All ETDs from UAB

Advisory Committee Chair

Jeremy J Day

Advisory Committee Members

Robinna G Lorenz

Erik D Roberson

David G Standaert

Linda S Overstreet-Wadiche

Document Type

Dissertation

Date of Award

2020

Degree Name by School

Doctor of Philosophy (PhD) Heersink School of Medicine

Abstract

Drug addiction is a chronic relapsing condition inflicting tremendous harm to individuals and society with ineffective treatment options available to most people. Drugs of abuse elevate dopamine levels in a brain region known as the nucleus accumbens (NAc), and activate gene programs considered essential for producing lasting synaptic and cell state changes which underlie the generation of addictive behavior. These gene expression changes remain poorly understood due to a complex heterogenous cellular architecture, and the rapidly fluctuating nature of the transcriptional processes themselves which make investigation difficult. Recent technical advances increase access to both profiling and manipulating these transcriptional changes to attribute functional value, but various challenges stood to be overcome to adapt them for use in this context. Here, we leverage single-nuclei sequencing to profile the transcriptional alterations occurring in the NAc during acute cocaine experience at the individual cell level. This allows insight into both the general cellular composition of the NAc and the inducible gene expression changes occurring during reward learning. Specifically, we identify the neuronal and non-neuronal subpopulations which become transcriptionally activated during drug experience, and define the gene expression signature of the early stages of reward learning within these individual cells. Additionally, we advance CRISPR/dCas9 technology capable of precise and accurate manipulation of gene transcription for ready functional use in the nervous system. This technology achieves robust targeted gene transcript specific suppression in post-mitotic neurons across diverse gene targets enabling the powerful and precise level of control required to assign functional valence to transcriptional events, such as those we characterize during reward learning. Finally, we identify and overcome a potential hurdle towards extending and utilizing light-inducible CRISPR/dCas9 technologies for functional interrogation of the rapid gene fluctuations which generate memory storage in the nervous system with temporal precision. Together, these investigations define the genome-wide transcriptional response of a vital heterogenous brain region with cellular precision during reward learning and extend capability to investigate essential processes underlying brain function. These advances and studies together generate a map of reward learning and provide the tools required to navigate and improve it to thereby identify therapeutic targets within the molecular processes which produce learning, memory, and addiction.

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