Chemogenetic stimulation of mouse central amygdala corticotropin-releasing factor neurons: Effects on cellular and behavioral correlates of alcohol dependence

Abstract

Background. Corticotropin-releasing factor (CRF) signaling in the central nucleus of the amygdala (CeA) plays a critical role in rodent models of excessive alcohol drinking.However, the source of CRF acting in the CeA during alcohol withdrawal remains to be identified. In the present study, we hypothesized that CeA CRF interneurons may represent a behaviorally relevant source of CRF to the CeA increasing motivation for alcohol via negative reinforcement.Methods. We tested this hypothesis in male mice and used chemogenetics to stimulate CeA CRF neurons in vitro and in vivo.Results. We first observed that Crh mRNA expression in the anterior part of the mouse CeA, at the junction with the interstitial nucleus of the posterior limb of the anterior commissure, correlates positively with alcohol intake in C57BL/6J males with a history of chronic binge drinking. We then found that chemogenetic activation of CeA CRF neurons in Crh-IRES-Cre mouse brain slices increases gamma-aminobutyric acid (GABA) release in the medial CeA in part via CRF1 receptor activation, indicating local CRF release. While chemogenetic stimulation of CeA CRF neurons exacerbated novelty-induced feeding suppression, as seen in C57BL/6J males withdrawn from chronic intermittent alcohol inhalation, it had no effect on voluntary alcohol consumption, following either acute or chronic manipulation.Conclusions. Altogether, these findings indicate that hyperactivity of CeA CRF neurons may contribute to elevated CeA GABA levels and negative affect during alcohol withdrawal but is not sufficient to drive alcohol intake escalation in dependent mice. amygdala (CeA) during alcohol withdrawal [12] and blockade of CRF1 signaling in the CeA reduces the anxiogenic-like effect of alcohol withdrawal [13,14], dependenceinduced increases in alcohol self-administration [15, 16], as well as heavy binge drinking [17]. CRF1 in the CeA also mediates the elevation of gamma-aminobutyric acid (GABA) 4 dialysate induced by alcohol dependence [18] and CRF1 overexpression in the CeA potentiates stress-induced reinstatement of alcohol seeking [19].Despite this wealth of converging evidence, the specific neurons responsible for the release of CRF in the CeA during alcohol withdrawal have not been identified. In the present study, we tested the hypothesis that these neurons are intrinsic to the CeA. This hypothesis was supported by evidence that Crh is upregulated in the CeA of alcoholdependent and post-dependent rats [18,20,21], along with our own findings (reported here) that Crh expression in the anterior part of the CeA correlates with alcohol intake in mice with a history of chronic alcohol drinking followed by abstinence. Importantly, psychological stress also upregulates Crh in the CeA [22][23][24] and CRF synthesis in the CeA plays a critical role in mediating anxiety-like behavior [25, 26]. In addition, CRF1expressing neurons in the CeA receive direct input from local interneurons [27] and CeA CRF neurons provide both local inhibitory GABA and excitatory CRF1-mediated sign...