((2S,3S)-[cis-2-(diphenylmethyl)-N-[(2-methoxyphenyl)-methyl]-1-az abicyclo[2.2.2]octan-3-amine]) (CP-96,345) noncompetitively inhibits substance P (SP) binding at the neurokinin-1 (NK-1) site and has been widely used to determine the extent of NK-1 activity in nociception. To test the selectivity of this compound in vivo regarding other putative nociceptive transmitters, such as excitatory amino acids, we compared the actions of CP-96,345 to those of ((2R,3R)-[cis-2-(diphenylmethyl)-N-[(2-methoxyphenyl)-methyl]-1-az abicyclo[2.2.2]octan-3-amine]), a less active isomer, on behavioral responses induced by SP, N-methyl-D-aspartate (NMDA) and kainic acid (KA) injected intrathecally in mice. When injected intrathecally, SP, NMDA or KA produce a caudally directed biting and scratching behavior that lasted for approximately 60 to 90 sec. At a dose as high as 2 nmol, CP-96,345 had no effect on responses induced by a single injection of 22.5 pmol of SP. In contrast, NMDA-induced behaviors were inhibited by CP-96,345 in a dose-related fashion beginning at a dose as low as 0.02 nmol. There was also an inhibitory effect of CP-96,345 on KA-induced activity that was not dose related. The more potent inhibitor of [3H] SP binding, (+)-(2S,3S)-3-(2-methoxybenzylamino)-2-phenylpiperidine (CP-99,994), was approximately 10 times more potent in inhibiting NMDA-induced activity than CP-96,345. CP-99,994 also inhibited NMDA-induced activity at doses that failed to inhibit SP-induced behavior. Also attenuated by CP-96,345 was the development of sensitization to the behavioral effects produced by repeated injections of KA and desensitization to repeated injections of SP, phenomena linked to an action of the N-terminus of SP. NMDA-induced behaviors and sensitization to KA were found to be sensitive to verapamil, consistent with their mediation by calcium. These results indicate that either CP-96,345 and CP-99,994 do not inhibit NK-1-induced activity in the mouse spinal cord, or that exogenously administered SP does not induce behavioral responses by an interaction with NK-1 receptors. Whether CP-96,345 acts by a mechanism that involves inhibition of calcium channels and/or SP N-terminal activity requires further testing.