Differential Regulation of two Molecular Forms of a μ-Opioid Receptor Type by Sodium Ions, Manganese Ions and by Guanyl-5′-YL Imidodiphosphate

Abstract

The rabbit cerebellum contains a very high proportion (up to 80 %) of μ-opioid receptor sites (Meunier, J.C., Kouakou, Y., Puget, A. and Moisand, C., Mol. Pharmacol. 24, 23–29, 1983). A membrane fraction derived therefrom is labeled either with the opioid agonist, ³H-etorphine or with the opioid antagonist, ³H-diprenorphine, and solubilized with digitonin. Centrifugation of the soluble extracts in linear sucrose gradients reveals that bound ³H-etorphine sediments faster than does bound ³H-diprenorphine: 12S vs 10S. Pre-incubation of membranes and radioligand in the presence of 120 mM NaCl results in considerably decreased recovery of the ³H-agonist in 12S form while recovery of the ³H-antagonist in 10S form is substantially increased. The opposite situation is observed when the membranes have been prelabeled with radioligand in the presence of 1 mM MnCl₂. Guanyl-5′-yl imidodiphosphate, a metabolically stable structural analog of GTP is found to selectively reduce recovery of labeled 12S receptors while it does not affect that of labeled 10S receptors. These data indicate that the μ-opioid receptor from rabbit cerebellum is capable of existing in two forms which differ in apparent molecular size: an “antagonist” (10s) form of apparent Mr z 230,000 which is stabilized in the presence of sodium ions and an “agonistl’ (12s) form of apparent Mr % 300,000 which, unlike the antagonist one, is sensitive to guanyl-5′-yl imidodiphosphate. It is thought that the form of larger apparent size represents the p-opioid receptor associated with a guanine nucleotide regulatory protein.