We investigated the effects of morphine on the noxious mechanical stimuli-induced release of immunoreactive substance P (iSP) from the rabbit dorsal horn in vivo. Systemic morphine in a dose of 10 mg/kg, but not 1 mg/kg, inhibited the iSP release induced by noxious stimuli, although both dosages inhibited the nociceptive responses of rabbits to the stimuli. The inhibitory effect of morphine (10 mg/kg) on the iSP release was partially or completely antagonized by the local application of naloxone or prazosin, respectively, to the dorsal horn. The local application of methysergide did not affect the morphine action. The inhibitory effect of morphine (10 mg/kg) on the iSP release was diminished in acute spinal animals. These results suggest that the inhibition of the evoked release of substance P participates only in the action of the larger dose of morphine, and that this inhibition of the substance P release may be mediated by opioid receptors and the noradrenergic system, but not the serotonergic system, in the spinal cord.