Zinc ions can totally block stereospecific binding of 3H-met-enkephalinamide (2-D-ala-5-L-methionine) to opiate receptor sites in synaptic membranes of the hippocampus, the cerebral cortex and the basal ganglia of the rat brain. Analysis of binding isotherms indicates that this inhibitory effect involves a decrease in both receptor affinity and the number of binding sites. Our data also suggest that the zinc ions react with essential SH-groups of the opiate receptor. The endogenous concentrations of zinc ions in the hippocampus, the cerebral cortex and the basal ganglia are compatible with the concentrations needed to inhibit opiate receptor binding in vitro. Especially the hippocampus contains a high concentration of zinc ion, which are localized exclusively in the giant boutons of the mossy fibers. Further, the hippocampal distribution of enkephalin and zinc ions is identical and confined to the mossy fiber zone. Hence, zinc ions may represent important modulators of opiate receptor binding in the central nervous system, particularly in the hippocampal mossy fiber zone.