RT Journal Article
SR Electronic
T1 Quantitation and Characterization of Human Platelet Alpha-Adrenergic Receptors Using [3H]Phentolamine
JF Molecular Pharmacology
JO Mol Pharmacol
FD American Society for Pharmacology and Experimental Therapeutics
SP 719
OP 728
VO 16
IS 3
A1 MICHAEL L. STEER
A1 JULIA KHORANA
A1 BERNARD GALGOCI
YR 1979
UL http://molpharm.aspetjournals.org/content/16/3/719.abstract
AB The binding of [3H]phentolamine to human platelet membranes was studied, and this ligand was found to bind to a single class of noncooperative sites with a dissociation constant (Kd) of 12 ± 3 nM. At saturation, 0.165 ± 0.060 pmoles were bound per mg protein, and each platelet was noted to possess 260 ± 95 binding sites. [3H]phentolamine binding was rapid (t1/2 < 15 sec) and reversible (t1/2 for dissociation <30 sec) at 30°. [3H]phentolamine binding was stereospecifically inhibited by epinephrine. Inhibition of [3H]phentolamine binding by catecholamines occurred with an order of potency of epinephrine > norepinephrine > isoproterenol and propranolol (1 µM) did not inhibit [3H]phentolamine binding. These observations indicate that [3H]phentolamine binds to sites having the characteristics of α-adrenergic receptors. Dihydroergocryptine was also noted to cause time and concentration-dependent inhibition of [3H]phentolamine binding. Comparison of these data to those previously reported using [3H]dihydroergocryptine as the binding probe (Newman, K. D., Williams, L. T., Bishopric, N. H. et al. J. Clin. Invest. 61: 395-402, 1978; Alexander, R. W., Cooper, B. and Handin, R. I. J. Clin. Invest. 61, 1136-1144, 1978) suggests that both [3H]phentolamine and [3H]dihydroergocryptine interact with the same binding sites. Dopamine and serotonin were found to inhibit [3H]phentolamine binding (Kd 1.9 and 15.1 mM respectively) indicating either that phentolamine can bind to receptors for dopamine and serotonin or these ligands bind to the platelet α-receptor. The binding of [3H]phentolamine to platelet membranes was not altered when GTP (0.01 mM) was present, but the nucleotide caused a tenfold shift in the inhibition curve for epinephrine, indicating that epinephrine was less avidly bound in the presence of GTP. ACKNOWLEDGMENT The authors gratefully acknowledge the assistance of Dr. W. Rein of Ciba-Geigy for help in obtaining [3H]phentolamine.