Agonist-, antagonist-, and inverse agonist-regulated trafficking of the delta-opioid receptor correlates with, but does not require, G protein activation.
In this study, we explored the relationship between ligand-induced regulation of surface delta opioid receptors and G protein activation. G protein activation was assessed with [(35)S]guanosine-5'-O-(3-thio)triphosphate (GTP gamma S) binding assays conducted at both 37 and 0 degrees C. Ligand-independent (constitutive) activity of the delta-receptor was readily observed when the [(35)S]GTP gamma S binding assay was performed at 37 degrees C. We identified a new class of alkaloid inverse agonists (RTI-5989-1, RTI-5989-23, RTI-5989-25), which are more potent than the previously described peptide inverse agonist ICI-174864 (N,N-diallyl-Tyr-Aib-Aib-Phe-Leu). Treatment with these inverse agonists for 18 h caused up-regulation of surface receptors. Eighteen-hour treatment with etorphine resulted in approximately 90% loss of surface receptor, whereas fentanyl, diprenorphine, and morphine caused between 20 and 50% loss. The abilities of ligands to modulate [(35)S]GTP gamma S binding at 37 degrees C showed a strong correlation with their abilities to regulate surface receptor number (r(2) = 0.86). Interestingly, the ability of fentanyl to activate G proteins was markedly temperature sensitive. Fentanyl showed no stimulation of [(35)S]GTP gamma S binding at 0 degrees C but was as efficacious as etorphine, morphine, and diprenorphine at 37 degrees C. Neither the ligand-induced receptor increases nor decreases were perturbed by pertussis toxin pretreatment, suggesting that functional G proteins are not required for ligand-regulated delta-opioid receptor trafficking.
Zaki
P A
PA
Department of Psychiatry and Biobehavioral Sciences, University of California, Los Angeles, California 90024-1759, USA.
Keith
D E
DE
Thomas
J B
JB
Carroll
F I
FI
Evans
C J
CJ
D001711:Biotransformation / Q000502:physiology; D002460:Cell Line; D002462:Cell Membrane / Q000378:metabolism; D004743:Enkephalin, Leucine / Q000031:analogs & derivatives / Q000494:pharmacology; D005434:Flow Cytometry; D019204:GTP-Binding Proteins / Q000378:metabolism*; D016244:Guanosine 5'-O-(3-Thiotriphosphate) / Q000494:pharmacology; D008024:Ligands; D009292:Narcotic Antagonists / Q000494:pharmacology; D037342:Pertussis Toxin; D017465:Receptors, Opioid, delta / Q000819:agonists* / Q000037:antagonists & inhibitors* / Q000235:genetics; D015398:Signal Transduction / Q000187:drug effects; D010566:Virulence Factors, Bordetella / Q000494:pharmacology
D008024:Ligands; D009292:Narcotic Antagonists; D017465:Receptors, Opioid, delta; D010566:Virulence Factors, Bordetella; D016244:Guanosine 5'-O-(3-Thiotriphosphate); D004743:Enkephalin, Leucine; C040643:N,N-diallyl-tyrosyl-alpha-aminoisobutyric acid-phenylalanyl-leucine; D037342:Pertussis Toxin; D019204:GTP-Binding Proteins