Hot plate. This test occurs approximately 2–3 h after the completion of the open field test. The lights in the testing area were turned off at least an hour prior to testing and animals were allowed to sit undisturbed in the darkened room. A lamp (15 W bulb) behind the hot plate (Hotplate Analgesia Meter, Model 39, IITC, Inc.) was faced away from the hot plate surface. A mirror was placed behind the hot plate so that the experimenter can observe the animal. The hot plate was maintained at 52°C. The mouse was placed on the center of the hot plate in a smoke gray Plexiglas bottomless cube and the built-in timer started. As soon as the animal elicited a pain response (i.e. paw licking, guarding, shaking or jumping), the timer was immediately stopped and the animal removed from the hot plate surface. If the animal did not show a response within 30 seconds, the test was stopped and the animal was assigned the 30 seconds maximum time as its response latency. Once an animal had been tested, it was placed in a holding cage until all animals from the home cage have been tested.
