Previous studies have shown that tolerance to the antinociceptive effect of morphine develops after a prolonged exposure, but its mechanisms remain unclear. In the present study, we examined whether anti-morphine antibody produced by chronic morphine exposure would contribute to the development of morphine antinociceptive tolerance in rats. Our results showed that anti-morphine antibody was present in rats rendered tolerance to antinociception after intrathecal morphine exposure for seven consecutive days. Superfusion of anti-morphine antibody onto spinal cord slice dose-dependently produced an inward excitatory current in spinal cord dorsal horn neurons using whole-cell patch-clamp recording, which surpassed morphine-induced outward inhibiting current. Co-administration of morphine with a monoclonal antibody (2.4G(2)) against Fc receptors for seven days significantly attenuated the production of anti-morphine antibody as well as the behavioral manifestation of morphine tolerance in same rats. These results indicate that anti-morphine antibody produced by morphine exposure may contribute to the development of morphine tolerance possibly through counteracting the inhibitory morphine effect on spinal cord dorsal horn neurons.