulnerability and resilience.

Project summary Behavioral strategies to cope with potentially threatening or aversive stimuli tend to cluster into two forms: active, escape-driven behaviors, or passive responses that may conserve energy or prevent detection by a predator. Since an individual's coping strategy is closely linked to long-term clinical outcomes after trauma exposure, a better understanding of the neurobiological basis of coping response could greatly impact public health for both men and women. One promising but under-studied area is the role of the endocannabinoid (eCB) system in modulating active vs. passive stress responses. In this project, we will define the contribution of eCB signaling in the medial prefrontal cortex (mPFC) to the mechanisms that underlie coping responses in both sexes. In particular, we will investigate the ability of eCB system to modulate two mPFC-brainstem pathways: mPFC-dorsal raphe (DR) and mPFC-periaqueductal gray (PAG). We will use a combination of behavioral, neuroanatomical, biochemical, and chemogenetic strategies to dissect these circuits. The results of this work will lead to a better understanding of sex-specific stress processes, and have the potential to identify novel therapeutic targets for stress-related mental illnesses.

Summary Women are twice as likely as men to develop Post-Traumatic Stress Disorder (PTSD) after a trauma, but the neurobiological basis for this discrepancy is poorly understood. While there is a great deal of evidence that trauma itself can impact the male and female brain in discrete ways, less attention has been paid to the potential for PTSD treatments to work in a sex-dependent manner. A better understanding of the mechanisms that specifically mediate PTSD recovery in women could lead to improved therapeutics and a higher success rate for symptom reduction. In particular, the neural processes by which ovarian hormones modulate extinction learning represent a promising area of focus. Here, we will investigate the influence of circulating estrogen on the structure and function of neural circuitry connecting the ventral tegmental area (VTA), infralimbic area (IL) of the medial prefrontal cortex (mPFC), and basolateral area of the amygdala (BLA) in rats. We propose a system by which high estrogen states facilitate IL-BLA connectivity and enhanced extinction retention through upstream modulation of VTA-IL DA release during extinction learning. To test this hypothesis, we will use a combination of neuroanatomical tracers, intersectional viral techniques, and 3D reconstructions, thereby defining fear extinction-associated neural activity and plasticity across the estrous cycle. This work will result in a multi-synaptic map of extinction circuitry in the female brain, potentially identifying novel mechanisms by which estrogen can modulate learning processes.

Project Summary Sex differences in the development and prevalence of mental illnesses are widespread, suggesting that the factors that promote disease risk and resilience may be distinct in men and women. The neurobiological mechanisms underlying this possibility are poorly understood, however, because the vast majority of pre- clinical animal research has been conducted in males. In addition, the behavioral outcome measures used to understand learned fear have focused predominantly on passive, or freezing behavior. We recently identified an active, escape-like conditioned fear response in rats (?darting?) that occurred almost exclusively in females and was predictive of improved extinction retention. A better understanding of potentially sexually dimorphic mechanisms that drive this behavior could open new avenues for treatment of disorders that are more prevalent in women, like PTSD. We propose here to define the neural circuitry that drives the selection of conditioned darting vs. freezing, and how that integrates with known extinction circuits to promote extinction retention. Our preliminary data implicate the infralimbic cortex (IL) in this role, and we will use pharmacological, chemogenetic, and viral techniques to probe how descending IL projections to the dorsal and ventral periaqueductal gray (PAG) may mediate a strategic switch between active and passive responding and drive long-term behavioral flexibility through putative collaterals in the amygdala. We will then use sophisticated machine vision and machine learning-based tools to define the broader behavioral repertoires within which darting exists, thereby identifying a multifaceted phenotype that could provide insight into the mechanisms underlying individual differences in stress resilience and vulnerability.

Summary Women are twice as likely as men to develop Post-Traumatic Stress Disorder (PTSD) after a trauma, but the neurobiological basis for this discrepancy is poorly understood. One hallmark symptom of PTSD is a re- experiencing of the trauma in safe situations, a ?generalization? phenomenon that can be studied using preclinical rodent models like Pavlovian fear conditioning. Because both women and female rodents exhibit a greater tendency to generalize fear in safe contexts, better insight into potential sex-dependent factors that disrupt aversive context processing could lead to the development of novel treatments for PTSD. Preliminary data from our lab points to a novel role for the endocannabinoid system in conferring a sex-specific susceptibility to contextual fear generalization. Specifically, we observe a TRPV1-mediated increase in contextual fear generalization in females, but not males. A deeper investigation into this provocative finding may open new avenues for therapeutic development for women with PTSD. But in order to identify key areas and mechanisms to target for manipulation, we must first conduct exploratory studies to help guide the direction of larger-scale interrogations. The work we propose here will first determine whether our behavioral effects are selectively mediated by either the dorsal or ventral hippocampus (Aim 1), and then examine potential sex differences in mechanisms of eCB- and fear conditioning-related synaptic plasticity (Aim 2). We will carry out these Aims using a combination of behavioral pharmacology, fluorescent microscopy, biochemistry, and high resolution neuronal structural analysis. Together, these experiments will identify potential mediators of our behavioral effects, opening up our model for more focused interrogation and providing insight into sex-specific mechanisms of fundamental learning and memory processes.