Acid-induced prolonged antinociceptive signaling in muscle. The withdrawal responses of mouse hind paws to a 0.2-mN bending force in mice before and after intramuscular acid injection in the gastrocnemius muscle. (A) In the dual acid injection scheme, mice showed transient hyperalgesia after the first intramuscular acid injection and chronic hyperalgesia after a second acid injection spaced 5 days apart. (B) A mix of acid saline with APETx2 and capsazepine abolished the transient hyperalgesia with the first injection and prevented the development of chronic hyperalgesia induced by the second acid injection 5 days later. (C,D) With the first acid injection, acid induced a prolonged antinociceptive effect lasting for 2 days, with ASIC3 and TRPV1 blocked by APETx2 and capsazepine, respectively, and a second acid injection at day 2 could not induce any hyperalgesic effect. Black arrows indicate when mice received the intramuscular acid injection. Red arrows indicate when mice received the co-injection of acid with APETx2 and capsazepine. B, baseline on day 0; D, day; WT, wild-type mice; CZP, capsazepine. *P < 0.05 compared with the response at baseline.