CFA-induced chronic pain and Von Frey hair test

Chronic inflammatory pain was induced by CFA intraplantar injection. Briefly, each B6 mouse was injected with either saline or CFA (20 μl) in the footpad of the right hind paw under isoflurane anesthesia on post-inoculation day (PID) 0. From PID14 to 18, saline- and CFA-injected mice were twice daily administered s.c. with vehicle, morphine (10 mg/kg), convallatoxin (0.5 mg/kg), or morphine (10 mg/kg) and convallatoxin (0.5 mg/kg) combined. Mechanical allodynia was subsequently evaluated using von Frey filaments (range 0.1–1 g; IITC Life Science). Mice were placed on a mesh floor with 5 × 5 mm holes, covered with a cup to prevent visual stimulation, and allowed to adapt for 1 h prior to testing. On the test days, withdrawal responses following hind paw stimulation were measured, and mechanical allodynia was defined as changes in the amount of pressure required to induce withdrawal. The filaments were applied to the middle of the plantar surface of the hind paw.