Skip to main content
NCBI home page
The Journal of Clinical Investigation logoLink to The Journal of Clinical Investigation
. 1978 May;61(5):1136–1144. doi: 10.1172/JCI109028

Characterization of the Human Platelet α-Adrenergic Receptor

CORRELATION OF [3H] DIHYDROERGOCRYPTINE BINDING WITH AGGREGATION AND ADENYLATE CYCLASE INHIBITION

R Wayne Alexander 1,2,3, Barry Cooper 1,2,3, Robert I Handin 1,2,3
PMCID: PMC372633  PMID: 207726

Abstract

Human platelets aggregate and undergo a release reaction when incubated with catecholamines. Indirect evidence indicates that these events are mediated through α-adrenergic receptors. We used [3H]dihydroergocryptine, an α-adrenergic antagonist, to identify binding sites on platelets that have the characteristics of α-adrenergic receptors. Catecholamines compete for the binding sites in a stereo-specific manner with the potency series of (−) epinephrine > (−) norepinephrine > (±) phenylephrine > (−) isoproterenol. The dissociation constant (Kd) of (−) epinephrine is 0.34 μM. Binding is saturable using a platelet particulate fraction but not with intact platelets. There are 0.130 pmol binding sites per milligram protein in fresh platelet membranes. This number represents approximately 100 binding sites per platelet. The Kd for [3H]-dihydroergocryptine was 0.003−0.01 μM. The α-adrenergic antagonist phentolamine (Kd = 0.0069 μM) was much more potent than the β-adrenergic antagonist (±) propranolol (Kd = 27 μM) in competing for the binding sites. The binding data were correlated with catecholamine-induced platelet aggregation and inhibition of basal and prostaglandin E1-stimulated adenylate cyclase. (−) Epinephrine was more potent than (−) norepinephrine in producing aggregation whereas (−) isoproterenol was ineffective. The threshold dose for inducing aggregation by (−) epinephrine was 0.46 μM. Phentolamine and dihydroergocyrptine blocked this response, whereas (±) propranolol had no effect. (−) Epinephrine and (−) norepinephrine inhibited basal and prostaglandin E1-stimulated adenylate cyclase in a dose-related manner. The concentration of (−) epinephrine inhibiting adenylate cyclase 50% was 0.7 μM. This inhibition was also blocked by phentolamine and dihydroergocryptine but not by (±) propranolol. The specificity of binding and the close correlation with α-adrenergic receptor-mediated biochemical and physiological responses suggest that the [3H]dihydroergocryptine binding site represents, or is closely related to, the human platelet α-adrenergic receptor. The ability to assay this receptor directly and to correlate these data with independently measured sequelae of receptor activation should facilitate increased understanding of the physiology and pathophysiology of the human platelet α-adrenergic receptor.

