Reginald D. Cannady, BS - US grants

Project Summary/Abstract The behavioral and molecular mechanisms underlying excessive alcohol drinking behavior and relapse are not fully understood and are vital for mapping the pathological course of alcoholism/dependence. Recent evidence indicates that chronic alcohol consumption leads to strengthening of excitatory synapses in key limbic brain regions that mediate reward and drinking behavior. This strengthening of synapses is highly dependent on the actions and synaptic incorporation of alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid receptors (AMPARs);fast action ion channel receptors that are activated by the excitatory neurotransmitter glutamate. However, the mechanistic role of enhanced AMPAR signaling in alcohol reinforcement and alcohol-seeking behavior remains unclear. Thus, the proposed experiments in this application seek to elucidate the behavioral and molecular mechanisms that underlie AMPAR-mediated increases in operant alcohol-self-administration and potentiated relapse-like behavior using a preclinical model of high alcohol consumption, the alcoholpreferring (P-) rat. Preliminary data indicate that enhancement of AMPAR signaling by pretreatment with aniracetam (positive allosteric modulator of AMPARs), increases operant alcohol self-administration and potentiates cue-induced reinstatement to alcohol seeking in P-rats, suggesting that enhanced AMPAR activity may be critical in facilitating increased drinking and susceptibility to relapse. Experiments will further characterize the role of enhanced AMPAR signaling in modulating operant self-administration and relapse-like behavior. AMPAR activity can be potentiated by post-translational modification (e.g. phosphorylation of the AMPAR GluR1 subunit amino acid residue 831;pGluR1{831}). Using immunohistochemistry techniques, experiments will map neuroadaptive changes in pGluR1{831} subunits in limbic brain regions after a history of alcohol self-administration or exposure to an alcohol-related cue during cue-induced reinstatement to determine brain regions that may influence enhanced AMPAR-mediated increases in self-administration and alcohol-seeking. We predict to see changes in pGluR1{831} in the nucleus accumbens (self-administration studies) or amygdala (reinstatement studies), and these regions will be targeted to investigate functional neuroanatomical control of enhanced AMPAR activity-mediated facilitation of alcohol self-administration and seeking behavior using aniracetam. Control experiments will address neuroanatomical and reinforcer specificity and non-specific locomotor effects. Lastly we will determine if aniracetam-induced increases in alcohol self-administration and seeking behavior are dependent on Ca2+/calmodulin-dependent protein kinase II (CamKII;known to phosphorylate GluR1 and enhance AMPAR activity) by blocking aniracetam-induced effects during self administration and reinstatement sessions with KN93 (CamKII inhibitor). Key findings from these studies will provide novel insight into AMPAR-related mechanisms in excessive alcohol drinking behavior and vulnerability to relapse.

Project Summary/ Abstract Individuals with alcohol use disorder (AUD) show deficits in cognitive function and an inability to regulate ethanol consumption and seeking. These deficits are driven by medial prefrontal cortex (mPFC) dysfunction. Ethanol dependence induced by chronic intermittent ethanol (CIE) vapor exposure disrupts expression of mPFC Kcnc1. This gene encodes voltage-dependent KV3.1 potassium channels, which are highly enriched in parvalbumin positive fast-spiking interneurons (PV+FSIs). Our preliminary data show that CIE-induced dysregulation of mPFC PV+FSI activity is restored by a novel KV3 channel positive modulator which also reduced drinking in dependent and non-dependent mice. These data implicate aberrant KV3 channel activity in PV+FSIs with dependence- induced deficits in mPFC function. We hypothesize that dependence-induced excessive drinking, ethanol reinforcement, and cue-induced relapse-like behavior are regulated by disruption of mPFC KV3 channel activity in PV+FSIs. Specific Aim 1 will examine functional neuroadaptations of KV3 channel activity in mPFC PV+FSIs of G42 (PV-GFP) transgenic mice exposed to CIE or Air, and test the effects of KV3 positive modulators to restore dependence-induced adaptations in PV+FSI function. We will also test if reduced KV3 channel expression is an underlying cellular mechanism driving aberrant KV3 channel activity in PV+FSIs. Specific Aim 2 will test if enhancing KV3 channel activity in the mPFC will reduce ethanol consumption in CIE and Air-exposed mice during home cage drinking sessions. Studies will also determine if KV3 positive modulation can restore mPFC PV+FSI activity during home cage drinking sessions in PV-Cre mice using in vivo fiber photometry techniques. Specific Aim 3 (R00 phase) will shift from studying home cage drinking to studying the role of mPFC KV3 channels in regulating the reinforcing effects of ethanol and relapse-like behavior by using operant conditioning procedures in dependent and non-dependent rats. Using in vivo fiber photometry, studies will test if positive modulation of KV3 increases Ca2+ transients in PV+FSIs in CIE-exposed PV-Cre rats, and in turn, decreases motivation to self-administer ethanol. Experiments will also determine if enhanced KV3 activity reduces ethanol-seeking behavior and modifies Ca2+ transients in PV+FSIs during cue-induced reinstatement tests in CIE-exposed PV-Cre rats. Collectively, these studies will validate KV3 channels as a novel target for the development of effective treatment options for ethanol dependence. In addition, the proposed experiments will provide training in emerging techniques used to selectively measure cellular function in sub-populations of cortical neurons of behaving transgenic animals.