Abstract A central problem in the treatment of ethanol (EtOH) addiction is the prevalence of relapse to EtOH use even after protracted intervals of forced or self-imposed abstinence. Advances have been made in elucidating the neurocircuitry that mediates craving and EtOH seeking, which provides insights into the neurobiological basis of relapse. Functional brain imaging in humans and studies that use c-fos expression as a marker of neural activation in rodents implicate interconnected cortical and limbic brain regions in response to drug cue-, drug priming-, and stress-induced reinstatement. Major components of this circuitry include the medial prefrontal cortex (mPFC), basolateral amygdala (BLA), central nucleus of the amygdala (CeA), bed nucleus of the stria terminalis (BNST), ventral tegmental area (VTA), nucleus accumbens (NAC), hippocampus, thalamus (THAL), and dorsal striatum. The hypocretin (Hcrt) system regulates a wide range of physiological processes, including feeding, energy metabolism, arousal, and stress, and is recruited by drugs of abuse. Of interest for the present proposal, recent studies have demonstrated a critical contribution of Hcrt in the modulation of stress and a possible anxiolytic effect of Hcrt receptor (Hcrt-r) antagonists. Furthermore, we have collected convincing data that EtOH exposure recruits the Hcrt system. Specifically, we found that downregulation of Hcrt mRNA was observed in the lateral hypothalamus (the major source of Hcrt production) of animals that had a history of EtOH dependence and that blockade of Hcrt-r selectively reversed EtOH seeking vs. natural reward seeking. Chronic drug use is well known to dysregulate stress responses that are mediated by corticotropin-releasing factor (CRF) in both the hypothalamic-pituitary-adrenal (HPA) axis and extrahypothalamic brain stress areas outside the HPA axis (e.g., CeA and BNST). With repeated cycles of drug use, the HPA axis becomes hyporesponsive, accompanied by an increase in the extrahypothalamic CRF stress system response (i.e., CRF-CRF1 receptors). Importantly, a Hcrt/CRF interaction exists, and it has been proposed that Hcrt modulation of CRF neurons participates in the chronic relapsing, negative affective states that characterize drug addiction. Converging lines of evidence from human and animal studies suggest that impairment of mPFC function due to drugs of abuse exposure is a key factor in the transition from goal-directed to compulsive drug seeking. The mPFC contains CRF interneurons, and Hcrt neurons project to the mPFC. This proposal will test the hypothesis that a history of EtOH dependence dysregulates Hcrt and its interaction with CRF in the mPFC, particularly the infralimbic area (IL), and if this dysfunction ...