The narcotic antagonist naloxone has been reported to decrease locomotor activity in the rat, presumably blocking endogenous opiate systems. Naloxone has a greater affinity for receptors which preferentially bind morphine and other opiate alkaloids as compared to receptors that bind endogenous opioid peptides. Diprenorphine, another pure opiate antagonist, binds with equal affinity to both receptor subtypes. Therefore, the effects of the narcotic antagonists naloxone and diprenorphine on spontaneous activity were compared in rats and mice, tested individually and in pairs. Only naloxone (10 mg/kg) affected spontaneous activity in rats tested individually, decreasing both gross and fine activity. In rats tested in pairs, naloxone (1.0 and 10 mg/kg) decreased both fine and gross activity, while diprenorphine (10 mg/kg) produced significant decreases only in fine activity. In mice tested individually, naloxone produced modest (nonsignificant) decreases in activity while diprenorphine (10 mg/kg) significantly enhanced activity. Neither opiate antagonist produced consistent effects on activity in paired mice. These results illustrate the species and situation dependence of the effects of opiate antagonists and point out the need for testing more than one narcotic antagonist in research designed to provide inferential information concerning possible physiological functions of endogenous opioid peptides.