In the present study, we studied the interaction between serotonergic (5-HTergic) and gamma-aminobutyric acid (GABA)-ergic systems in the modulation of analgesia from morphine, a mu-opioid agonist, and U50,488, a kappa-opioid agonist. All experiments were performed in mice using the 49 degrees C tail-withdrawal assay. The benzodiazepine receptor agonist, diazepam, the serotonin synthesis inhibitor, para-chlorophenylalanine (p-CPA), and the 5-HT(1A) receptor agonist, 8-OH-DPAT, were all found to attenuate morphine and U50,488 analgesia. In each case, the attenuation was itself blocked by treatment with L-5-HTP, a serotonin precursor, bicuculline, a GABA(A) receptor antagonist or picrotoxin, a GABA(A)-gated chloride channel blocker. Neither L-5-HTP nor the GABA(A) receptor antagonists were found to affect morphine or U50,488 analgesia per se. Thus, these findings indicate that a benzodiazepine-GABAergic agent (diazepam) attenuates opioid analgesia through the serotonergic system, and antiserotonergic agents (8-OH-DPAT, p-CPA) attenuate opioid analgesia through the GABAergic system. The intimate interactions between GABA and serotonin in the present study further suggest that these neurotransmitters work in complex ways together rather than alone in the modulation of opioid analgesia.