Inflammatory pain model in mice.
Inflammation was induced with intraplantar injection of 10 µl CFA (diluted 1:4 with saline; Sigma-Aldrich) to the left hind paw. The PWL to noxious heat was measured using a system described previously (Bai et al., 2010). The injected hind paw showed edema and erythema indicating inflammation (Bai et al., 2010).
Neuropathic pain model in rats.
The L5 spinal nerve of adult, male Sprague-Dawley rats (300–350 g; Harlan Bioproducts for Science) was ligated as described previously (Guan et al., 2008). The animals were anesthetized with isoflurane (2%; Abbott Laboratories). The left L5 spinal nerve was tightly ligated with a 6–0 silk suture and cut distally. The muscle layer was closed with 4–0 silk suture and the skin closed with metal clips.
Inflammatory pain model in rats.
Inflammation was induced with CFA suspended in an oil/saline 1:1 emulsion and 0.1 mg was injected s.c. (Mycobacterium) into the plantar surface of the left hind paw. The injection produced intense tissue inflammation of the hind paw characterized by erythema, edema, and hyperalgesia that were confined to the injected hind paw.
Pain measurement in rats.
Hypersensitivity to punctuate mechanical stimulation in rats was determined with the up-down method using a series of von Frey filaments (0.38, 0.57, 1.23, 1.83, 3.66, 5.93, 9.13, and 13.1 g) as described previously (Chaplan et al., 1994). The von Frey filaments were applied for 4–6 s to the test area between the footpads on the plantar surface of the hind paw. If a positive response occurred (e.g., abrupt paw withdrawal, licking, and shaking), the next smaller von Frey hair was used; if a negative response was observed, the next higher force was used. The test was continued until: (1) the responses to five stimuli were assessed after the first crossing of the withdrawal threshold or (2) the upper/lower end of the von Frey hair set was reached before a positive/negative response had been obtained. The PWT was determined according to the formula provided by Dixon (1980).
Measurement of thermal pain hypersensitivity in mice and rats.
Thermal pain hypersensitivity was determined by measuring PWL to radiant heat stimuli (Hargreaves et al., 1988) with a plantar stimulator analgesia meter (model 390; IITC). Animals were placed under individual plastic boxes on a heated glass floor (30°C) and allowed to habituate for at least 30 min before testing. Both hind paws were tested three times, with >2 min between trials. A cut-off time of 20 s was used to avoid sensitization and damage to the skin. The mean PWL of the three trials was used for data analysis.
Mice were i.p. injected with the vehicle or cannabinoids. After 15 min, the spontaneous locomotor activity of mice was measured in a standard home cage in a 12-station photobeam activity system (Opto-M3 Activity Meter; Columbus Instruments) where the animals were placed individually 30 min after injection of drugs. Using infrared beams, activity was monitored in the horizontal directions. The total number of ambulatory beam breaks was recorded for 30 min and stored every 10 s.