Acupotomy Alleviates Pain and Attenuates Spinal Neuronal Hyperexcitability in Knee Osteoarthritis Rats: Implication of the CaMKII/GluA1 Axis

Meng Wu,1,* Jinyan Teng,2,* Ziyi Huang,1 Ruizhe Xu,1 Xiaoming Gao,1 Xin Hao,3 Zhigang Li,1 Zhaoheng Liu41School of Acupuncture, Moxibustion and Tuina, Beijing University of Chinese Medicine, Beijing, People’s Republic of China; 2Department of Acupuncture and Moxibustion, Hubei Provincial Hospital of Traditional Chinese Medicine, Wuhan, Hubei, People’s Republic of China; 3Department of Acupuncture and Physiotherapy, Beijing Electric Power Hospital of State Grid Corporation of China, Beijing, People’s Republic of China; 4School of Life Sciences, Beijing University of Chinese Medicine, Beijing, People’s Republic of China*These authors contributed equally to this workCorrespondence: Zhigang Li; Zhaoheng Liu, Email [email protected]; [email protected]: To investigate the spinal mechanisms underlying the analgesic effects of acupotomy on chronic pain and central sensitization in rats with monosodium iodoacetate (MIA)-induced knee osteoarthritis (KOA).Methods: Following MIA-induced KOA and a three-week acupotomy intervention in rats, pain behaviors and joint pathology were assessed. Spinal RNA-sequencing was performed to identify differentially expressed genes. Key targets (CaMKII and GluA1) were validated at both mRNA and protein levels using RT-qPCR and Western blotting. Furthermore, immunofluorescence and whole-cell patch-clamp electrophysiology were employed to evaluate the activation and excitability of spinal substantia gelatinosa (SG) neurons.Results: Acupotomy significantly alleviated mechanical and thermal hyperalgesia and reduced joint degeneration in KOA rats. RNA-seq analysis revealed the modulation of 220 genes in the spinal cord. These DEGs were significantly enriched in calcium signaling and synaptic transmission pathways, highlighting CaMKII and Gria1 as potential key molecules. Molecular validations confirmed that acupotomy downregulated CaMKII and Gria1 mRNA expression and the phosphorylation ratios of CaMKII and GluA1. Furthermore, acupotomy reduced the activation of p-CaMKII-positive neurons in the spinal SG region and dampened their abnormally high-frequency action potential discharges.Conclusion: Acupotomy intervention ameliorates peripheral joint degeneration and exerts significant analgesia in KOA, which is accompanied by a reduction in the intrinsic hyperexcitability of spinal SG neurons. Although direct causal validation utilizing targeted inhibitors is warranted, our findings suggest that this analgesic effect is closely associated with the downregulation of the CaMKII/GluA1 signaling axis, providing a plausible central mechanistic framework.Keywords: acupotomy, knee osteoarthritis, RNA-seq, spinal dorsal horn, electrophysiology