5R)-4-Allyl-2,5

ABSTRACT Among the different mechanisms underlying opioid tolerance, receptor desensitization would represent a major cellular adaptation process in which the role of receptor internalization is still a matter of debate. In the present study, we examined desensitization of the human ␦-opioid receptor (hDOR) produced by endogenous opioid peptides Leu-enkephalin (Tyr-Gly-Gly-PheLeu) and Met-enkephalin (Tyr-Gly-Gly-Phe-Met), and the contribution of internalization in this process. Results obtained with natural peptides were compared with those produced by a synthetic opioid agonist, SNC-80 . After a 30-min treatment, we observed a different regulation of hDOR between agonists. SNC-80 produced a stronger and faster desensitization and was associated with a loss of opioid binding sites by 50%. SNC-80 also caused a marked hDOR down-regulation by 30% as observed by Western blot. Immunocytochemistry revealed that SNC-80 induced a complete redistribution of hDOR from cell surface into intracellular compartments, whereas a partial internalization was visualized upon enkephalin exposure. In constrast, a stronger hDOR recycling and resensitization were measured after enkephalin treatment compared with SNC-80. These data strongly suggested a differential sorting of the internalized receptors caused by enkephalins and SNC-80 that was further confirmed by chloroquine as a lysosomal degradation blocker and monensin as a recycling endosome inhibitor. Finally, by preventing hDOR internalization with 0.5 M sucrose, we demonstrated that hDOR internalization contributes partially to desensitization. In conclusion, hDOR desensitization depends both on its internalization and its sorting either to the recycling pathway or to lysosomes. The three opioid receptors, namely, ␦-, -, and -opioid receptors, are regulated not only by endogenous opioid peptides but also by numerous pharmacologically active drugs, particularly those used in the clinical management of pain 35 S]guanosine 5Ј-O-(3-thio)triphosphate binding. Desensitization could result from uncoupling between opioid receptors and G proteins, whereas contribution of a decrease of opioid receptor number is less clear in this process. For example, a prolonged treatment (24 h) of the R1.1 cell line by (trans)-3,4-dichloro-N-methyl-N-[2-(1-pyrrolidinyl)-cyclohexyl]benzeneacetamide (U50,488) reduced the number of -opioid receptors by 50% without any significant Article, publication date, and citation information can be found a