Antinociception was assessed using the hotplate latency test. Individual mice were placed on the hotplate, set at 52 ± 0.02°C, and observed for the first behavioral signs of nociception (time to lick/flick their front and hind paws and jump from the hotplate). A cut-off time of 60 s was used to minimize tissue damage. Predosing reaction times were the average of the three readings taken approximately 15 min apart. After subcutaneous administration of morphine (3 mg/kg) (or saline) antinociceptive testing was performed at the following times: 15, 30, 45, 60, 75, 90 and 180 min. Hotplate latency times were then converted to % maximal possible effect (MPE) using the following formula28: %MPE=(Post drug latency)−(pre drug latency)(Maximum latency)−(pre drug latency)×100
