Acute Ethanol Exposure Models and Gut Health

Two models of acute ethanol exposure were used to model a controlled acute dose. (1) A single oral gavage of ethanol (5 gm/kg body weight) or maltose was provided to the mice. The dosage was chosen based on a previously established acute ethanol exposure model which has been published by us previously, and this model has also been utilized as an established model by the National Institute on Alcohol Abuse and Alcoholism (NIAAA) to model for acute binge drinking [37 (link)]. This model results in systemic effect of ethanol after 6 h of ingestion. Mice were randomized to receive normal saline (100 µL; control) or tributyrin (100 µL; 2.5 mM) three days prior to the ethanol gavage that contained normal saline or tributyrin (5 mM) as part of the ethanol gavage per previous supplement allocation. Mice were euthanized 6 h after the acute ethanol and maltose challenge. Control mice received saline gavage to control the element of stress-related microbiome changes. (2) Mice received clindamycin subcutaneously (1.4 mg/kg body weight) for 3 days to induce disruption in gut microbiota. On day 4, a single oral gavage of ethanol (5 gm/kg body weight) or maltose was provided to mice. Mice were randomized to receive normal saline (100 µL) or tributyrin (100 µL; 2.5 mM) concurrently with the clindamycin treatment. Tributyrin (5 mM) or normal saline was provided in the oral ethanol gavage per previous supplement allocation. Mice were euthanized 6 h after the acute ethanol challenge. Mice were anesthetized and blood was collected from the posterior vena cava and plasma was separated and stored at −80 °C. Intestinal sections (jejunum, cecum) were dissected, and the cecum and jejunum were snap frozen in liquid nitrogen, and the jejunum was also stored for RNA extraction (RNALater, Ambion, Austin, TX, USA) or frozen for tissue sectioning and histological analysis (optimal cutting temperature compound (OCT), Sakura Finetek, Torrence, CA, USA). Fecal pellets were expelled from the proximal colon and immediately plated for bacterial growth.