BETA-ADRENERGIC-RECEPTOR ASSAY IN PERMEABLE CELLS

We developed a method for assaying agonist binding to study the coupli ng of beta-adrenergic receptors to G proteins in an environment that r esembles an intact cell. This method, devised using sapinin-permeable cell suspensions, permits investigation of beta-adrenergic receptor be havior under conditions where the receptor appears to be tightly coupl ed to Os protein to activate adenylyl cyclase. In this communication, we compare the binding characteristics of beta-adrenergic receptors in permeable cell suspensions and cell membrane preparations. The cell l ines examined here, by means of [I-125]pindolol binding, were C6 gliom a cells and NCB hybrid cells. Scatchard plots for permeable cells were linear with correlation coefficients greater than 0.97 and the best f it was by the single site model. Furthermore, Bmax values of C6 cells and NCB cells were consistent with those of membranes. However, IC50 v alues of isoproterenol competition for [I-125]pindolol binding sites a nd the extent of GppNHp-induced changes of IC50 were not identical in permeable cells and cell membranes, suggesting that beta-adrenergic re ceptors behave differentially in the undisrupted cellular milieu than in a membrane preparation. Our newly devised method permits one to obt ain further information about receptor coupling in a nearly intact cel l.