Single-nucleus RNA sequencing of rostral ventromedial medulla in mice with trigeminal neuralgia.

Aim: To investigate the transcriptional changes and cell interactions following trigeminal neuralgia (TN) in different cell types in rostral ventromedial medulla (RVM).Methods: In this study, trigeminal neuropathic pain was induced in mice by ligating the left infraorbital nerve. Ten days after nerve ligation, we performed single-nucleus RNA sequencing of the RVM cells from the Sham and TN groups.Results: We identified neurons, astrocytes, microglial cells, oligodendrocytes, oligodendrocyte progenitor cells, Purkinje cells, neuroblasts, endothelial cells and fibroblasts in RVM tissue. After analyzing the cell-type-specific transcriptional changes in the RVM following nerve ligation, we found that the number of neurons and Purkinje cells in the RVM increased. Furthermore, the differentially expressed genes (DEGs) in neurons and astrocytes between the two groups was identified. The downregulated DEGs in neurons were significantly enriched in GABAergic synapse (such as Gabrg3 and Gabra2). The upregulated DEGs in neurons were enriched in glutamatergic synapse and voltage-gated ion channels. The downregulated DEGs in astrocytes involved in regulation of the postsynaptic membrane and synaptic membrane were enriched. Notably, the CellChat analysis highlights the Slit2-Robo 1/2 signaling pathway as a key route of communication between neurons and astrocytes after nerve ligation.Conclusions: This study analyzed cell-type-specific transcriptional responses to pain, and identified the possible key genes involved in TN. We inferred cell-state-specific communication in various cell types. Our findings provide potential targets, such as Slit2-Robo 1/2, Gabrg3, Gabra2, Grm4, Grik2, Cadps2 and Camk4 on therapeutic strategies for neuropathic or orofacial pain.