Electroacupuncture Relieves Neuropathic Pain by Inhibiting Degradation of the Ecto-Nucleotidase PAP in the Dorsal Root Ganglions of CCI Mice

Abstract

Electroacupuncture (EA) is an effective treatment for relieving neuropathic pain, however, its underlying effect mechanisms remain unclear. We hypothesised that EA alleviates pain by inhibiting degradation of the ecto-nucleotidase prostatic acid phosphatase (PAP) and facilitating ATP dephosphorylation in dorsal root ganglions (DRGs). To test this, we applied EA in male C57 mice subjected to chronic constriction injury (CCI) and assessed extracellular ATP and 5′-nucleotidease expression in DRGs. Specifically, we used a luminescence assay, quantitative reverse transcriptase–polymerase chain reaction, western blotting, immunohistochemistry and nociceptive-related behavioural changes to gather data, and we tested for effects after PAP expression was inhibited with an adeno-associated virus (AAV). Moreover, membrane PAP degradation was investigated in cultured DRG neurons and the inhibitory effects of EA on this degradation were assessed using immunoprecipitation. EA treatment alleviated CCI surgery induced mechanical allodynia. Furthermore, extracellular ATP decreased significantly in both the DRGs and dorsal horn of EA-treated mice. PAP protein but not mRNA increased in L4–L6 DRGs, and inhibition of PAP expression via AAV microinjection reversed the analgesic effect of EA. Membrane PAP degradation occurred through a clathrin-mediated endocytosis pathway in cultured DRG neurons; EA treatment inhibited the phosphorylation of adaptor protein complex 2, which subsequently reduced the endocytosis of membrane PAP. In summary, EA treatment alleviated peripheral nerve injury-induced tactile allodynia in mice by inhibiting membrane PAP degradation via reduced endocytosis and subsequently promote ATP dephosphorylation in DRGs. Our findings provide new insights into the underlying mechanisms of pain relief provided by acupuncture. Funding Statement: This study was supported by grants from the National Natural Science Foundation of China 414 (Nos. 81874371, 81803853, 81771133, 81970995), Shanghai Shenkang Hospital Development Center Founding (SHDC12017X11), and Shanghai municipal Education Commission-Gaofeng Clinical Medicine Support [20191903], Shanghai Municipal Key Clinical Specialty (shslczdzk03601)，State Key Laboratoy of Neuroscience(SKLN-201803). Declaration of Interests: The authors have declared that no competing interests exist regarding the publication of this paper. Ethics Approval Statement: All of the experiments were approved by the Animal Care and Use Committee of Renji Hospital, Shanghai Jiao Tong University School of Medicine, and all protocols followed the policies issued by the National Institutes of Health and the International Association for the Study of Pain.