C-FOS MAPPING OF NEURAL CIRCUITS ENGAGED BY ENHANCED BINGE-LIKE ETHANOL DRINKING IN C57BL/6J MICE

Alcohol consumption is a leading cause of preventable death worldwide, with binge drinking being a highly prevalent harmful pattern. This study evaluated a modified four-bottle Drinking in the Dark (DID) preclinical paradigm to enhance binge-like ethanol intake and induce neural activation assessed by c-Fos expression. Male C57BL/6J mice underwent a 4-day DID protocol with 2 h of ethanol access daily, except on day 4 when access was extended to 4 h. In Experiment 1, ethanol consumption (5%, 10%, 20%, or 40%) was measured using 1–4 bottles and compared with water intake in young (8–9 weeks) and adult (20 weeks) mice. Experiment 2 focused on young mice, as young adulthood shows the highest binge consumption and heightened neuroplasticity, and examined neuronal activation following the 4-day DID procedure with four bottles of 20% ethanol or water; brains were collected after 2 h or 4 h of access on test day (day 4). Ethanol intake reached ~5 g/kg within 2 h and 8–9 g/kg over 4 h. c-Fos significantly increased in the central amygdala, nucleus accumbens, and arcuate nucleus at both 2 and 4 h, with no differences between time points. These patterns indicate selective engagement of reward- and homeostasis-related circuits, with the nucleus accumbens and arcuate nucleus potentially reflecting endogenous opioid system involvement. Overall, the modified multi-bottle DID model robustly increases ethanol intake and induces neural activation patterns consistent with binge-like and excessive drinking, providing a relevant preclinical model of high-risk alcohol consumption. Funding: UJIB2021/28.