A single-cell RNA-seq survey of the action profile of general anaesthetics on the human prefrontal cortex cells

The cellular and molecular actions of general anaesthetics to induce anaesthesia state and also cellular signalling changes for subsequent potential “long term” effects remain largely elusive although great efforts have been made to study these in vitro, ex vivo and in vivo settings. General anaesthetics were reported to act on voltage-gated ion channels and ligand-gated ion channels. Here we used single-cell RNA-sequencing complemented with whole-cell patch clamp and calcium transient techniques to examine the gene transcriptome and ion channels profiling of sevoflurane and propofol, both commonly used clinically, on human embryonic primary prefrontal cortex (PFC) mixed cell cultures. Both propofol and sevoflurane at clinically relevant dose/concentration promoted “microgliosis” but only sevoflurane changed microglia cell similarity. Propofol and sevoflurane each extensively but transiently altered transcriptome profiling 2 hours after anaesthetics exposure across microglia, excitatory neurons, interneurons, astrocytes and oligodendrocyte progenitor cells. Within the excitatory neurons and microglia, exemplary ion-gated and ligand-gated ion channels related genes response to either anaesthetic included SCN1A, CACNAB2, KCNA1, GABRR2 and GRINA1 amongst many others. Utilising scRNA-seq as a robust and high-throughput tool, our work may provide a comprehensive blueprint for future mechanistic studies of general anaesthetics in clinically relevant settings. Single-cell RNA-sequencing complemented with whole-cell patch clamp and calcium transient techniques to examine the gene transcriptome and ion channels profiling of sevoflurane and propofol, both commonly used clinically, on human embryonic primary prefrontal cortex (PFC) mixed cell cultures.

尽管学界已在体外、离体及体内实验模型中对全身麻醉剂（general anaesthetics）展开了大量研究，但其诱导麻醉状态的细胞与分子机制，以及后续潜在"long term"效应相关的细胞信号传导变化，在很大程度上仍未明确。已有研究表明，全身麻醉剂可作用于电压门控离子通道（voltage-gated ion channels）与配体门控离子通道（ligand-gated ion channels）。本研究采用单细胞RNA测序（single-cell RNA-sequencing）联合全细胞膜片钳（whole-cell patch clamp）与钙瞬变（calcium transient）技术，针对临床常用的七氟烷（sevoflurane）与丙泊酚（propofol），在人类胚胎原代前额叶皮层（PFC）混合细胞培养体系中，对其基因转录组与离子通道谱进行分析。在临床相关剂量/浓度下，丙泊酚与七氟烷均可诱导"microgliosis"（小胶质细胞活化），但仅七氟烷会改变小胶质细胞的细胞相似性。暴露于麻醉剂2小时后，丙泊酚与七氟烷均会在小胶质细胞、兴奋性神经元（excitatory neurons）、中间神经元（interneurons）、星形胶质细胞（astrocytes）及少突胶质前体细胞（oligodendrocyte progenitor cells）中，广泛且一过性地改变转录组谱特征。在兴奋性神经元与小胶质细胞中，响应两类麻醉剂的典型离子门控与配体门控离子通道相关基因包括SCN1A、CACNAB2、KCNA1、GABRR2及GRINA1等众多基因。本研究以单细胞RNA测序作为可靠的高通量研究工具，其成果可为未来在临床相关场景下开展全身麻醉剂的机制研究提供全面的研究框架。本研究采用单细胞RNA测序联合全细胞膜片钳与钙瞬变技术，针对临床常用的七氟烷与丙泊酚，在人类胚胎原代前额叶皮层混合细胞培养体系中分析其基因转录组与离子通道谱。