Single-Nucleus Transcriptomics Reveals Glial Metabolic-Immune Rewiring and Intercellular Signaling Disruption in...

Chronic migraine (CM) is a debilitating neurological disorder, yet the glial-specific mechanisms underlying its pathophysiology in the trigeminal nucleus caudalis (TNC)-a critical hub for craniofacial pain processing-remain poorly understood. Here, we employed single-nucleus RNA sequencing (snRNA-seq) to resolve cell-type-specific transcriptional landscapes in a nitroglycerin (NTG)-induced CM rat model, with a particular focus on microglia and astrocytes. We identified 19 transcriptional clusters representing nine major cell types, among which reactive microglia (NTG-Mic) and astrocytes (NTG-Asts) were markedly expanded. The NTG-Mic displayed a glycolysis-dominant, complement-enriched state, whereas the NTG-Asts exhibited concurrent activation of amino acid transport and cytokine signaling pathways. Pseudotime trajectory analysis revealed bifurcated glial activation paths, with NTG driving both cell types toward terminal reactive states. Intercellular communication inference uncovered suppressed homeostatic interactions (e.g., CSF1-CSF1R) alongside enhanced proinflammatory signaling (e.g., FGF1-FGFR2, PTN-SDC4), particularly affecting neuron-glia and glia-glia crosstalk. Together, these findings define a high-resolution atlas of glial reprogramming in CM, implicating state-specific metabolic-immune transitions and dysregulated glial communication as potential targets for therapeutic intervention.