AbstractA major limitation of the most widely used current animal models of alcohol dependence is that they use forced exposure to ethanol including ethanol-containing liquid diet and chronic intermittent ethanol (CIE) vapor to produce clinically relevant blood alcohol levels (BAL) and addiction-like behaviors. We recently developed a novel animal model of voluntary induction of alcohol dependence using ethanol vapor self-administration (EVSA). In this model, naive outbred rats given intermittent access to alcohol vapor self-administration exhibit BAL in the 150-300 mg% range and develop somatic signs of withdrawal during acute abstinence. However, it is unknown whether EVSA leads to an escalation of alcohol drinkingper se, and whether such escalation is associated with neuroadaptations in brain regions related to stress, reward, and habit. To address these issues, we compared the levels of alcohol drinking during withdrawal between rats passively exposed to alcohol (CIE) or voluntarily exposed to EVSA and measured the number of Fos+ neurons during acute withdrawal (16 h) in the central nucleus of the amygdala (CeA), dorsomedial striatum (DMS), dorsolateral striatum (DLS), nucleus accumbens core (Nacc), periaqueducal grey area (PAG), lateral Habenula (HbL), and the paraventricular nucleus of the thalamus (PVT). The rats were first trained to orally self-administer alcohol in standard operant chambers and then divided in 4 groups (CIE, CI-Air, EVSA and Air-SA) and exposed to intermittent ethanol vapor (passive or active) or intermittent air (passive or active) for 8 h/day, 3 days a week. CIE and EVSA rats exhibited similar BAL (150-300 mg% range) and similar escalation of alcohol drinking during withdrawal, while no changes in terms of drinking were observed in the air exposed rats. CIE and EVSA also increased the motivation for alcohol compared to their respective air control groups under a progressive ratio schedule of reinforcement. Acute withdrawal from EVSA and CIE recruited a similar number of Fos+ neurons in the CeA, however, acute withdrawal from EVSA recruited a higher number of Fos+ neurons in every other brain region analyzed compared to acute withdrawal from CIE. Moreover, acute withdrawal from EVSA specifically recruited the DMS and PVT, a pattern not observed in CIE rats.In summary, these results demonstrate that EVSA produces similar escalation of alcohol drinking, motivation to drink, and blood-alcohol levels than the CIE model, while letting animals voluntary initiate alcohol exposure and maintain alcohol dependence. Moreover, while the behavioral measures of alcohol dependence between the voluntary (EVSA) and passive (CIE) model was similar, the recruitment of neuronal ensembles during acute withdrawal was very different with a higher recruitment of Fos+ neurons in key brain regions important for stress, reward and habit-related processes. The EVSA model may be particularly useful to unveil the neuronal networks and pharmacology responsible for the voluntary induction and maintenance of alcohol dependence and may improve translational studies by providing preclinical researchers with an animal model with better face validity for alcohol use disorder.