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ID 25994 Poster Board 467With over 50 million people affected in the US alone, mental health disorders take a high toll on our country. Further, women have a predisposition to developing stress-related disorders and demonstrate more anxiety-like behaviors following stress cues compared with men. Anxiety and fear responses are heavily regulated by the amygdala, a brain region that plays a crucial role in mood behaviors and emotional memory processes. Mu (MOR), kappa (KOR), and delta (DOR) opioid receptors are present in high concentrations in such stress-related areas of the brain and mediate communication with specific neurotransmitters involved in emotional processing and stress responses. Enkephalin (ENK), an agonist at both MOR and DOR, promotes a basal hedonic state and is known to attenuate stress-induced anxiety-like behavior. In contrast, the endogenous KOR agonist dynorphin (DYN) promotes an "anti-reward" response, through inhibition of dopamine release and initiation of a dysphoric cycle that leads to irritability, anxiety, anhedonia, and loss of reward motivation. Accordingly, KOR neurotransmission mediates the effects of stress and leads to heightened drug reinstatement in rodent models. This study utilized a highly validated, ethologically relevant model, witness stress (WS), whereby females observed social defeat between two males from a confined region of an aggressive resident's cage. Previous data from our lab suggests that WS increases anxiety-like responding in females as determined by increases in hypervigilant burying responses during stress exposure as well as during re-exposure to the stress context. Further, we determined that these behavioral responses were paralleled by increases in KOR expression within the amygdala, supporting the involvement of the opioid receptor systems in the stress susceptibility of females. We next aimed to determine if exposure to witness stress (WS)-related cues alters ENK and DYN consistent with maintenance of anxiety-like behaviors in two regions of the amygdala, the basolateral amygdala (BLA) and the central amygdala (CeA), in females with a history of WS. This is an especially important consideration considering research regarding females within this topic has not yet been explored. To test this hypothesis, we first exposed female rats to a single WS exposure and brain tissue was collected two hours later. In the second experiment, rats experienced five days of WS and were subsequently exposed to WS-related cues prior to tissue collection. Immunofluorescence was used to detect levels of ENK and DYN in the CeA and BLA in response to both acute stress and to the WS context. In alignment with previous literature and current rationale, we expect that ENK expression will be lower in the BLA of stressed females while, in turn, DYN expression should be increased in the CeA of these same rats to create a dysphoric effect following stress-cue exposure. In correspondence with these results, future studies will employ behavioral pharmacology to determine if injecting t...

ID 28676 Poster Board 412Neuropsychiatric disorders that result from stress, especially post-traumatic stress disorder, are highly detrimental to both the patient and society and disproportionally affect females. One hallmark symptom of this disorder, that is also more often observed in females, is hypervigilance in response to reminders of the initial stressful/traumatic event.. Recent evidence points towards heightened neuroimmune signaling in response to stress as a mechanistic factor underlying this behavioral dysfunction. Previous experiments in our lab have determined that social stress, specifically vicarious witness stress (WS), leads to increases in neuroimmune signaling within the locus coeruleus (LC) of females, as indicated by increased microglial activation and IL-1b proinflammatory cytokine release. Further, neuroimmune factors are known to activate LC neurons while hyperactivity of LC neurons has been identified as a maladaptive factor in clinical and preclinical studies of anxiety. Taken together, increased proinflammatory signaling may be a key factor in the increased susceptibility to developing neuropsychiatric disorders following stress exposure that is observed in females. Therefore, the goal of this project was to determine the functional and behavioral impact of blocking microglial activity within the LC during WS exposure on subsequent hypervigilance-related outcomes. These experiments utilized chemogenetic DREADDs techniques with a virus engineered to specifically target microglia (AAV-CD68-Gi). The first experiment was designed to determine the impact of intra-LC microglial inactivation on anxiety-like behavior during stress exposure. Female rats were injected with this inhibitory virus and, following 3-4 weeks to allow for viral expression, rats were injected with either CNO to activate the DREADDs virus or vehicle one hour prior to exposure to a single session of WS. Brain tissue was collected two hours following stress exposure and behavior during the session along with the activation state of the microglia was analyzed. A second study treated female rats with AAV-CD68-Gi within the LC prior to five daily WS exposures with CNO/vehicle injections before each session. Subsequently, rats underwent testing in marble burying and the stress context to determine the impact of microglial blockade on subsequent hypervigilant responses to stress-related stimuli. Finally, a separate subset of surgically naïve rats were solely treated with CNO or vehicle to ensure that CNO alone had no effect on behaviors during stress. Importantly, these studies quantified microglial morphology using Iba1 and confirmed that DREADD activation with CNO prevented microglial activation and further verified that CNO did not affect behaviors during stress. Additionally, these experiments concluded that DREADD-mediated microglial inhibition prevents the emergence of the hypervigilant burying behaviors we have previously observed both during stress and in response to the stress context. Finally, cfos immunohistochemistr...

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