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Recent large genome-wide association studies (GWAS) have identified multiple confident risk loci linked to addiction-associated behavioral traits. Genetic variants linked to addiction-associated traits lie largely in non-coding regions of the genome, likely disrupting cis-regulatory element (CRE) function. CREs tend to be highly cell type-specific and may contribute to the functional development of the neural circuits underlying addiction. Yet, a systematic approach for predicting the impact of risk variants on the CREs of specific cell populations is lacking. To dissect the cell types and brain regions underlying addiction-associated traits, we applied LD score regression to compare GWAS to genomic regions collected from human and mouse assays for open chromatin, which is associated with CRE activity. We found enrichment of addiction-associated variants in putativeCREsmarked by open chromatin in neuronal (NeuN+) nuclei collected from multiple prefrontal cortical areas and striatal regions known to play major roles in reward and addiction. To further dissect the cell type-specific basis of addiction-associated traits, we also identified enrichments in human orthologs of open chromatin regions of mouseneuronal subtypes: cortical excitatory, D1, D2, and PV. Lastly, we developed machine learning models from mouse cell type-specific regions of open chromatin to further dissect human NeuN+ open chromatin regions into cortical excitatory or striatal D1 and D2 neurons and predict the functional impact of addiction-associated genetic variants. Our results suggest that differentneuronal subtypeswithin the reward system play distinct roles in the variety of traits that contribute to addiction.Significance StatementWe combine statistical genetic and machine learning techniques to find that the predisposition to for nicotine, alcohol, and cannabis use behaviors can be partially explained by genetic variants in conserved regulatory elements within specific brain regions and neuronal subtypes of the reward system.This computational framework can flexibly integrateopen chromatindata across species to screen for putative causal variants in a cell type-and tissue-specific manner across numerous complex traits.

Neuron subtype dysfunction is a key contributor to neurologic disease circuits, but identifying associated gene regulatory pathways is complicated by the molecular complexity of the brain. For example, parvalbumin-expressing (PV+) neurons in the external globus pallidus (GPe) are critically involved in the motor deficits of dopamine-depleted mouse models of Parkinson's disease, where cell type-specific optogenetic stimulation of PV+neurons over other neuron populations rescues locomotion. Despite the distinct roles these cell types play in the neural circuit, the molecular correlates remain unknown because of the difficulty of isolating rare neuron subtypes. To address this issue, we developed a new viral affinity purification strategy, Cre-Specific Nuclear Anchored Independent Labeling, to isolate Cre recombinase-expressing (Cre+) nuclei from the adult mouse brain. Applying this technology, we performed targeted assessments of the cell type-specific transcriptomic and epigenetic effects of dopamine depletion on PV+and PV–cells within three brain regions of male and female mice: GPe, striatum, and cortex. We found GPe PV+neuron-specific gene expression changes that suggested increased hypoxia-inducible factor 2α signaling. Consistent with transcriptomic data, regions of open chromatin affected by dopamine depletion within GPe PV+neurons were enriched for hypoxia-inducible factor family binding motifs. The gene expression and epigenomic experiments performed on PV+neurons isolated by Cre-Specific Nuclear Anchored Independent Labeling identified a transcriptional regulatory network mediated by the neuroprotective factor Hif2a as underlying neural circuit differences in response to dopamine depletion.SIGNIFICANCE STATEMENTCre-Specific Nuclear Anchored Independent Labeling is an enhanced, virus-based approach to isolate nuclei of a specific cell type for transcriptome and epigenome interrogation that decreases dependency on transgenic animals. Applying this technology to GPe parvalbumin-expressing neurons in a mouse model of Parkinson's disease, we discovered evidence for an upregulation of the oxygen homeostasis maintaining pathway involving Hypoxia-inducible factor 2α. These results provide new insight into how neuron subtypes outside the substantia nigra pars compacta may be compensating at a molecular level for differences in the motor production neural circuit during the progression of Parkinson's disease. Furthermore, they emphasize the utility of cell type-specific technologies, such as Cre-Specific Nuclear Anchored Independent Labeling, for isolated assessment of specific neuron subtypes in complex systems.