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Chronic migraine is a disabling neurological disorder with complex mechanisms. The trigeminal nucleus caudalis (TNC) is a critical relay in migraine pathogenesis, yet its cellular and molecular underpinnings remain unclear. We applied single-nucleus RNA sequencing (snRNA-seq) to the TNC of a nitroglycerin (NTG) -induced chronic migraine mouse model and controls. Major neuronal classes were annotated, followed by subclustering of GABAergic neurons. We assessed cell-type abundance changes, reconstructed transcriptional trajectories, and applied network- and pathway-level analyses including high-dimensional weighted gene co-expression network analysis (hdWGCNA), pseudotime modeling, and CellChat-based intercellular communication profiling. We identified a previously unrecognized GABAergic subpopulation characterized by high Nox4 expression (GABANox4). This subpopulation was significantly reduced in NTG-treated mice compared with controls (p = 0. 018), a finding further validated by immunofluorescence staining. hdWGCNA identified a migraine-associated co-expression module enriched in GABANox4 neurons, which was further supported by disease enrichment analysis. Pseudotime analysis showed that GABANox4 neurons diverged into distinct transcriptional states under NTG treatment. Intercellular communication analysis revealed enhanced crosstalk of GABANox4 neurons with astrocytes, endothelial cells, and OPCs in the NTG group, mediated by NTG-enriched ligand–receptor pairs such as Agrn–Dag1, Ncam1–Ncam2, and endothelial-derived Ncam1–L1cam, whereas VEH-specific interactions such as Pdgfa–Pdgfra and Pdgfa–Pdgfrb were diminished. Our integrative single-cell analysis identifies GABANox4 neurons as a vulnerable and communication-active GABAergic subpopulation in the TNC that is selectively reduced and transcriptionally reprogrammed in chronic migraine. The disruption of inhibitory signaling and rewiring of ligand–receptor networks highlight this population as a potential cellular driver of migraine pathophysiology and a candidate target for therapeutic intervention.
Xu et al. (Tue,) studied this question.