The role of cerebroside sulfate in opiate action and binding was studied by examining the effect of decreasing the availability of the glycolipid in vivo on morphine analgetic activity and receptor affinity. Available cerebroside sites were decreased either by injecting Azure A at a dose with selective high affinity for sulfo-lipids, or by using "jimpy" mice, a genetic leukodystropic mutant mouse with a deficiency in brain sulfatides. Injections of Azure A (4.4 MG/KG) intracerebroventricularly produced a 2-fold increase in the AD50 of morphine (tail-flick test) within 2 hr. The genetic mutation also resulted in a decreased sensitivity to morphine; the morphine AD50 was 6- to 11-fold higher in jimpy mice than in their normal littermates. Azure A produced a dose-dependent inhibition of opiate binding to synaptosomal plasma membranes. There was 85% inhibition of [3H]morphine binding with 0.2 micrometer Azure A, and 40% inhibition of [3H]naloxone binding with 5 micrometer Azure A. The inhibition of morphine binding was competitive and was demonstrated to be different from that elicited by Na+. Synaptic membranes from jimpy mice exhibited a decrease in number of binding sites for morphine. Binding of Azure A to cerebroside sulfate, associated with or in the proximity of the receptor sites, was suggested by the fact that in the presence of 0.2 micrometer Azure A, no inhibition of [3H]morphine binding to synaptosomal plasma membranes of jimpy mice could be demonstrated whereas 31% inh-bition was observed in membranes from control littermates. Based on the findings, it is concluded that cerebroside sulfate strategically located may have a role in binding morphine and mediating its effects.