Involvement of DNA Methylation in Alcohol Withdrawal-Induced Behavioral Changes in Rats

Alcohol use disorder (AUD) poses a significant burden to global health and the economy. Individuals with AUD often self-medicate with alcohol to alleviate the negative affective symptoms of withdrawal, such as anxiety, which often results in a relapse to heavy drinking. The identification of novel therapeutic targets that can reduce withdrawal-induced increases in anxiety and drinking can help alleviate the burden of AUD. DNA methylation is an epigenetic mechanism that is dysregulated in AUD. Inhibitors of the enzymes that promote DNA methylation (DNMTs) have been used to treat certain forms of cancer, however their potential to treat symptoms of alcohol withdrawal is less understood. To address this gap, we used a rat model of alcohol dependence to explore the role of DNA methylation in both withdrawal-induced anxiety-like behavior and changes in alcohol drinking. We found that acute withdrawal produces subregion-specific increases in DNMT1 and DNMT3b protein and mRNA in the amygdala – a key region regulating anxiety. Systemic administration of the FDA-approved DNMT inhibitor, 5-azacytidine (5-AzaC), reversed the withdrawal-induced increase in anxiety-like behavior exhibited by these rats. Administration of 5-AzaC also reduced ethanol intake in a home cage drinking paradigm in both dependent and non-dependent rats. Taken together, these results suggest that DNA methylation is a potential therapeutic target to prevent relapse in AUD. In addition to heavy, uncontrolled drinking, significant time spent seeking out alcohol is also a common symptom of AUD, yet home cage drinking paradigms are unable to capture this behavior. Instead, operant ethanol self-administration paradigms are used to discriminate between appetitive (seeking) and consummatory (taking) behaviors. Curiously, however, most studies infer consumption indirectly from appetitive responding. To address this shortcoming, we compared operant behavior across three strains of outbred rats using a paradigm that directly measures both appetitive and consummatory behaviors. Our data uncovered three unique operant phenotypes and revealed that appetitive responding is correlated with consumption in only a subset of rats. These data suggest that direct measurement of consumption during operant self-administration is crucial. This approach will be useful for examining the effect of DNMT inhibition on both seeking and taking in the future.