2.4 |. Two-bottle choice test for ethanol consumption and preference ratio

Twenty-two male and sixteen female adolescent rats were tested for ethanol consumption and preference ratio using the intermittent two-bottle choice with faded sucrose paradigm. Testing started on approximately PND 29–35, with age-at-onset similarly distributed between groups. Animals underwent a total of twelve 18-hr drinking sessions (1500–0900 hr the next day) that occurred three times a week (every other day with the weekend off) for 4 weeks. During each session, animals were single-housed in standard size cages and had ad libitum access to lab chow and two drinking bottles: one containing 5% (v/v) ethanol solution in water and the other containing water. The ethanol solution was sweetened with 1% sucrose during week 1 and 0.5% sucrose during week 2, while there was no sweetener during weeks 3 and 4. This faded sweetening paradigm was adapted from previous experiments (Fabio, Macchione, Nizhnikov, & Pautassi, 2015; Samson, 1986), because it effectively enhances ethanol acceptance in rodent ethanol consumption models (Carnicella, Ron, & Barak, 2014; Samson, 1986; Simms et al., 2008). Rats were weighed before every drinking session and drinking bottles were weighed before and after.

Ethanol consumed (g/kg) was calculated as follows: (ethanol solution consumed * 0.04) ÷ body weight. Ethanol preference-to-water ratio (%) was calculated as follow: (weight of ethanol solution consumed ÷ total (water and ethanol) solution consumed) * 100.