Drp1/HK1/NLRP3-Associated Pathway Contributes to Neuronal Pyroptosis-Related Signaling in Neuropathic Pain.

Neuropathic pain (NP) is a prevalent and debilitating chronic pain condition within the nervous system. Recent studies have implicated pyroptosis as a potential mechanism underlying NP. Therefore, this study investigates NLRP3-mediated neuronal pyroptosis and its regulation in NP. A spared nerve injury (SNI) model was established in C57BL/6 mice to induce NP. Mechanical allodynia and thermal hyperalgesia were assessed using von Frey filaments and the hot plate test. Nissl staining was performed on spinal cord sections to evaluate neuronal morphology. Ipsilateral lumbar spinal cord tissues were collected for Western blot analysis, while transverse spinal cord sections were used for immunofluorescence and transmission electron microscopy to assess dorsal horn localization and cellular morphology. The functional roles of HK1 and Drp1 were examined through intrathecal delivery of HK1-overexpressing adeno-associated virus (AAV-HK1) and Drp1-targeted siRNA. SNI induced persistent pain hypersensitivity and dorsal horn neuronal morphological alterations. This was accompanied by a time-dependent upregulation of NLRP3, ASC, GSDMD-N, and IL-1β post-SNI, with immunofluorescence evidence supporting prominent neuronal localization of pyroptosis-related signals. Concurrently, HK1 expression was progressively downregulated, while Drp1 was upregulated in neurons. Restoration of HK1 via intrathecal AAV delivery suppressed pyroptosis and alleviated pain hypersensitivity. Conversely, Drp1 knockdown restored HK1 levels, reduced pyroptosis markers, and alleviated pain hypersensitivity. Mitochondrial fractionation revealed reduced mitochondrial HK1 and increased mitochondrial Drp1 following SNI. Drp1 upregulation is functionally associated with HK1 downregulation and NLRP3/GSDMD-N-related neuronal pyroptosis in SNI-induced NP. The Drp1/HK1/NLRP3-associated pathway may represent a potential regulatory mechanism and therapeutic target for neuropathic pain.