Pharmacological Delta1- and Delta2-Opioid Receptor Subtypes in the Human Neuroblastoma Cell Line SK-N-BE: No Evidence for Distinct Molecular Entities.
Résumé
The two pharmacological δ-opioid receptor subtypes, δ1 and δ2, have been defined on the basis of pharmacological tools but remain to be characterized at the molecular level, since only a single cDNA has been cloned. The present study aimed to investigate the pharmacological properties of δ1- and δ2-opioid subtypes expressed in the human neuroblastoma cell line SK-N-BE and to characterize their putative corresponding mRNAs. Binding experiments using “selective” δ1- and δ2-opioid agonists and antagonists revealed the presence of two binding sites, demonstrating the presence of these δ1-opioid subtypes as they were previously described. The activation of these pharmacological subtypes by the selective agonists induced the incorporation of [α-32P]azidoanilide-GTP into Gαi2/Gα0 subunits with the same efficiency and potency and inhibited adenosine 3′,5′-cyclic monophosphate (cAMP) accumulation with similar efficiency, while their sustained activation for 15 min induced a cross-desensitization. The “selective” δ1 and δ2 antagonists, 7-benzylidenenaltrexone and naltrindole benzofuran, respectively, were found to be as potent in blocking the inhibition of cAMP accumulation induced by both [D-Pen2,5]enkephalin and Tyr-D-Ala-Phe-Asp-Val-Val-Gly-NH2. The possibility that δ-opioid subtypes could arise from alternative splicing was ruled out by reverse transcription–polymerase chain reaction (RT–PCR) experiments and the sequencing of PCR products, which revealed the presence of a single transcript encoding for the δ-opioid receptor. Different possibilities which could account for the δ-opioid receptor heterogeneity observed in the SN-N-BE cell line are discussed.