The human brain has enormously complex cellular diversity and connectivities fundamental to our neural functions, yet difficulties in interrogating individual neurons has impeded understanding of the underlying transcriptional landscape. We developed a scalable approach to sequence and quantify RNA molecules in isolated neuronal nuclei from post-mortem brain, generating 3,227 sets of single neuron data from six distinct regions of the cerebral cortex. Using an iterative clustering and classification approach, we identified 16 neuronal subtypes that were further annotated on the basis of known markers and cortical cytoarchitecture. These data demonstrate a robust and scalable method for identifying and categorizing single nuclear transcriptomes, revealing shared genes sufficient to distinguish novel and orthologous neuronal subtypes as well as regional identity within the human brain.

Lysophosphatidic acid (LPA) is a small, ubiquitous phospholipid that acts as an extracellular signaling molecule by binding to and activating at least five known G protein-coupled receptors (GPCRs): LPA(1)-LPA(5). They are encoded by distinct genes named LPAR1-LPAR5 in humans and Lpar1-Lpar5 in mice. The biological roles of LPA are diverse and include developmental, physiological, and pathophysiological effects. This diversity is mediated by broad and overlapping expression patterns and multiple downstream signaling pathways activated by cognate LPA receptors. Studies using cloned receptors and genetic knockout mice have been instrumental in uncovering the significance of this signaling system, notably involving basic cellular processes as well as multiple organ systems such as the nervous system. This has further provided valuable proof-of-concept data to support LPA receptors and LPA metabolic enzymes as targets for the treatment of medically important diseases that include neuropsychiatric disorders, neuropathic pain, infertility, cardiovascular disease, inflammation, fibrosis, and cancer.

Detailed characterization of the cell types in the human brain requires scalable experimental approaches to examine multiple aspects of the molecular state of individual cells, and computational integration of the data to produce unified cell-state annotations. Here we report improved high-throughput methods for single-nucleus Droplet-based sequencing (snDrop-seq) and single-cell transposome hypersensitive-site sequencing (scTHS-seq). We used each method to acquire nuclear transcriptomic and DNA accessibility maps for >60,000 single cells from the human adult visual cortex, frontal cortex, and cerebellum. Integration of these data revealed regulatory elements and transcription factors that underlie cell-type distinctions, providing a basis for studying complex processes in the brain, such as genetic programs coordinating adult remyelination. We also mapped disease-associated risk variants to specific cellular populations, providing insights into normal and pathogenic cellular processes in the human brain. This integrative multi-omics approach permits more detailed single-cell interrogation of complex organs and tissues.

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