Opto-seq reveals input-specific immediate-early gene induction in ventral tegmental area cell types.

The ventral tegmental area (VTA) is a critical node in circuits governing motivated behavior and is home to diverse populations of neurons that release dopamine, gamma-aminobutyric acid (GABA), glutamate, or combinations of these neurotransmitters. The VTA receives inputs from many brain regions, but a comprehensive understanding of input-specific activation of VTA neuronal subpopulations is lacking. To address this, we combined optogenetic stimulation of select VTA inputs with single-nucleus RNA sequencing (snRNA-seq) and highly multiplexed in situ hybridization to identify distinct neuronal clusters and characterize their spatial distribution and activation patterns. Quantification of immediate-early gene (IEG) expression revealed that different inputs activated select VTA subpopulations, which demonstrated cell-type-specific transcriptional programs. Within dopaminergic subpopulations, IEG induction levels correlated with differential expression of ion channel genes. This new transcriptomics-guided circuit analysis reveals the diversity of VTA activation driven by distinct inputs and provides a resource for future analysis of VTA cell types.