A Bahi1,2, JL Dreyer3

Anxiety disorders frequently co-occur with alcohol use disorder (AUD), and mesolimbic cholinergic signaling has been implicated in the regulation of affective and reward-related behaviors. The muscarinic M4 receptor (M4R) is highly expressed in striatal circuitry, including the nucleus accumbens (NAc), where it may function as an inhibitory modulator of behavioral output. However, its role within the NAc in regulating anxiety-like and ethanol-related behaviors remains unclear. Adult mice received bilateral intra-accumbal injections of viral vectors expressing shRNA targeting M4R or a control construct. Anxiety-like behavior was assessed using the elevated plus maze (EPM) and open field (OF) tests. Voluntary ethanol intake and preference were subsequently measured using a two-bottle choice paradigm. Endpoint M4 mRNA expression within the NAc was quantified using real-time PCR, and expression levels were correlated with behavioral parameters. M4R loss-of-function in the NAc significantly increased anxiety-like behavior, indicated by reduced open-arm exploration in the EPM and decreased center time in the OF test. Following anxiety assessment, accumbal M4-deficient mice exhibited elevated voluntary ethanol intake and preference compared to controls. Saccharin and quinine consumption were unchanged, suggesting enhanced ethanol drinking was not attributable to altered taste sensitivity. Importantly, the degree of M4R mRNA reduction correlated positively with anxiety-like measures and ethanol intake. These findings demonstrate that M4R in the NAc serve as a critical inhibitory regulator of anxiety-like behavior and ethanol consumption, identifying accumbal M4R signaling as a potential therapeutic target for comorbid anxiety disorders and AUD.

1Department of Basic Medical Sciences, College of Medicine, Ajman University, Ajman, UAE

2Center of Medical & Bio-Allied Health Sciences Research, Ajman University, Ajman, UAE

3Division of Biochemistry, Department of Medicine, University of Fribourg, Fribourg, Switzerland