Apparent precoupling of kappa- but not mu-opioid receptors with a G protein in the absence of agonist.
Rabbit and guinea-pig cerebellum membranes contain a very high (greater than 80%) proportion of mu- and kappa-opioid receptors, respectively. Rabbit (mu) and guinea-pig (kappa) cerebellum membranes were (i) labeled either with the opiate agonist, [3H]etorphine (Kd = 0.1-0.2 nM), or with the opiate antagonist, [3H]diprenorphine (Kd = 0.1 nM), in the absence or presence of Na+ and/or 5'-guanylylimidodiphosphate (GppNHp), (ii) solubilized with digitonin (1%, w:v) and (iii) the radioactivity in the soluble extracts analyzed by ultracentrifugation in sucrose gradients. In the soluble extracts from rabbit cerebellum (mu) membranes, bound [3H]etorphine sedimented faster (S20,w congruent to 12S) than bound [3H]diprenorphine (10S), while in those from guinea-pig cerebellum (kappa) membranes, bound [3H]etorphine and bound [3H]diprenorphine sedimented at the same position (12S). Na+ selectively decreased recovery of the bound tritiated agonist in the two soluble preparations. When they had been generated in the presence of GppNHp but in the absence of Na+, the [3H]etorphine complexes of the mu- and kappa-opioid receptors as well as the [3H]diprenorphine complex of the kappa-opioid receptor were all recovered at position 10S, indicating that GppNHp had induced a decrease of the apparent molecular size of the two types of opioid receptors. These data are interpreted in terms of mu- and kappa-opioid receptors being capable of physically interacting with a G protein (GTP binding regulatory protein) yet, unlike the mu-opioid receptor which does so only in the presence of an agonist, the kappa-opioid receptor appears to be precoupled with a G protein.
Frances
B
B
Unité de Neuropharmacologie Moléculaire, C.N.R.S. UPR 8221, Toulouse, France.
Puget
A
A
Moisand
C
C
Meunier
J C
JC
D000818:Animals; D002499:Centrifugation, Density Gradient; D002531:Cerebellum / Q000378:metabolism / Q000648:ultrastructure; D004072:Digitonin / Q000494:pharmacology; D004174:Diprenorphine / Q000378:metabolism*; D005048:Etorphine / Q000378:metabolism*; D019204:GTP-Binding Proteins / Q000378:metabolism*; D006165:Guanylyl Imidodiphosphate / Q000494:pharmacology; D006168:Guinea Pigs; D007700:Kinetics; D008566:Membranes / Q000378:metabolism / Q000648:ultrastructure; D009292:Narcotic Antagonists; D011817:Rabbits; D011957:Receptors, Opioid / Q000378:metabolism*; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D012964:Sodium / Q000494:pharmacology; D014316:Tritium
D009292:Narcotic Antagonists; D011957:Receptors, Opioid; D017473:Receptors, Opioid, kappa; D017450:Receptors, Opioid, mu; D014316:Tritium; D004174:Diprenorphine; D006165:Guanylyl Imidodiphosphate; D005048:Etorphine; D012964:Sodium; D019204:GTP-Binding Proteins; D004072:Digitonin