Functional and morphological adaptation of medial prefrontal corticotropin releasing factor receptor 1-expressing neurons in male mice following chronic ethanol exposure

Abstract

Chronic ethanol dependence and withdrawal activate corticotropin releasing factor (CRF)-containing GABAergic neurons in the medial prefrontal cortex (mPFC), which tightly regulate glutamatergic pyramidal neurons. Using male CRF1:GFP reporter mice, we recently reported that CRF1-expressing (mPFC^CRF1+) neurons predominantly comprise mPFC prelimbic layer 2/3 pyramidal neurons, undergo profound adaptations following chronic ethanol exposure, and regulate anxiety and conditioned rewarding effects of ethanol. To explore the effects of acute and chronic ethanol exposure on glutamate transmission, the impact of chronic alcohol on spine density and morphology, as well as persistent changes in dendritic-related gene expression, we employed whole-cell patch-clamp electrophysiology, diOlistic labeling for dendritic spine analysis, and dendritic gene expression analysis to further characterize mPFC^CRF1+ and mPFC^CRF1− prelimbic layer 2/3 pyramidal neurons. We found increased glutamate release in mPFC^CRF1+ neurons with ethanol dependence, which recovered following withdrawal. In contrast, we did not observe significant changes in glutamate transmission in neighboring mPFC^CRF1− neurons. Acute application of 44 mM ethanol significantly reduced glutamate release onto mPFC^CRF1+ neurons, which was observed across all treatment groups. However, this sensitivity to acute ethanol was only evident in mPFC^CRF1− neurons during withdrawal. In line with alterations in glutamate transmission, we observed a decrease in total spine density in mPFC^CRF1+ neurons during dependence, which recovered following withdrawal, while again no changes were observed in mPFC^CRF− neurons. Given the observed decreases in mPFC^CRF1+ stubby spines during withdrawal, we then identified persistent changes at the dendritic gene expression level in mPFC^CRF1+ neurons following withdrawal that may underlie these structural adaptations. Together, these findings highlight the varying responses of mPFC^CRF1+ and mPFC^CRF1− cell-types to acute and chronic ethanol exposure, as well as withdrawal, revealing specific functional, morphological, and molecular adaptations that may underlie vulnerability to ethanol and the lasting effects of ethanol dependence.