Viral-Mediated Knockdown of Nucleus Accumbens Shell PAC1 Receptor Promotes Excessive Alcohol Drinking in Alcohol-Preferring Rats

Abstract

Alcohol use disorder (AUD) is a chronic, relapsing disorder whose genetic and environmental susceptibility components are not fully understood. Neuropeptidergic signaling has been repeatedly implicated in modulating excessive alcohol drinking, especially within sub-regions of the striatum. Here, we investigated the potential involvement of the selective receptor for pituitary adenylate cyclase-activating polypeptide (PACAP), PAC1R, in the nucleus accumbens shell (NAcc Shell) in excessive alcohol drinking in alcohol-preferring rats, an established animal model of the genetic propensity for alcoholism. Scr:sP alcohol-preferring rats were trained to operantly self-administer alcohol and then either an AAV virus short-hairpin RNA (shRNA) targeted to knockdown PAC1R, or an AAV control virus were microinfused into the NAcc Shell. NAcc Shell PAC1R shRNA knockdown virus was confirmed to significantly decrease PAC1R levels in the NAcc Shell. The effects of NAcc Shell PAC1R shRNA knockdown on ethanol self-administration were investigated using a Fixed Ratio (FR) 1 and a Progressive Ratio (PR) schedule of reinforcement. The effect of PAC1R knockdown on self-administration of an alternative reinforcer, saccharin, was also assessed. The results showed that the reduction in PAC1R in the NAcc Shell led to excessive ethanol drinking, increased preference for ethanol, and higher motivation to drink. NAcc Shell PAC1R shRNA knockdown did not comparably increase saccharin self-administration, suggesting selectivity of action. These data suggest that NAcc Shell PAC1R may serves as a “brake” on alcohol drinking, and thereby the loss of function of PAC1R leads to excessive alcohol consumption. Therefore, the PACAP/PAC1R system may represent a novel target for the treatment of AUD.

The Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) System of the Central Amygdala Mediates the Detrimental Effects of Chronic Social Defeat Stress in Rats

“…The needle was kept in place for an additional 10 min following infusion to prevent backflow. The ability of this specific AAV-shRNA construct to knockdown PAC1R expression in rats was previously confirmed (Minnig et al, 2021;Minnig et al, 2022). After surgeries, rats were allowed at least 3 weeks of recovery before the start of the CSDS procedure to allow for maximum transfection.…”

Pituitary Adenylate Cyclase-Activating Polypeptide (PACAP) of the Bed Nucleus of the Stria Terminalis Mediates Heavy Alcohol Drinking in Mice

“…PAC1R positive cells quantification. The number of PAC1R expressing cells was quantified using an unbiased stereological approach as previously described (Ferragud et al, 2020) (Ferragud et al, 2021;Minnig et al, 2021). The STLD of the BNST was outlined with an Olympus PlanApo N x2 objective (numerical aperture 0.08), and counting was done using an Olympus UPlanFL N x20 objective (num.…”

“…Suggesting a likely involvement of the PACAP system in AUD, a single nucleotide polymorphism of the PACAP gene was linked to high alcohol intake in a European population (Kovanen et al, 2010); furthermore, a specific genotype of the PAC1R was associated with problematic alcohol use in women (Dragan et al, 2017). In the preclinical realm, the PACAP/PAC1R system has also been shown to be involved in the actions of alcohol by pharmacological and genetic deletion studies (Feany and Quinn, 1995;Tanaka et al, 2004;Tanaka et al, 2010;Gargiulo et al, 2020;Minnig et al, 2021;Minnig et al, 2022). In particular, in the BNST, it has recently been shown that rats made dependent via chronic exposure to alcohol vapors display increased levels of PACAP, and that the pharmacological antagonism of PAC1R in this area is able to block dependence-induced excessive alcohol intake as well as anxiety-like behavior in these rats (Ferragud et al, 2021).…”

Pharmacological modulation of ventral tegmental area neurons elicits changes in trigeminovascular sensory processing and is accompanied by glycemic changes: Implications for migraine

“…Sections were then mounted with DAPI. Primary antibodies were chosen based on recent published studies (40)(41)(42)(43) and the specificity was demonstrated by the suppliers. Additionally, samples were processed with the omission of primary antibodies, and the staining of brain areas known to contain each receptor was performed.…”