Correlating anatomy and function with gene expression in individual neurons by combining in vivo labeling, patch clamp and single cell RNA-seq

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 we 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. With this method we can determine the genetic expression pattern of functionally and anatomically identified individual neurons. single cortical neurons were patch clamped and the RNA harvested; single neuron mRNA profiles were generated by deep sequencing将神经元划分为不同类型是一项持续推进的研究工作，旨在揭示并阐明神经系统组成成分的多样性。近年来可采用的研究方法使得我们能够检测哺乳动物大脑皮层（cerebral cortex）中单个神经元的基因表达模式，进而构建出有效的分类方案。若要全面深入理解神经元的多样性，需将此类基因分类方案与传统分类参数（如位置、轴突投射或感觉刺激响应特性）相结合。本文介绍了一种可将单个神经元的基因表达与其位置、轴突投射或感觉响应特性相关联的方法：先根据神经元的解剖学或功能特性在活体内对其进行标记，随后使用膜片钳电极（patch clamp pipettes）在体外单独收集其RNA，用于RNA测序（RNA-seq）。通过该方法，我们能够确定经功能与解剖学鉴定的单个神经元的基因表达模式。本研究对单个皮层神经元实施了膜片钳（patch clamp）操作并收集其RNA；通过深度测序（deep sequencing）生成了单个神经元的mRNA表达谱。