Abstract. The neural substrates underlying alcohol use disorder (AUD), remain poorly understood in part due to lack of translational models that recapitulate phenotypes from the human condition. Biosynthesis of the GABAergic neurosteroid, allopregnanolone (Allo) in corticolimbic neurons, regulates stress sensitivity and induces a potent anxiolytic action. In a rodent model of chronic intermittent ethanol (CIE) exposure, decreased expression of Allo biosynthetic enzymes, 5α-reductase type I (5α-RI) and 3α-hydroxy-steroid dehydrogenase (3α-HSD) is associated with Allo level downregulation in the hippocampus (HIP) and cerebellum. Consistently, in AUD postmortem brain, cerebellum Allo levels and neurosteroidogenic proteins and enzymes, such as the translocator protein (TSPO), 5α-RI and 3α-HSD expression decreased in association with aberrant epigenetic marks. Alcohol-induced epigenetic modifications (e.g., DNA hyper/hypomethylation) on transcriptomics and their impact on neurosteroidogenic gene expression, neurosteroid levels, and anxiety are poorly understood. Allo biosynthesis can be upregulated in brain areas that modulate anxiety and alcohol reward by stimulating the epigenetically modifiable nuclear receptor, peroxisome proliferator-regulated receptor (PPAR)-α by the endogenous modulator, palmitoylethanolamide (PEA). Intriguingly, chronic alcohol exposure decreases PPAR- α expression, while stimulation of PPAR-α by PEA decreases both anxiety and alcohol intake. The molecular mechanisms underlying these effects remain unclear. Our preliminary and published results suggest that alcohol-induced aberrant regulation of PPAR-α may affect anxiety via decreasing Allo content. Hypothesis: Chronic alcohol exposure alters methylation/demethylation dynamics that downregulate corticolimbic PPAR-α expression and allopregnanolone biosynthesis, and elevated anxiety. In male and female rats, we will: (AIM 1) Examine the effect of 14-day CIE exposure (EtOH), ethanol acute (24h, W24h) and protracted (7 days, W7d) withdrawal on the epigenetic regulation of PPAR-α expression and downstream effects on neurosteroidogenic enzyme expression; (AIM 2) Investigate the effects of CIE exposure and ethanol acute and protracted withdrawal on the brain content of Allo; and (AIM 3) Study the pharmacoepigenetics of PEA and fenofibrate on Allo biosynthesis and anxiety after CIE exposure and acute and protracted ethanol withdrawal. This study may unveil CIE-induced neurobiological alterations and suggest treatment targets for alcohol withdrawal symptoms.

AUD is a debilitating neuropsychiatric disorder and the third-leading preventable cause of death in the US, yet remains without efficient treatments. The GABAergic neurosteroid, allopregnanolone (Allo) is decreased in AUD postmortem brain and in rat models of alcohol dependence. PPAR-α is an epigenetically modifiable neuronal target that regulates Allo biosynthesis and anxiety, reduces alcohol drinking, and may thus provide a pharmacological target for symptoms of alcohol withdrawal.