Date of Award

Spring 4-9-2025

Embargo Period

4-21-2025

Document Type

Dissertation

Degree Name

Doctor of Philosophy (PhD)

Department

Neuroscience

College

College of Graduate Studies

Additional College

College of Medicine

First Advisor

Christopher W. Cowan

Second Advisor

Peter W. Kalivas

Third Advisor

Thomas Jhou

Fourth Advisor

Donald R. Menick

Fifth Advisor

Makoto Taniguchi

Sixth Advisor

Danyelle M. Townsend

Abstract

Substance Use Disorder (SUD) is a relapsing disorder characterized by repeated seeking and use of psychoactive substances despite negative consequences to the individual. Epigenetic changes within the medium spiny neurons (MSNs) of the nucleus accumbens (NAc) are thought to underly the lasting propensity to relapse in SUD. The epigenetic enzyme histone deacetylase 5 (HDAC5) in NAc MSNs limits the formation of reward-cue associations in rodent models of drug seeking, but not sucrose seeking, leading to decreased relapse-like behavior. However, whether HDAC5 reduces drug seeking behavior through its own deacetylase activity or through the recruitment of co-repressors is unknown. Furthermore, these HDAC5-mediated behavioral changes are associated with a decrease in MSN intrinsic excitability and the downregulation of numerous genes associated with transmembrane cation transport. One of these genes, Scn4b, encodes a sodium channel auxiliary subunit that functions to increase action potential frequency in MSNs. We hypothesized that HDAC5 limits relapse-like behavior through reducing MSN intrinsic excitability via the deacetylase-dependent downregulation of Scn4b.

To test this hypothesis, we first characterized a conserved cysteine-containing structural zinc-binding motif within HDAC5’s enzymatic domain using targeted mass spectrometry, molecular dynamics modeling, and enzymatic and transcriptional activity assays. We found that HDAC5 exhibits high basal levels of an intramolecular disulfide bond at this motif. Mutation of the involved cysteine residues selectively disrupts HDAC5’s intrinsic deacetylase activity without disrupting co-factor association or altering subcellular localization, indicating a critical involvement of these cysteine residues in the intrinsic deacetylase activity of HDAC5.

We next assessed the role of HDAC5’s intrinsic deacetylase activity in relapse-like cocaine seeking behavior and NAc MSN electrophysiology. Using viral-mediated gene transfer in a rat cocaine self-administration paradigm, we found that HDAC5’s intrinsic deacetylase activity is required to limit cue-induced reinstatement of cocaine seeking. This behavioral effect was associated with a deacetylase-dependent reduction of NAc MSN intrinsic excitability, indicating that HDAC5’s enzymatic activity regulates relapse-like behavior through the modulation of NAc MSN firing rate.

Finally, we investigated the HDAC target gene, Scn4b, as a critical mediator of HDAC5’s electrophysiological and behavioral effects. Short hairpin RNA-mediated reduction of NAc Scn4b transcript levels limited MSN intrinsic excitability and reduced cue-induced reinstatement of cocaine seeking, while having no effect on sucrose seeking behavior.

Taken together, the data in this dissertation provide new insights into the molecular mechanism of HDAC5’s limiting of relapse-like behavior. It positions NAc MSN intrinsic excitability as a major factor underlying the establishment of lasting drug-cue, but not natural reward-cue, associations and reveals Scn4b to be a critical HDAC5 target gene mediating these processes.

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