Study: A transcriptomic taxonomy of adult mouse visual cortex (VISp) 1679 cells

Tasic B, Menon V, Nguyen TN, Kim TK, Jarsky T, et al. (2016) Adult mouse cortical cell taxonomy revealed by single cell transcriptomics. Nat Neurosci 19: 335-346. DOI: 10.1038/nn.4216

Nervous systems are composed of various cell types, but the extent of cell type diversity is poorly understood. Here, we construct a cellular taxonomy of one cortical region, primary visual cortex, in adult mice based on single cell RNA-sequencing. We identify 49 transcriptomic cell types including 23 GABAergic, 19 glutamatergic and seven non-neuronal types. We also analyze cell-type specific mRNA processing and characterize genetic access to these transcriptomic types by many transgenic Cre lines. Finally, we show that some of our transcriptomic cell types display specific and differential electrophysiological and axon projection properties, thereby confirming that the single cell transcriptomic signatures can be associated with specific cellular properties.

This study utilizes 1,679 cells collected from the primary visual cortex of 56 day old adult male mice.

Below: Workflow overview. (a) Experimental workflow starts with the isolation, sectioning and micro-dissection of the primary visual cortex from a transgenic mouse. The tissue samples are converted into a single-cell suspension, single cells are isolated by FACS, poly(A)-RNA from each cell is reverse transcribed (RT), cDNA is amplified and fragmented, and sequenced on a next-generation sequencing (NGS) platform. (b) Analysis workflow starts with the definition of high variance genes and iterative clustering based on two different methods, PCA (shown here) and WGCNA, and cluster membership validation using a random forest classifier. Cells that are classified consistently into one cluster are called ‘core’ cells, while cells that are mapped to more than one cluster are labeled ‘intermediate cells.’ After the termination criteria are met, clusters from the two methods are intersected, and iteratively validated until all core clusters contain at least 4 cells. (Supplementary Fig. 3, Methods). (c) The final 49 clusters were assigned an identity based on cell location (Fig. 2) and marker genes (Fig. 3). Each type is represented by a color bar with the name and number of core cells representing that type. The violin plots represent distribution of mRNA expression on a linear scale, adjusted for each gene (max. RPKM on the right), for major known marker genes: Snap25 (pan-neuronal); Gad1 (pan-GABAergic); Vip, Sst and Pvalb (GABAergic); Slc17a7 (pan-glutamatergic); Rorb (mostly L4 and L5a); Foxp2 (L6); Aqp4 (astrocytes); Pdgfra (oligodendrocyte precursor cells, OPCs); Mog (oligodendrocytes); Itgam (microglia); Flt1 (endothelial cells) and Bgn (smooth muscle cells, SMC).

Methodology overview figure

 

More details in: Tasic, et al. Nature Neuroscience (2016).