α7 nAChR PAMs attenuate CCD‑induced neuropathic pain, with modulation of the NLRP3/IL‑1β pathway: insights from single‑cell transcriptomics and CHRNA7 structural simulation

Neuropathic pain (NP) remains a refractory clinical condition with unclear genetic and molecular mechanisms. The cholinergic system and NLRP3 pathway are critically involved in pain regulation, but their roles in chronic dorsal root ganglion compression (CCD)-induced radicular pain remain to be fully elucidated. This study aims to investigate the analgesic effects and underlying mechanisms of α7 nAChR positive allosteric modulators (PAMs) in NP. Single-cell transcriptomic datasets from mouse dorsal root ganglion (DRG) were combined with integrated supervised machine-learning to screen core molecular signatures associated with pain. A CCD rat model and LPS-stimulated DRG cells were established. Pain behaviors were assessed by von Frey and hot-plate tests. Protein expression was measured by western blotting, immunofluorescence, and ELISA. Apoptosis was evaluated by flow cytometry. Protein stability was analyzed by AlphaFold3 and molecular dynamics simulation. Single-cell transcriptomic analysis revealed that CHRNA7 and NLRP3/IL-1β pathway-related genes were expressed in overlapping cell types, including satellite glial cells. Integrated machine‑learning further confirmed that these molecules constitute core molecular signatures distinguishing pain‑associated cellular states. PNU-120596 and NS-1738 significantly alleviated mechanical and thermal hypersensitivity in CCD rats, an effect blocked by the α7 nAChR antagonist methyllycaconitine (MLA). Both PAMs modulated the NLRP3/IL-1β pathway in the ACC and DRG, accompanied by changes in CHRNA7 protein levels. In LPS-induced DRG cells, PNU-120596 and NS-1738 similarly modulated the NLRP3/IL-1β pathway and reduced apoptosis effects reversed by MLA. Molecular dynamics simulations showed that the CHRNA7 W77G mutation leads to structural instability and conformational loosening. This study identifies cell-type-level expression signatures of CHRNA7 and the NLRP3/IL-1β pathway in mouse DRG cell subpopulations. α7 nAChR PAMs produce prominent analgesia against CCD-provoked neuropathic pain, concomitant with modulation of the NLRP3/IL-1β pathway. Our findings provide transcriptomic insights and potential therapeutic targets for neuropathic pain.