Interaction of Opiates with Opioid Binding Sites in the Bovine Adrenal Medulla: II. Interaction with K Sites

Abstract

Abstract: In this study we examined the interaction of opiates with K binding sites in the bovine adrenal medulla. [ 3 H]Ethylketocyclazocine (EKC), [ 3 H]etorphine, and [ 3 H]bremazocine stereoselective bindings were used to assay these interactions. The K sites were found to be heterogeneous: [ 3 H]bremazocine identified with high affinity all subtypes of these sites. [ 3 H]EKC, in the presence of saturating concentrations of [D‐Ala 2 , D‐Leut]‐enkephalin (DADLE) (5μ M ), was used to identify K 1 sites, on which dynorphin A (1–13) bound with high affinity. Either [ 3 H]EKC or [ 3 H]etorphine in the presence of 5μ M DADLE identified the K 2 subtype. This subtype was found to interact with β‐endorphin and especially with the octapeptide Met 5 ‐enkephalyl‐Arg 6 ‐Gly 7 ‐Leu 8 . Furthermore, [ 3 H]etorphine identified in the bovine adrenal medulla a third high‐affinity component, in the presence of 5 μ M DADLE. This residual interaction was found to be equally stereoselective and presenting K selectivity. Met 5 ‐enkephalyl‐Arg 6 ‐Phe 7 interacted preferentially with this site. The three K subtypes interacted differentially with monovalent (Na + , K + , and Li + ) and divalent (Ca 2+ , Mg 2+ , and Mn 2+ ) ions by modification of the apparent concentration of the accessible sites and/or by changes of the apparent K D for radioligands. Modifying agents (proteolytic enzymes, thiol‐modifying reagents, and A 2 ‐phospholipase) produced different effects on each subtype of the K site, suggesting a different protein (or protein‐lipid?) composition.