Chemogenetic Activation of Parasubthalamic nucleus Corticotropin‐Releasing Factor Neurons recapitulates Alcohol Withdrawal‐Related Behavioral Symptomatology

Abstract

Negative reinforcing effects of alcohol, driven by negative affect during withdrawal, are thought to be crucial to drinking escalation in alcohol use disorders. While the brain regions and neurotransmitter signaling affected by chronic alcohol use are well characterized, our understanding of the specific neural circuits driving drinking escalation and negative affect during withdrawal remains limited. Animal studies have highlighted a critical role of corticotropin‐releasing factor (CRF) signaling in the behavioral symptomatology of alcohol dependence. Hence, the present study sought to examine the role of a cluster of CRF neurons located in the parasubthalamic nucleus (PSTN), a small region of the posterior hypothalamus, which has received little attention in behaviors related to alcohol dependence. Adult male Crh‐IRES‐Cre mice were bilaterally injected in the PSTN with a viral vector encoding the excitatory designer receptor hM3D(Gq) fused to the fluorescent reporter mCherry upon Cre recombination. Injection of the hM3D(Gq) agonist clozapine‐N‐oxide (CNO, 1 mg/kg) significantly increased alcohol and saccharin consumption but did not change food and water consumption compared to vehicle. CNO had no effect in mice expressing mCherry only. CNO also increased locomotor and digging activity but did not alter open arm exploration in the elevated plus maze. CNO also decreased grooming duration in the splash test and immobility duration in the tail suspension test. Interestingly, the increased alcohol consumption, digging activity, active coping and reduced grooming all recapitulate phenotypes observed in mice withdrawn from chronic intermittent ethanol vapor inhalation, a model of alcohol dependence. However, CNO injection caused a significant increase in nociceptive thresholds in both the tail immersion and tail pressure tests, which is at odds with the hyperalgesia typically associated with alcohol withdrawal.The PSTN sends a strong projection to the central amygdala (CeA), a region that plays a critical role in the transition to alcohol dependence. We therefore tested whether the effects of PSTN CRF neuron chemogenetic activation may be mediated by the activation of CeA neurons carrying CRF1 receptors. However, CNO did not affect alcohol drinking, locomotion and digging in Crhr1‐IRES‐Cre mice expressing hM3D(Gq) in the CeA. Combined with our recent finding that PSTN neurons become activated during alcohol withdrawal, our results suggest that PSTN CRF neurons contribute to the behavioral symptomatology of ethanol withdrawal, but CRF1 signaling in the CeA may not be relevant to their mechanism of action.Support or Funding InformationThis work was funded by the following grants from the National Institutes of Health: AA026685 (CC), P60 AA006420 (CC), and University of Benin, Benin City, Nigeria, (AO).