Single-Cell Transcriptomic Analysis of Somatosensory Neurons Uncovers Temporal Development of Neuropathic Pain

Peripheral nerve injury could lead to chronic neuropathic pain. Understanding transcriptional changes induced by nerve injury could provide fundamental insights into the complex pathogenesis of neuropathic pain. Gene expression profiles of dorsal root ganglia (DRG) under neuropathic pain condition have been studied. However, little is known about transcriptomic changes in individual DRG neurons after peripheral nerve injury. Here we performed single-cell RNA sequencing on dissociated mouse DRG cells after spared nerve injury (SNI). In addition to DRG neuron types also found under normal conditions, we identified three SNI-induced neuron clusters (SNIICs) characterized by the expression of Atf3/Gfra3/Gal (SNIIC1), Atf3/Mrgprd (SNIIC2) and Atf3/S100b/Gal (SNIIC3). These SNIICs were originated from Cldn9+/Gal+, Mrgprd+ and Trappc3l+ DRG neuron types. Interestingly, SNIIC2 was switched to SNIIC1 by increasing Gal and reducing Mrgprd expression 2 days after nerve injury. Inferring the gene regulatory networks underlying nerve injury, we revealed that activated transcription factor Atf3 and Egr1 in SNIICs could enhance Gal expression while activated Cpeb1 in SNIIC2 might suppress Mrgprd expression within 2 days after SNI. Furthermore, we screened the transcriptomic changes in the development of neuropathic pain to identify the potential analgesic targets. We revealed that the expression of cardiotrophin-like cytokine factor 1, which could activate the astrocytes in the dorsal horn of spinal cord, was increased in SNIIC1 neurons and contributed to SNI-induced mechanical allodynia. Therefore, our results provide a new framework to understand the changes in neuron types and the dynamics of molecular and cellular mechanisms underlying the development of neuropathic pain. Overall design: Dorsal root ganglion cells profile of C57 mice in Control, SNI 6h, SNI 24h, SNI 2d, SNI 7d, SNI 14d, and SNI 28d. Please note that The "barcode_of_raw_UMI_counts_1.txt" is the barcode list for each cell in "raw_UMI_counts_1.txt" The "barcode_of_raw_UMI_counts_2.txt" is the barcode list for each cell in "raw_UMI_counts_2.txt" A part of processed data in "Smart-seq2_FPKM_metadata.txt" and "Smart-seq2_FPKM.txt" was obtained from GSE63576 and listed in "smart-seq2_data_from GSE63576.txt".