P2X7 on mast cells participates in peripheral pain and serves as a potential target for salicylic acid and aspirin analgesia

Background: Extracellular ATP signaling through excitatory and calcium-permeable P2X receptor channels is considered as a critical player in pain generation and maintenance. P2X7 has attracted much attention over the past decade because of its prominent role in driving inflammatory processes. The role of P2X7 in mast cells in peripheral pain remains unclear. Methods: P2X expression in mouse peritoneal mast cells was detected by RT-PCR. The subtypes of P2X receptors in mouse peritoneal mast cells were determined with a series of blockers by using calcium imaging and electrophysiology. The regulation of inflammatory factors mediated by different P2X subtypes were detected by ELISA and real-time PCR. The role of mast cells and P2X7 receptor in peripheral pain was explored by behavioral assays, pathological analysis and real-time PCR. Several anti-inflammatory small molecules were screened based on P2X7 in mast cells by using calcium imaging, electrophysiology and molecular docking. Results: We found that ATP was significantly increased in inflammatory pain. Mouse peritoneal mast cells expressed P2X1, P2X3, P2X4 and P2X7 and could be activated by different concentrations of extracellular ATP, which could be blocked by specific ion channel antagonists. In particularly, high concentration of ATP could induce mast cells to release inflammatory mediators such as histamine, IL-1β and CCL3 through P2X7 receptor. Furthermore, peripheral pain induced by high concentration of ATP could be alleviated by P2X7 blockers or mast cell defects. Interestingly, salicylic acid and aspirin could attenuate the inward current, the release of inflammatory factors and peripheral pain induced by ATP with high concentration. Furthermore, salicylic acid and aspirin also inhibited the inward current evoked by P2X7 agonist BZATP. Molecular docking results showed that salicylic acid and aspirin had affinity to the cytoplasmic GDP-binding region of P2X7. Conclusions: We concluded that P2X7 on mast cells involved in peripheral pain. Salicylic acid and aspirin could inhibit the activity of P2X7 via interacting with the GDP binding region. P2X7 receptor was a potential target for salicylic acid and aspirin analgesia.