The contribution of neuronal and microglia proinflammatory signaling in insular cortex on escalated ethanol self-administration

Abstract
The induction of pro-inflammatory signaling is a key pathologic feature of alcohol use disorder (AUD). This
includes increased expression of Toll-like receptor (TLR) signaling systems as well as activation of a variety of
proinflammatory cytokines. Since many TLRs are involved in AUD pathology, it is important to study downstream
mediators that are shared by multiple TLRs, in circuits that regulate AUD-related behaviors. Therefore, it is
important that we find we find that agonists to TLR3 and 7 as well as chronic ethanol exposure all result in robust
downstream induction of the proinflammatory transcription factor interferon regulator factor-7 (IRF7) in the
anterior insular cortex (aIC) and nucleus accumbens core (AcbC). This induction in these brain regions is relevant
as our work implicates aIC→AcbC circuitry as critical in the maintenance of alcohol self-administration.
Interestingly, IRF7 was primarily localized in neurons along with pro-inflammatory activation of microglia.
Microglia are key initiators of inflammatory signaling in brain and are capable of promoting neuronal and circuit
adaptations to ethanol. Microglial pro-inflammatory cytokines further alter neuronal excitatory/inhibitory (E/I)
balance – an overall theme of the UNC Alcohol Research Center (ARC) – as well as functional connectivity.
Therefore, we hypothesize that pro-inflammatory microglia in aIC promote neuronal IRF7 and subsequent
excitatory outflow from the aIC to AcbC to promote alcohol self-administration. The overall hypothesis of the
present project is that neuronal IRF7 induction and pro-inflammatory microglial activation as a consequence of
alcohol vapor exposure together alter E/I balance across aIC→Acb circuitry to promote increases in alcohol self-
administration. Experiments in Aim 1 assess IRF7 signaling in aIC projection neurons and determine if IRF7
promotes escalations in alcohol self-administration following alcohol vapor exposure. Experiments in Aim 2 will
examine proinflammatory microglia signaling in the aIC including E/I balance using gene expression analyses
and determine the functional role of aIC proinflammatory microglia in vapor-induced increases in alcohol self-
administration. Lastly, the collaborative studies in Aim 3 will directly determine the effects of the alcohol vapor
exposure on the activity of aIC→Acb projection neurons using electrophysiology in collaboration with Research
Component 1 and investigate the role of microglia on brain-wide connectivity across binge ethanol models using
resting state functional connectivity magnetic resonance imaging (rs-fcMRI) and magnetic resonance
spectroscopy (MRS) in collaboration with Research Components 3 and 4 and the Scientific Resource Core. The
overall goal of the UNC ARC is to increase understanding of molecular and cellular pathogenesis in alcohol use
disorder. To this end, the studies in Research Component 2 will determine the molecular consequences of
chronic high-dose alcohol exposure via vapor exposure on IRF7 signaling and microglial activation and how
these changes promote subsequent escalations in alcohol self-administration.

Grant Number: 5P60AA011605-29
NIH Institute/Center: NIH
Principal Investigator: Joyce Besheer