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Research Component 5:  Cortical Dynorphinergic Circuits in Ethanol Drinking

Primary Investigators: Dr. Thomas Kash, Dr. Monserrat Navarro

Excessive ethanol consumption is a major public health issue that can drive a range of serious health problems. Ethanol has been shown to have both aversive and rewarding properties, and it has been hypothesized that tolerance to the aversive properties of ethanol contributes to excessive intake. The neural circuits and mechanisms that are responsible for the aversive properties of ethanol are not well understood. The anterior insular cortex (aIC) is a brain structure that has been shown to be involved in multiple aspects of ethanol consumption; however, its role in regulation of the aversive properties remains unknown. Notably, recent studies have found that the aIC is a critical substrate for regulation of both innate and learned aspects of taste aversion. Based on this, our central hypothesis is that aversive properties of ethanol are encoded in the insular cortex, and that chronic ethanol disrupts excitatory/inhibitory (E/I) balance in the aIC, disrupting the aversive properties of ethanol, contributing to escalated ethanol consumption.

Aim 1. Test the hypothesis that the retrieval of ethanol-induced conditioned taste aversion (CTA) enhances GABAergic inhibition onto aIC efferents to basolateral amygdala outputs (aICBLA). The aICBLA is required for acquisition and retrieval of CTA to lithium chloride (LiCl). Intact aIC Parvalbumin (aICPV) inhibitory interneuron function is required for retrieval of CTA. It is unknown if these requirements generalize to the retrieval of the conditioned aversive properties of ethanol. In this aim, we will use slice electrophysiology to explore plasticity in these aIC circuit elements during ethanol CTA retrieval. We hypothesize that there will be a reduction in E/I balance in this pathway due to an upregulation of GABAergic transmission following ethanol CTA. Finally, we will use chemogenetic inhibition approaches to explore the necessity of PV interneurons during retrieval of ethanol CTA. These experiments will determine the role of defined aIC circuit elements in conditioned aversive properties of ethanol.

Aim 2. Test the hypothesis that chronic ethanol exposure disrupts CTA retrieval via dysregulation of aICPV-mediated inhibition onto aICBLA efferents. In our preliminary results, we found that long-term ethanol drinking leads to escalated ethanol consumption, reduced ethanol-induced CTA, and increased E/I balance in aIC layer 5 pyramidal neurons. We hypothesize that this change in E/I balance is linked with tolerance to the aversive properties of ethanol and will be reflected in the inability to undergo ethanol CTA-induced plasticity.  We will use electrophysiology to explore plasticity in both aICPV and aICBLA neurons during retrieval of CTA following long-term ethanol drinking. We will then determine if chemogenetic activation of aICPV will restore CTA retrieval and reduce excessive ethanol consumption. Finally, we will use fiber photometry to assess the dynamics of the aIC outputs to the BLA during both ethanol consumption and retrieval of CTA.  These experiments will identify how aIC circuits link excessive ethanol intake with tolerance to the aversive properties of ethanol.

Aim 3. Conduct Center-wide collaborative studies to identify mechanisms of chronic ethanol -induced dysregulation of neural networks. Numerous studies across the ARC have identified circuit components that play critical roles in multiple aspects of ethanol-related behaviors. In this collaborative aim, we will examine if chronic ethanol exposure induced alterations in these key circuit components. First, we will use slice physiology combined with channelrhodopsin-assisted circuit mapping to explore changes in frontal-limbic connectivity between aIC, BLA, and Acb. In addition, utilizing the expertise of the Scientific Resource Core, we will use multi-site fiber photometry to measure E/I dynamics in the aIC and BLA during both ethanol consumption and ethanol CTA. Finally, we will collaborate with Research Component 3 to investigate how adolescent alcohol exposure alters aIC and prelimbic cortex function. Together, these studies will assess interspecies generality of alcohol-induced changes in functional network activity and corresponding mechanistic neural processes.

The goal of the UNC-ARC is to increase understanding of circuit pathogenesis in alcohol use disorder (AUD). To address this goal, Research Component 5 seeks to identify a novel circuit mechanism that drives tolerance to the aversive properties of ethanol, which is a hallmark of AUD. This work has significant potential to guide development of novel therapeutic approaches.