Full text

PDF
1136

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Barthel W., Markwardt F. Aggregation of blood platelets by biogenic amines and its inhibition by antiadrenergic and antiserotoninergic agents. Biochem Pharmacol. 1974 Jan 1;23(1):37–45. doi: 10.1016/0006-2952(74)90311-6. [DOI] [PubMed] [Google Scholar]
  2. Buchanan G. R., Martin V., Levine P. H., Scoon K., Handin R. I. The effects of "anti-platelet" drugs on bleeding time and platelet aggregation in normal human subjects. Am J Clin Pathol. 1977 Sep;68(3):355–359. doi: 10.1093/ajcp/68.3.355. [DOI] [PubMed] [Google Scholar]
  3. Carvalho A. C., Colman R. W., Lees R. S. Platelet function in hyperlipoproteinemia. N Engl J Med. 1974 Feb 21;290(8):434–438. doi: 10.1056/NEJM197402212900805. [DOI] [PubMed] [Google Scholar]
  4. Cheng Y., Prusoff W. H. Relationship between the inhibition constant (K1) and the concentration of inhibitor which causes 50 per cent inhibition (I50) of an enzymatic reaction. Biochem Pharmacol. 1973 Dec 1;22(23):3099–3108. doi: 10.1016/0006-2952(73)90196-2. [DOI] [PubMed] [Google Scholar]
  5. Cherrington A. D., Assimacopoulos F. D., Harper S. C., Corbin J. D., Park C. R., Exton J. H. Studies on the alpha-andrenergic activation of hepatic glucose output. II. Investigation of the roles of adenosine 3':5'-monophosphate and adenosine 3':5'-monophosphate-dependent protein kinase in the actions of phenylephrine in isolated hepatocytes. J Biol Chem. 1976 Sep 10;251(17):5209–5218. [PubMed] [Google Scholar]
  6. Cooper B., Gregerman R. I. Hormone-sensitive fat cell adenylate cyclase in the rat. Influences of growth, cell size, and aging. J Clin Invest. 1976 Jan;57(1):161–168. doi: 10.1172/JCI108256. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Davis J. N., Strittmatter W. J., Hoyler E., Leefkowitz R. J. [3H]dihydroergocryptine binding in rat brain. Brain Res. 1977 Aug 26;132(2):327–336. doi: 10.1016/0006-8993(77)90425-5. [DOI] [PubMed] [Google Scholar]
  8. Greenberg D. A., Prichard D. C., Snyder S. H. Alpha-noradrenergic receptor binding in mammalian brain: differential labeling of agonist and antagonist states. Life Sci. 1976 Jul 1;19(1):69–76. doi: 10.1016/0024-3205(76)90375-1. [DOI] [PubMed] [Google Scholar]
  9. Jakobs K. H., Saur W., Schultz G. Reduction of adenylate cyclase activity in lysates of human platelets by the alpha-adrenergic component of epinephrine. J Cyclic Nucleotide Res. 1976 Nov-Dec;2(6):381–392. [PubMed] [Google Scholar]
  10. LOWRY O. H., ROSEBROUGH N. J., FARR A. L., RANDALL R. J. Protein measurement with the Folin phenol reagent. J Biol Chem. 1951 Nov;193(1):265–275. [PubMed] [Google Scholar]
  11. Levine P. H. An acute effect of cigarette smoking on platelet function. A possible link between smoking and arterial thrombosis. Circulation. 1973 Sep;48(3):619–623. doi: 10.1161/01.cir.48.3.619. [DOI] [PubMed] [Google Scholar]
  12. Mills D. C., Roberts G. C. Effects of adrenaline on human blood platelets. J Physiol. 1967 Nov;193(2):443–453. doi: 10.1113/jphysiol.1967.sp008369. [DOI] [PMC free article] [PubMed] [Google Scholar]
  13. O'BRIEN J. R. SOME EFFECTS OF ADRENALINE AND ANTI-ADRENALINE COMPOUNDS ON PLATELETS IN VITRO AND IN VIVO. Nature. 1963 Nov 23;200:763–764. doi: 10.1038/200763a0. [DOI] [PubMed] [Google Scholar]
  14. Robison G. A., Butcher R. W., Sutherland E. W. Adenyl cyclase as an adrenergic receptor. Ann N Y Acad Sci. 1967 Feb 10;139(3):703–723. doi: 10.1111/j.1749-6632.1967.tb41239.x. [DOI] [PubMed] [Google Scholar]
  15. Ruffolo R. R., Jr, Fowble J. W., Miller D. D., Patil P. N. Binding of [3H]dihydroazapetine to alpha-adrenoreceptor-related proteins from rat vas deferens. Proc Natl Acad Sci U S A. 1976 Aug;73(8):2730–2734. doi: 10.1073/pnas.73.8.2730. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Salomon Y., Londos C., Rodbell M. A highly sensitive adenylate cyclase assay. Anal Biochem. 1974 Apr;58(2):541–548. doi: 10.1016/0003-2697(74)90222-x. [DOI] [PubMed] [Google Scholar]
  17. Salzman E. W., Neri L. L. Cyclic 3',5'-adenosine monophosphate in human blood platelets. Nature. 1969 Nov 8;224(5219):609–610. doi: 10.1038/224609a0. [DOI] [PubMed] [Google Scholar]
  18. Shattil S. J., Anaya-Galindo R., Bennett J., Colman R. W., Cooper R. A. Platelet hypersensitivity induced by cholesterol incorporation. J Clin Invest. 1975 Mar;55(3):636–643. doi: 10.1172/JCI107971. [DOI] [PMC free article] [PubMed] [Google Scholar]
  19. Strittmatter W. J., Davis J. N., Lefkowitz R. J. alpha-Adrenergic receptors in rat parotid cells. I. Correlation of [3H]dihydroergocryptine binding and catecholamine-stimulated potassium efflux. J Biol Chem. 1977 Aug 10;252(15):5472–5477. [PubMed] [Google Scholar]
  20. Williams L. T., Mullikin D., Lefkowitz R. J. Identification of alpha-adrenergic receptors in uterine smooth muscle membranes by [3H]dihydroergocryptine binding. J Biol Chem. 1976 Nov 25;251(22):6915–6923. [PubMed] [Google Scholar]

Articles from Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation

RESOURCES