Three Weeks of Binge Alcohol Drinking Generates Increased Alcohol Front-Loading and Robust Compulsive-Like Alcohol Drinking in Male and Female C57BL/6J Mice

Abstract

Background: Current models of compulsive-like quinine-adulterated alcohol (QuA) drinking in mice, if improved, could be more useful for uncovering the neural mechanisms of compulsive-like alcohol drinking. The purpose of these experiments was to further characterize and improve the validity of a model of compulsive-like QuA drinking in C57BL/6J mice. We sought to determine whether compulsive-like alcohol drinking could be achieved following 2 or 3 weeks of Drinking-in-the-Dark (DID), whether it provides evidence for a robust model of compulsive-like alcohol drinking by inclusion of a water control group and use of a highly concentrated QuA solution, whether repeated QuA exposures alter compulsive-like drinking, and whether there are sex differences in compulsive-like alcohol drinking.

Methods: Male and Female C57BL/6J mice were allowed free access to either 20% alcohol or tap water for 2 hours each day for approximately 3 weeks. After 2 or 3 weeks, the mice were given QuA (500 μM) and the effect of repeated QuA drinking sessions on compulsive-like alcohol drinking was assessed. 3-minute front-loading, 2 hour binge-drinking, and blood alcohol concentrations were determined.

Results: Compulsive-like QuA drinking was achieved after 3 weeks, but not 2 weeks, of daily alcohol access as determined by alcohol history mice consuming significantly more QuA than water history mice and drinking statistically nondifferent amounts of QuA than nonadulterated alcohol at baseline. Thirty-minute front-loading of QuA revealed that alcohol history mice front-loaded significantly more QuA than water history mice, but still found the QuA solution aversive. Repeated QuA exposures did not alter these patterns, compulsive-like drinking did not differ by sex, and BACs for QuA drinking were at the level of a binge.

Conclusions: These data suggest that compulsive-like QuA drinking can be robustly achieved following 3 weeks of DID and male and female C57BL/6J mice do not differ in compulsive-like alcohol drinking.

Keywords: Alcohol; Aversion-Resistant; Binge; Compulsion; Quinine.

Fig. 1.

Absence of compulsive-like alcohol drinking phenotype after 2 weeks of DID. Male and female C57BL/6J mice consumed either alcohol or water for 14 days. On day 15, all mice were given QuA (QuA Test 1; n = 7 to 9/group). (A) Average daily consumption of either alcohol or water across the 14 days prior to QuA1 in male and female mice. Two-way RM ANOVA of day and sex in the alcohol history mice revealed a significant main effect of day (***p = 0.0006) and sex (*p = 0.01). (B) QuA consumption in male and female water and alcohol history mice on QuA Test 1. Two-way ANOVA of sex and drinking history on QuA Test 1 revealed a main effect of sex (**p = 0.0023). (C) Alcohol consumption at baseline and QuA consumption at QuA Test 1 for male and female alcohol history mice. Two-way RM ANOVA of day (baseline vs. QuA Test 1) and sex in alcohol history mice on consumption revealed a significant main effect of sex (****p = 0.0003) and day (**p = 0.0045). D. Unpaired t-test of QuA consumption on Test 1 displayed as percent baseline between male and female alcohol history mice did not reveal a significant difference. Note the change in y-axis between Figures (A) and (B-C). Data are displayed as mean ± SEM.

Fig. 2.

Presence of compulsive-like alcohol drinking phenotype after 3 weeks of DID. The same male and female C57BL/6J mice from Fig. 1 consumed either alcohol or water for an additional 7 days. On day 22, all mice were given QuA (QuA Test 2; n = 7 to 9/group). (A) Average daily consumption of either alcohol or water across the 21 days prior to QuA Test 2 in male and female mice. Two-way RM ANOVA of day and sex in the alcohol history mice revealed a significant main effect of day (***p = 0.0004) and sex (**p = 0.0033). (B) QuA consumption in male and female water and alcohol history mice on QuA Test 2. Two-way ANOVA of sex and drinking history on QuA Test 2 revealed a main effect of drinking history (**p = 0.0016). (C) Alcohol consumption at baseline and QuA consumption at QuA Test 2 for male and female alcohol history mice. Two-way RM ANOVA of day (baseline vs. QuA Test 2) and sex in alcohol history mice on consumption revealed a significant main effect of sex (*p = 0.0495). (D) Unpaired t-test of QuA consumption on Test 2 displayed as percent baseline between male and female alcohol history mice did not reveal a significant difference. Note the change in y-axis between Figures (A) and (B-C). Data are displayed as mean ± SEM.

Fig. 3.

Repeated QuA exposures do not alter compulsive-like QuA drinking. A separate group of male and female C57BL/6J mice consumed either alcohol or water for 21 days. On days 22, 24, and 26, all mice were given QuA (QuA Test 1 to 3; n = 12/group). (A) Average daily consumption of either alcohol or water across the 21 days prior to QuA Test Days 1 to 3 in male and female mice. Two-way RM ANOVA of day and sex in the alcohol history mice revealed a significant main effect of day (****p < 0.0001) and sex (*p = 0.032). (B) QuA drinking in male and female water and alcohol history mice across QuA Test Days. Three-way RM ANOVA of sex, drinking history, and QuA Test Day on QuA consumption revealed a significant main effect of QuA Test Day (***p = 0.0004), sex (***p = 0.0001), and drinking history (***p = 0.0002). (C) Consumption on baseline and QuA drinking sessions across QuA Test Days in male and female alcohol history mice. Three-way RM ANOVA of QuA Test Day, sex, and day (respective baseline vs. QuA Test Day) on consumption revealed a significant main effect sex (***p = 0.0001). (D) QuA consumption as percent baseline in male and female alcohol history mice. Two-way RM ANOVA of QuA Test Day and sex displayed as percent baseline consumption for each QuA Test Day revealed no significant main effects or interactions. Data are displayed as mean ± SEM.

Fig. 4.

Repeated QuA exposures alter front-loading of compulsive-like QuA drinking. 30-minute front-loading of QuA in the same group of male and female C57BL/6J mice as in Fig. 3. On days 22, 24, and 26, all mice were given QuA and 30-minute bottle readings were taken (QuA Test 1 to 3; n =−12/group). (A) Three-way RM ANOVA of QuA Test Day, sex, and drinking history on 30-minute QuA consumption revealed a significant main effect of test day (****p < 0.0001), sex (**p = 0.0067), and drinking history (****p < 0.0001). (B) Three-way RM ANOVA of QuA test day, sex, and day (i.e. respective baseline vs. QuA Test Day) on 30-minute QuA front-loading revealed a significant effect sex (**p = 0.009), QuA test day (****p < 0.0001), and day (respective baseline vs. QuA Test Day) (****p < 0.0001). (C) Two-way RM ANOVA of QuA Test Day and sex displayed as percent baseline consumption for each QuA Test Day revealed no significant main effects or interactions. Data are displayed as mean ± SEM.

Fig. 5.

Blood Alcohol Concentrations on QuA Test Day 3. BACs were taken immediately after DID on QuA Test Day 3 (n = 12/group). (A) Two-way RM ANOVA of sex and drinking history on BACs revealed a significant main effect of drinking history (*p = 0.0183). Data are displayed as mean ± SEM. (B) Pearson’s correlation of 30-minute front-loading of QuA on Test Day 3 and BACs taken immediately after the 2-hour DID session did not reveal a significant relationship. Pearson’s correlation of 2-hour QuA consumption on Test Day 3 and BACs taken immediately after the 2-hour DID revealed a significant relationship (****p < 0.0001).