Multiomic profiling of cell types mediating opioid use disorder in rats

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Project Summary The misuse and abuse of prescription pain relievers, such as oxycodone, contributed to the unprecedented opioid epidemic in the United States. The opioid crisis has devastating consequences on public health including a surge in opioid misuse and related overdoses. Research is urgently needed to develop better treatments for opiate addiction. Despite substantial knowledge of the pharmacokinetic and behavioral effects of oxycodone in various animal models, only a small number of candidate genes and neuroanatomical systems affected by opioids have been studied. Recent technological advances in the field of single cell genomics are promising avenues for the unbiased discovery and characterization of brain cell types that respond to opioids. In response to this RFA, we leverage an innovative multi-omics methodology (Single Cell Multiome ATAC + Gene Expression) to map the transcriptome and epigenome from the same cell across thousands of cells in brain regions relevant to the effects of opioid exposure. To this aim we will use a rat model of extended access to oxycodone intravenous self-administration that recapitulates several neuroadaptations also observed in humans with opioid use disorders (OUD). This approach provides an exceptional opportunity to systematically explore the cellular diversity of the opioid system and, at the same time, the causative mechanisms that regulate cellular states based on the associations between epigenetic changes and the expression of target genes in individual cells. We will integrate this innovative multi-omics methodology with rigorous computational approaches to explore the cellular organization the opioid system in multiple brain regions and different stages of OUD progression (initial exposure, escalation of use, acute withdrawal, prolonged abstinence, and cue- induced relapse). We have provided strong preliminary that support the feasibility of our proposed plan for the following aims. In Aim 1, we will collect brain tissues at different stages of the extended access to oxycodone intravenous self-administration (ivsa) protocol and we will generate single cell genomics data from both male and female rats that are exposed to either saline or oxycodone. In Aim 2, we will integrate these transcriptomic and epigenomic datasets to identify changes in cellular states, genes and upstream regulators that are associated with different stages of oxycodone use. This approach will facilitate the identification of linkages between cis- regulatory elements and target genes. In Aim 3, we will validate key cell type-specific findings by RNA-FISH and identify the top 3 target genes for functional validation. To this aim, we will use a viral-mediated CRISPR- Cas9 system to modulate addictive behaviors in rat models of oxycodone self-administration. The results of this study will enable future studies that may identify new targets for treatment and prevention of OUD.