Journal of Inflammation Research (Sep 2020)
Spinal TLR4/P2X7 Receptor-Dependent NLRP3 Inflammasome Activation Contributes to the Development of Tolerance to Morphine-Induced Antinociception
Abstract
Haiyan Wang1 ,* Yu Zhang1 ,* Xiaqing Ma,1 Wenying Wang,1 Xiaotao Xu,1 Min Huang,1 Liang Xu,2 Haibo Shi,3 Tifei Yuan,4 Wei Jiang,1 Aizhong Wang,1 Tao Xu1,5 1Department of Anesthesiology, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai 200233, People’s Republic of China; 2Heart Health Center, East Hospital, Tongji University School of Medicine, Shanghai 200120, People’s Republic of China; 3Shanghai Key Laboratory of Sleep Disordered Breathing, Shanghai 200233, People’s Republic of China; 4Shanghai Key Laboratory of Psychotic Disorders, Shanghai Mental Health Center, Shanghai Jiao Tong University School of Medicine, Shanghai 200030, People’s Republic of China; 5Department of Anesthesiology, Tongzhou People’s Hospital, Nantong 226300, People’s Republic of China*These authors contributed equally to this work.Correspondence: Tao XuDepartment of Anesthesiology, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai 200233, People’s Republic of ChinaTel +86 21 24058309Fax +86 21 24058330Email [email protected] WangDepartment of Anesthesiology, Shanghai Jiao Tong University Affiliated Sixth People’s Hospital, Shanghai 200233, People’s Republic of ChinaEmail [email protected]: Long-term use of morphine induces antinociceptive tolerance and limits its clinical efficacy. Neuroinflammation in the spinal cord is thought to play a pivotal role in the development of morphine tolerance. Toll-like receptor 4 (TLR4) and P2X7 receptor (P2X7R) are key modulators of neuroinflammation. Recent studies show that the Nod-like receptor protein 3 (NLRP3) inflammasome play a crucial role in microglia-mediated neuroinflammation. Thus far, the mechanism underlying NLRP3 inflammasome activation during morphine-induced tolerance is not yet fully understood. Therefore, we sought to investigate the mechanisms of NLRP3 inflammasome activation and its role in the development of morphine-induced tolerance.Methods: Repeated morphine treatment through intrathecal injection (15 μg once daily for 7 days) was given to establish antinociceptive tolerance in mice. Tail-flick latency was used to evaluate morphine-induced antinociception. NLRP3 knockout mice were used to assess the role of NLRP3 inflammasome in morphine tolerance. TLR4 knockout mice and A438079, a P2X7R antagonist, were used to assess the role of TLR4 and P2X7R in chronic morphine-induced NLRP3 inflammasome activation. Western blot and immunofluorescence were used for quantitative comparison.Results: Repeated morphine treatment increased the expression of NLRP3. Knockout of NLRP3 attenuated morphine-induced tolerance and suppressed morphine-induced activation of microglia. Knockout of TLR4 alleviated morphine tolerance and chronic morphine-induced upregulation of spinal NLRP3. Inhibition of spinal P2X7R with A438079 not only prevented the development of morphine-induced tolerance but also inhibited repeated morphine treatment-induced upregulation of spinal NLRP3. Furthermore, spinal NLRP3, TLR4 and P2X7R were collectively colocalized with the microglia marker Iba1.Conclusion: This study demonstrates that the NLRP3 inflammasome in microglia plays a crucial role in morphine tolerance and that both TLR4- and P2X7R-dependent pathways are required for NLRP3 inflammasome activation over the course of the development of morphine-induced tolerance. Our results provide a new perspective for the targeted treatment of morphine-induced tolerance.Keywords: morphine, tolerance, NLRP3, TLR4, P2X7R, neuroinflammation
