In vivo occupation of mouse brain opioid receptors by endogenous enkephalins: blockade of enkephalin degrading enzymes by RB 101 inhibits [3H]diprenorphine binding.

With the aim of possibly studying the local activity of brain enkephalinergic pathways by autoradiography and positron emission tomography, preliminary competition experiments of [3H]diprenorphine binding in mouse brain were carried out after i.v. administration of the first systemically-active mixed inhibitor of enkephalin degrading enzymes RB 101 (N(R,S)-2-benzyl-3[(S)-(2-amino-4-methylthiobutyldithio]-1-oxoprop yl]- L-phenylalanine benzyl ester). Although devoid of affinity for the opioid binding sites, RB 101 inhibited the [3H]diprenorphine binding to the opioid receptors in a dose-dependent manner. This effect, very likely due to an RB 101-induced increase in extracellular levels of enkephalins, reached a plateau at a dose of 10 mg/kg, where almost 30% displacement was observed. Intravenous administration of either 5 or 20 mg/kg of RB 101 in mice submitted to warm-swim stress led to an additional [3H]diprenorphine displacement, which reached 45% compared to unstressed controls. This ceiling effect could account for the reported minimal morphine-like side effects induced by mixed inhibitors. A large increase in endogenous enkephalin levels induced by RB 101, associated or not with stress, was also indirectly demonstrated by the analgesic responses elicited by i.v. injection of the mixed inhibitor. This effect was blocked by naloxone but not by the delta antagonist naltrindole (NTI), supporting a preferential implication of mu receptors in supraspinal analgesia. Taken together, these results suggest that RB 101 could be used to determine the precise in vivo localization of enkephalinergic pathways recruited by various stimuli.