%0 Journal Article %A CHHABIRANI MUKHERJEE %A ROBERT J. LEFKOWITZ %T Regulation of Beta Adrenergic Receptors in Isolated Frog Erythrocyte Plasma Membranes %D 1977 %J Molecular Pharmacology %P 291-303 %V 13 %N 2 %X The ability of beta adrenergic agonists to inactivate reversibly the beta adrenergic receptor binding sites in isolated frog erythrocyte plasma membranes has been studied. When membranes were exposed to isoproterenol at 25°,40-70% of the beta adrenergic receptors were rapidly lost, i.e., could no longer be assayed with (-)-[3H]dihydroalprenolol. Although the rate of this process was considerably more rapid in membranes than previously found in whole cells, it was still slower than the rate of stimulation of the membrane-bound adenylate cyclase by isoproterenol. In agreement with previous findings in whole cells, the decreased receptor binding in membranes was associated with a fall in receptor number with no significant change in affinity of the remaining receptors. The specificity of this process in membranes was that of a beta adrenergic receptor-mediated effect. The order of potency of agonists in inducing the apparent fall in receptor number was (-)-isoproterenol> (-)-epinephrine>> (-)-norepinephrine. (-)-Isoproterenol was 1000 times more potent than (+)-isoproterenol. Beta adrenergic antagonists competitively antagonized the agonist-induced effect. (-)-Propranolol was 100 times more potent than (+)-propranolol in this regard. Among a series of 11 beta adrenergic agents tested, the ability to decrease receptor number maximally was directly correlated with maximum ability to stimulate adenylate cyclase (intrinsic activity); r = 0.93, p < 0.001. However, adenosine 3',5'-monophosphate (cAMP) did not appear to be involved in this desensitization process, since no substrate ATP was present during the incubations of membranes with isoproterenol, and cAMP at concentrations up to 0.5 mM did not reproduce the effect. Interventions which uncouple beta adrenergic receptors and adenylate cyclase, such as solubilization or filipin treatment of membranes, prevented the agonist-induced fall in receptor number. Guanine nucleotides and analogues, such as 5'-guanylylimidodiphosphate, completely prevented the agonist effect. Nucleotides also completely restored receptor number to normal after the agonist-induced lowering had occurred. Several sulfhydryl reagents, including N-ethylmaleimide, p-hydroxymercuribenzoate, and dithiothreitol, markedly inhibited the agonist-induced fall in receptor number and also caused partial or complete restoration of receptor number toward normal. A wide variety of other group-specific reagents, as well as several oxidizing and reducing agents, were without effect. These data suggest the necessity for beta receptor-adenylate cyclase coupling as a primary requisite for the regulatory effect of agonists on the beta receptors. cAMP formation does not appear to be required. ACKNOWLEDGMENT The authors wish to thank Dr. Marc G. Caron for assistance with the studies of solubilized preparations. %U https://molpharm.aspetjournals.org/content/molpharm/13/2/291.full.pdf