INVESTIGATING THE ROLE OF DOPAMINE D3 RECEPTOR-EXPRESSING NEURONS IN WAKEFULNESS

Our understanding of how the brain regulates activity by reorganizing neural networks across the sleep–wake cycle remains incomplete. Because wakefulness and behavior interact in complex ways, it is crucial to identify the specific cell populations that both control arousal and shape behavioral outcomes. The goal of this study was therefore to identify neuronal populations that not only initiate arousal but also influence how behavior emerges from brain activity. We performed single-nucleus RNA sequencing on the ventral striatum under sleep, wake, and sleep deprivation conditions, and also included Sik3 mutant mice with chronically elevated sleep need. This allowed us to identify a distinct population of neurons in the islands of Calleja that express D3 receptors (D3Rs) and display a transcriptional profile associated with wakefulness. This population is molecularly separable from the D1/D3 co-expressing populations that are known to promote wakefulness. We hypothesized that this cell population may regulate arousal and complex behaviors. To target this population, we identified two marker genes selectively expressed in D3R-positive neurons of the islands of Calleja and generated new Cre-driver mouse lines specific to this cell population. We confirmed D3R and marker-gene expression using in situ hybridization, and then employed Cre-dependent AAV-hM3Dq/hM4Di to chemogenetically modulate these neurons while simultaneously measuring extracellular dopamine. This approach enables us to define how activity in this cell population contributes to arousal regulation and behavioral state control.