On postnatal week 10, the MS rats were single housed and randomly assigned to water drinking (MS15W, n = 10; MS360W, n = 10) or free-choice water/alcohol drinking (MS15E, n = 10; MS360E, n = 20) groups. To simulate human episodic drinking, a voluntary drinking paradigm with repeated drinking and non-drinking days in between was employed [29]. The rats exposed to ethanol had free choice between non-sweetened ethanol (5 or 20 % made from Ethanol 96 %; Solveco AB, Rosersberg, Sweden) and water for three consecutive days a week with drug-free days in-between. The first week the rats had free access to 5 % ethanol for 24 h/day, the next week they had limited access to 5 % for 2 h/day, and the following five weeks access to 20 % ethanol in 2 h sessions for three consecutive days a week. Water drinking controls had two bottles with water. After 5 weeks of access to 20 % ethanol and/or water, the rats were decapitated immediately after a 2-h drinking session. Trunk blood was collected, and the brain regions VTA, Acb, Amy, dStr, mPFC, and CCx were removed, immediately frozen, and stored at −80 °C.
