Supplementary Material for: PRECLINICAL EVALUATION OF SODIUM BUTYRATE’S POTENTIAL TO REDUCE ALCOHOL CONSUMPTION: A DOSE ESCALATION STUDY IN C57BL/6J MICE IN ANTIBIOTIC-ENHANCED BINGE-LIKE DRINKING MODEL

Introduction: In our earlier efforts to establish gut-brain axis during alcohol use disorder (AUD), we have demonstrated that supplementation of C57BL/6J male mice with 8 mg/ml sodium butyrate, a major short-chain fatty acid, in drinking water reduced ethanol intake and neuroinflammatory response in antibiotic (ABX)-enhanced voluntary binge-like alcohol consumption model, drinking in the dark (DID). Methods: To further evaluate the pre-clinical potential of SB, we have set a dose-escalation study in C57BL/6J male mice to test effects of ad libitum 20 mg/ml SB (SB20) and 50 mg/ml SB (SB50) and their combinations with ABX in the DID procedure for 4 weeks. Effects of these SB concentrations on ethanol consumption and bodily parameters were determined for the duration of the treatments. At the end of study, blood, liver and intestinal tissues were collected to study any potential adverse effects ad to measure blood ethanol concentrations. Results: Increasing SB concentrations in the drinking water caused a loss in the protective effect against ethanol consumption and produced adverse effects on body and liver weights, reduced overall liquid intake. The hypothesis that these effects were due to aversion to SB smell/taste at these high concentrations were further tested in a follow up proof-of-concept study with intragastric gavage administration of SB. The higher gavage dose (320 mg/kg) caused reduction in ethanol consumption without any adverse effects. Conclusion: Overall, these findings added more support for the therapeutic potential of SB in management of AUD, given a proper form of administration.