Mechanisms of alcohol's effects on prefrontal cortex circuits - Abstract The prefrontal cortex (PFC) is essential for moderating drinking and regulating affective changes in alcohol use disorder (AUD). PFC activity is dynamically modulated by inhibitory interneurons that express vasoactive intestinal peptide (VIP-INs). Unique among interneurons, VIP-INs preferentially target other inhibitory neurons to indirectly enhance principal neuron activity. Here, we show preliminary data that chemogenetic stimulation of PFC VIP-INs reduces social approach behavior in control mice. Conversely, we find that chemogenetic inhibition of VIP-INs can enhance social preference following chronic drinking (intermitent access, IA ethanol). Thus, we believe that VIP-INs represent an an exciting cellular target to improve social and affective behaviors in AUD. Towards that end, the goal of this proposal is to better define the mechanisms through which alcohol and drinking affects VIP-IN function. One likely mechanism through which ethanol may modulate VIP-INs is by inhibiting KCNQ channels. Indeed, we provide preliminary data that ex vivo ethanol enhances VIP-IN excitability in prelimbic (PL)-PFC, that ethanol-induced hyperexcitability is occluded by a KCNQ channel inhibitor, and that ethanol directly inhibits KCNQ channel currents in VIP-INs but not pyramidal cells. KCNQ channels assemble as tetramers, and the common KCNQ3 subunit preferentially forms heteromeric channels with other subunits. Interestingly, we provide evidence that VIP-INs do not express KCNQ3, allowing KCNQ5 and/or KCNQ2 subunits to form heterodimeric KCNQ2/5 or homomeric channels. In this proposal, we hypothesize that long-term drinking decreases VIP-IN excitability through a homeostatic upregulation of KCNQ channel function. We will use CRISPR/SaCas9 molecular approaches to systematically test that unique expression of KCNQ subunits (low KCNQ3 and high KCNQ5) confers VIP-INs with high sensitivity to ethanol and to its long-term adaptations. Finally, we will test that adaptations to VIP-INs regulate drinking and associated affective disturbances. Aim 1: Test that KCNQ subunit stoichiometry governs ethanol’s actions in distinct cell types Aim 2: Test that long-term drinking alters KCNQ channel function on VIP-INs and ethanol sensitivity Aim 3: Test that PL-PFC KCNQ channels drive VIP-IN hypo-excitability following IA ethanol and abstinence- associated affective disturbances These studies will further characterize how PFC disinhibitory interneurons mediate responses to ethanol and drinking. The mechanistic CRISPR/SaCas9 approach will provide unprecedented insight into how specific KCNQ subunits mediate ethanol’s effects in native tissue, with potential to inform and motivate a new wave of drug discovery or repurposing for AUD medication development.
