Acute withdrawal signatures are associated with delayed affective-like vulnerability after chronic intermittent ethanol exposure in rats

Alcohol use disorder is associated with persistent depression and anxiety, but vulnerability after withdrawal is heterogeneous. We examined whether acute withdrawal measures were associated with delayed affective-like and activity-related outcomes after chronic intermittent ethanol exposure in rats. Male and female Sprague-Dawley rats received five cycles of 10% v/v ethanol liquid diet or pair-fed control diet.

Acute withdrawal signs were assessed 12 to 24 h after the final ethanol cycle, and behavior was measured one month later using sucrose preference, open field (OF), and elevated plus maze (EPM) testing. Alcohol-exposed rats showed reduced sucrose preference, EPM open-arm exploration, OF total distance, and OF center-zone distance. Depending on the measure, 28.3% to 56.7% of alcohol-exposed rats met control-derived criteria for delayed behavioral abnormalities.

Principal component analysis of five neurological withdrawal signs yielded a single Neurological Severity Score (NSS), explaining 50.4% of variance. Among the three prespecified withdrawal measures, body temperature showed the most consistent associations with multiple delayed outcomes, including depression-like behavior, hypoactivity, sensitive anxiety, and sensitive comorbidity. Temperature-only models showed moderate internal discrimination, with optimism-corrected AUCs of 0.773 for sensitive anxiety and 0.735 for hypoactivity.

NSS and weight change did not show consistent independent associations. These findings identify body temperature as the most informative candidate withdrawal measure among those assessed in this cohort. Independent replication is required, and the association does not establish causality or clinical predictive value.

We sincerely thank Anna Oleshko from the Department of Biology and Methods of Teaching Biology, A. Makarenko Sumy State Pedagogical University, Sumy, Ukraine, and Olha Shapoval from the Department of Biochemistry and Physiology, Faculty of Biology and Ecology, Oles Honchar Dnipro National University, Dnipro, Ukraine, for their valuable guidance and assistance with behavioral testing and evaluation. This research received no specific grant from any funding agency in the public, commercial, or not-for-profit sectors.

These authors contributed equally: Alexander Zlotnik, Benjamin F. These authors jointly supervised this work: Amit Frenkel, Matthew Boyko. Department of Anesthesiology and Critical CareSoroka University Medical Center, Ben-Gurion University of the Negev, Beer-Sheva, 84101, Israel Alexander Zlotnik, Shahar Negev, Michael Dubilet, Yakov Perlov, Nitzan Hertz, Amit Frenkel & Matthew Boyko Department of Anesthesiology and Perioperative Medicine, Mayo Clinic, Jacksonville, FL, 32224, USA Nursing Research Unit, Soroka University Medical Center Ben-Gurion of the Negev, Beer-Sheva, 84101, Israel Department of Orthopedic Surgery, Soroka University Medical Center, Ben-Gurion University of the Negev, Beer-Sheva, 84101, Israel The authors declare no competing interests.