Glaucine suppresses IL-16/CD4 to alleviate neuropathic pain via spinal p56-NMDA receptor signaling.

Abstract

Effective drugs for neuropathic pain are urgently needed. Natural compound glaucine from Corydalis Rhizoma inhibits spinal sensitization and alleviates neuropathic pain, yet its exact analgesic mechanism remains unclear. A rat model of neuropathic pain was established via chronic constriction injury (CCI) of the sciatic nerve. Behavioral tests (von Frey and Hargreaves tests) were used to assess neuropathic pain symptoms by evaluating mechanical allodynia and thermal hyperalgesia. Primary spinal neurons, glia, and peritoneal macrophages were cultured for in vitro analysis. Molecular pathways were examined using RNA sequencing, Western blot, qRT-PCR, and immunofluorescence. Glaucine significantly suppressed spinal IL-16 expression in CCI rats, and its analgesic effects were reversed by exogenous IL-16 administration. RNA-seq and protein assays confirmed IL-16 upregulation in injured peripheral nerves and spinal cord. Critically, inhibiting IL-16/CD4 signaling at peripheral and spinal levels alleviated neuropathic pain. Dual immunofluorescence in vivo and in vitro revealed IL-16/CD4 localization in neurons, and spinal IL-16 inhibition reduced neuronal activation. Calcium fluorescence imaging further demonstrated that spinal IL-16 blockade reduced spinal neuronal excitability through restraining the NMDA receptor. The IL-16 induced phosphorylation of NMDA receptor was mediated by lymphocyte-specific protein tyrosine kinase (p56). Additionally, peripheral macrophages were identified as a source of IL-16 after nerve injury. Glaucine alleviates neuropathic pain by inhibiting the IL-16-p56-NMDA axis which promotes pain through enhanced NMDA activity and central sensitization. The targeted inhibition highlights glaucine as a novel therapeutic strategy for alleviating neuropathic pain.