Project Summary/Abstract: Adolescent alcohol use is a known risk factor for the development of alcohol use disorders (AUDs); latent maladaptive plasticity associated with adolescent use likely underlies this increased risk in adults. The bed nucleus of the stria terminalis (BNST) is a sexually dimorphic brain region and critically involved in stress and negative affect-associated relapse in AUDs, and therefore is a prime target for this maladaptive plasticity. Using a model of adolescent intermittent ethanol exposure (AIE) in male and female mice, we find disrupted BNST glutamatergic transmission and plasticity, but the mechanisms differ across sex (GluN2B- NMDAR-mediated in males and mGluR1/5-mediated in females). When tested during adulthood, both forms of BNST plasticity return to control levels but exposure to restraint stress produced a re-emergence of AIE-induced that were sex-specific. AIE also increased glutamate release in the BNST and this effect appears be mitigated through modulation of presynaptic CRFR1. AIE and adult stress also produce sex-specific negative affect behavioral phenotypes (novelty-induced hypophagia test in females, foot shock-induce freezing in males). As the BNST sits at the intersection between cortical inputs and downstream amygdalar and hypothalamic targets, this enhanced drive to the BNST and subsequent inter BNST plasticity likely sensitize the stress response and the expression of negative affect behaviors. Using a combination of molecular, electrophysiological, genetic, and behavioral approaches, we will evaluate the ability of AIE to prime the BNST to adult stress sex-specific manner. I hypothesize that that AIE and adult stress act to synergistically enhance glutamate release in the BNST via activation of CRFR1 in male mice but not female mice (Aim 1). Modulation of CRFR1 transmission will be examined for its ability to block the effects of stress on glutamate release and plasticity and evaluate stress sensitive inputs. Next, I propose that the sex difference in BNST plasticity occur in distinct projection regions (CeA, LH, or VTA). Finally, BNST-specific deletion, BNST DREADD activation, and pharmacological antagonism of GluN2B-NMDARs and mGluR5s (depending on the sex) will be used to assess the role of these receptors in the development of sex-specific negative affect phenotypes. The goal of this work is to identify potential pharmacotherapies to treat AUDs and our work suggest that these likely need to be distinct for males and females.