Researchers combine in vivo labeling, patch clamp, and single cell RNA-seq to correlate anatomy and function with gene expression in individual neurons

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The classification of neurons into distinct types is an ongoing effort aimed at revealing and understanding the diversity of the components of the nervous system. Recently available methods allow us to determine the gene expression pattern of individual neurons in the mammalian cerebral cortex to generate powerful categorization schemes. For a thorough understanding of neuronal diversity such genetic categorization schemes need to be combined with traditional classification parameters like position, axonal projection or response properties to sensory stimulation.

Here UCSD researchers describe a method to link the gene expression of individual neurons with their position, axonal projection, or sensory response properties. Neurons are labeled in vivo based on their anatomical or functional properties and, using patch clamp pipettes, their RNA individually harvested in vitro for RNAseq. The researchers validate the methodology using multiple established molecularly and anatomically distinct cell populations and explore molecular differences between uncharacterized neurons in mouse visual cortex. Gene expression patterns between L5 neurons projecting to frontal or contralateral cortex are distinct while L2 neurons differing in position, projection, or function are molecularly similar. With this method we can determine the genetic expression pattern of functionally and anatomically identified individual neurons.

 Single cell RNA-seq from patched neurons distinguishes
spatially distributed glutamatergic cells

rna-seq

Neurons from spatially separate vertical layers of mouse visual cortex (L2, as defined by the SepW1-Cre line; L6, defined as cells of deeper L6 at the border to the white matter) and CA1 pyramidal cells from dorsal (CA1d) and ventral (CA1v) poles of the mouse hippocampus were patch-clamped and the interior of the cell sucked into the tip of the patch pipette and subsequently expelled into a small volume tube. CDNA libraries of individual cells were generated and sequenced using high-throughput sequencing.


Pfeffer CK, Beltramo R. (2017) Correlating Anatomy and Function with Gene Expression in Individual Neurons by Combining in Vivo Labeling, Patch Clamp, and Single Cell RNA-seq. Front Cell Neurosci 11:376. [article]

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