Cell type-specific barcoding reveals the single-neuron projectional architecture of the mouse midbrain dopaminergic...

Brain-wide neural circuits are formed by the diverse axonal branching patterns of neurons of different cell types. Here we introduce POINTseq (projections of interest by sequencing), a cell type-specific barcoded connectomics method that uses selective barcoding and sequencing to rapidly map single-cell projections of a cell type of interest for thousands of neurons per animal. POINTseq leverages pseudotyping of Sindbis virus and a specific alphavirus-cellular receptor pair to make Sindbis infections cell type specific. It thus integrates MAPseq-style high-throughput barcoded projection mapping with the established viral-genetic neural circuit analysis toolbox. We validated POINTseq by mapping genetically and projection-defined cell populations in the mouse motor cortex. We then applied POINTseq to midbrain dopaminergic neurons and reconstructed the brain-wide single-cell projections of 5,902 dopaminergic neurons in ventral tegmental area (VTA) and substantia nigra pars compacta (SNc). These neurons cluster into over 25 connectomic cell types, vastly exceeding the known diversity of dopaminergic cells, and form stereotyped projection motifs that may mediate parallel dopamine signaling. This data constitutes the anatomical substrate on which the diverse functions of dopamine in the brain are built.