scRNAseq analysis of mouse WT and Fmr1KO dorsal root ganglions

Sensory hypersensitivity and somatosensory deficits represent the core symptoms of Fragile X syndrome (FXS).These alterations are believed to arise from changes in cortical sensory processing, while potential deficits in the function of peripheral sensory neurons or surrounding satellite glial cells (SGCs) residing in dorsal root ganglia remain unexplored. We found that cultured peripheral sensory neurons exhibit pronounced hyperexcitability in Fmr1 KO mice evident in markedly increased firing rate and decreased spike threshold. These alterations were caused primarily by increased input resistance, which aroused from decreased HCN channel-mediated current Ih. EM analyses also revealed major structural defects in neuron-SGC association. Single-cell RNAseq demonstrated extensive transcriptional changes in both neurons and SGCs indicative of defects in neuronal maturation/differentiation and neuron-SGC communication. These results reveal a hyper-excitable state of peripheral sensory neurons in Fmr1 KO mice with contributions from intrinsic alterations and from disrupted neuron-glia association and communication. We have applied the high-throughput single-cell mRNA sequencing technique, using the Chromium Single Cell Gene Expression Solution (10x Genomics) to mouse DRG, 2 biological replicates for each sample.感觉超敏与躯体感觉缺陷是脆性X综合征（Fragile X syndrome, FXS）的核心症状。目前学界普遍认为此类异常源于皮层感觉加工过程的改变，但背根神经节（dorsal root ganglia, DRG）内外周感觉神经元及其周围卫星胶质细胞（satellite glial cells, SGCs）的功能潜在缺陷仍未得到系统探索。本研究发现，Fmr1基因敲除（Fmr1 KO）小鼠的体外培养外周感觉神经元表现出显著的兴奋性增高，具体体现为放电频率显著升高与动作电位阈值降低。此类异常主要由输入电阻升高所致，而该变化源于超极化激活环核苷酸门控通道（HCN channel）介导的内向电流Ih的活性减弱。电子显微镜分析还揭示了神经元与卫星胶质细胞的联结存在显著结构缺陷。单细胞RNA测序结果显示，神经元与卫星胶质细胞均出现广泛的转录组改变，提示神经元成熟、分化以及神经元-胶质细胞通信过程均存在缺陷。上述结果表明，Fmr1基因敲除小鼠的外周感觉神经元处于过度兴奋状态，该状态由神经元内在改变以及神经元-胶质细胞联结与通信紊乱共同介导。本研究采用高通量单细胞mRNA测序技术，借助Chromium单细胞基因表达解决方案（10x Genomics）对小鼠背根神经节进行测序，每个样本设置2个生物学重复。