The effects of mu- and delta-preferring agonists on adenylate cyclase activity have been investigated in vitro in homogenates of guinea pig cochleas. Morphine, Leu-enkephalin, D-Ala2, N-methyl-Phe4, Gly-ol5-enkephalin (DAGO) and D-Ser2-Leu-enkephalin-Thr (DSLET) each inhibited the synthesis of cyclic AMP. This effect was reversed by naloxone which had a greater affinity in blocking the effect of the mu-preferring agonists (morphine, DAGO) than in blocking the effect of the delta-preferring agonists (Leu-enkephalin, DSLET). Finally, no additive effects were observed when various combinations of two agonists were used. These results indicate that opioid receptors exist in the guinea pig cochlea and that they are negatively linked to adenylate cyclase. The different affinities shown by naloxone to reverse the inhibition induced by the mu- and delta-preferring agonists suggest that morphine and DAGO act through mu-receptors, whereas Leu-enkephalin and DSLET act through delta-receptors. Since no additive effects have been found when combining two different agonists, it can be hypothesized that the mu- and delta-receptors are coupled to the same pool of adenylate cyclase. It may be proposed from these findings that in vivo enkephalins inhibit the synthesis of cyclic AMP via mu- and delta-receptors. However, whether this effect occurs at a presynaptic level (within opioid-containing olivocochlear varicosities) or at the postsynaptic level (within dendrites of the primary auditory neurons) remains to be determined.