Apparent precoupling of κ- but not μ-opioid receptors with a G protein in the absence of agonist

Abstract

Rabbit and guinea-pig cerebellum membranes contain a very high (> 80%) proportion of μ- and κ-opioid receptors, respectively. Rabbit (μ) and guinea-pig (κ) cerebellum membranes wre (i) labeled either with the opiate agonist. [ 3H]etorphine (K d = 0.1–0.2 nM), or with the opiate antagonist. [ 3diprenorphine (K d = 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 (μ) membranes, bound [ 3H]etorphine sedimented faster (s 20 ≅ 12S) that bound [ 3H]diprenorphine (10s), while is those from guinea-pig cerebellum (κ) 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 [ 3etorphine complexes of the μ- and κ-opioid receptors as well as the [ 3Hdiprenorphine complex of the κ-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 μ- and κ-opioid receptors being capable of physically interacting with a G protein (GTP binding regulatory protein) yet, unlike the μ-opioid receptor which does so only in the presence of an agonist, the κ-opioid receptor appears to be precoupled with a G protein.