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Thromb Haemost 1999; 81(01): 111-117
DOI: 10.1055/s-0037-1614427
Review Article
Schattauer GmbH

Evidence for Two Distinct G-protein-coupled ADP Receptors Mediating Platelet Activation

Hans-Michael Jantzen
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
,
Laurent Gousset
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
,
Vinay Bhaskar
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
,
Diana Vincent
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
,
Albert Tai
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
,
Elwood E. Reynolds
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
,
Pamela B. Conley
1   From COR Therapeutics, Inc., South San Francisco, CA, USA
› Author Affiliations
Further Information

Correspondence to:

H.-M. Jantzen
COR Therapeutics, Inc., 256 E.
Grand Ave.
South San Francisco CA 94080 USA
Phone: +16502446831   
Fax: +16502449208   

Publication History

Received10 June 1998

Accepted after revision09 September 1998

Publication Date:
08 December 2017 (online)

Also available at
 
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Summary

The identity of the receptors mediating platelet activation by ADP remains elusive. To distinguish between platelet ADP receptor subtypes, the effects of antagonists on platelet responses and the cloned P2Y1receptor, a putative platelet ADP receptor, have been investigated. 2-methylthio-AMP (2MeSAMP), an inhibitor of ADP-dependent platelet aggregation, antagonized ADP-mediated inhibition of adenylyl cyclase, competed with binding of [3H]2-methylthio-ADP and inhibited the stimulation of [35S]GTP γS binding. 2MeSAMP did not inhibit platelet shape change and was only a weak antagonist of intracellular calcium mobilization in platelets or in cells expressing the cloned human P2Y1receptor. By contrast, the P2Y1receptor antagonist adeno-sine 3’,5’-diphosphate (A3P5P) inhibited ADP-induced platelet aggregation, completely abolished shape change, but did not antagonize ADP effects on cyclic AMP generation or [3H]2-methylthio-ADP binding. However, A3P5P antagonized intracellular calcium mobilization in platelets and cells expressing the cloned P2Y1receptor. Furthermore, using a specific monoclonal antibody and flow cytometry, P2Y1receptor protein was detected on human platelets. These results support the existence of two G protein-coupled ADP receptors mediating platelet aggregation, one of which is coupled to Giproteins and blocked by 2MeSAMP, whereas the second receptor is similar or identical to P2Y1and coupled to Gq.


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  • References

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    CrossrefPubMedGoogle Scholar
  • 2 Hourani SMO, Hall DA. Receptors for ADP on Human Blood Platelets. Trends Pharmacol Sci 1994; 15: 103-8.
    CrossrefPubMedGoogle Scholar
  • 3 Mills DCB. ADP Receptors on Platelets. Thromb Haemost 1996; 76: 835-6.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 4 Puri RN, Colman RW. ADP-Induced Platelet Activation. Crit Rev Biochem Mol Biol 1997; 3: 437-502.
    PubMedGoogle Scholar
  • 5 Boarder MR, Hourani SMO. The regulation of vascular function by P2 receptors: multiple sites and multiple receptors. Trends Pharmacol Sci 1998; 19: 99-107.
    CrossrefPubMedGoogle Scholar
  • 6 Sage SO. Calcium Entry Mechanisms in Human Platelets. Exp Physiol 1997; 82: 807-23.
    CrossrefPubMedGoogle Scholar
  • 7 Greco NJ, Yamamoto N, Jackson BW, Tandon NN, Moos M, Jamieson GA. Identification of a nucleotide-binding site on glycoprotein IIb. J Biol Chem 1991; 266: 13627-33.
    PubMedGoogle Scholar
  • 8 Harden TK, Boyer JL, Nicholas RA. P2 Purinergic Receptors: Subtype-Associated Signaling Responses and Structure. Annu Rev Pharmacol Toxicol 1995; 35: 541-79.
    CrossrefPubMedGoogle Scholar
  • 9 North RA, Barnard EA. Nucleotide receptors. Curr Opin Neurobiol 1997; 7: 346-57.
    CrossrefPubMedGoogle Scholar
  • 10 Fredholm BB, Abbracchio MP, Burnstock G, Dubyak GR, Harden TK, Jacobson KA, Schwabe U, Williams M. Towards a revised nomenclature for P1 and P2 receptors. Trends Pharmacol Sci 1997; 18: 79-82.
    PubMedGoogle Scholar
  • 11 Leon C, Hechler B, Vial C, Leray C, Cazenave JP, Gachet C. The P2Y1 Receptor is an ADP Receptor Antagonized by ATP and Expressed in Platelets and Megakaryoblastic Cells. FEBS Lett 1997; 403: 26-30.
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  • 12 Gachet C, Hechler B, Leon C, Vial C, Leray C, Ohlmann P, Cazenave JP. Activation of ADP Receptors and Platelet Function. Thromb Haemost 1997; 78: 271-5.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 13 Hechler B, Vigne P, Leon C, Breittmeyer J-P, Gachet C, Frelin C. ATP Derivatives Are Antagonists of the P2Y1 Receptor: Similarities to the Platelet ADP Receptor. Mol Pharmacol 1998; 53: 727-33.
    CrossrefPubMedGoogle Scholar
  • 14 Daniel JL, Dangelmaier C, Jin J, Ashby B, Smith JB, Kunapuli SP. Molecular Basis for ADP-induced Platelet Activation. J Biol Chem 1998; 273: 2024-9.
    CrossrefPubMedGoogle Scholar
  • 15 Jin J, Daniel JL, Kunapuli SP. Molecular Basis for ADP-induced Platelet Activation. J Biol Chem 1998; 273: 2030-4
    CrossrefPubMedGoogle Scholar
  • 16 Savi P, Bornia J, Sabel V, Delfaud M, Herbert JM. Characterization of P2x1 Purinoceptors on Rat Platelets: Effect of Clopidogrel. Br J Haematol 1997; 98: 880-6.
    CrossrefPubMedGoogle Scholar
  • 17 Vial C, Hechler B, Leon C, Cazenave JP, Gachet C. Presence of P2X1 Purinoceptors in Human Platelets and Megakaryoblastic Cell Lines. Thromb Haemost 1997; 78: 1500-4.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 18 Fratantoni JC, Poindexter BJ. Measuring Platelet Aggregation with Micro-plate Reader. Am J Clin Pathol 1990; 94: 613-7.
    CrossrefPubMedGoogle Scholar
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    CrossrefPubMedGoogle Scholar
  • 20 Schachter JB, Li Q, Boyer JL, Nicholas RA, Harden TK. Second Messenger Cascade Specificity and Pharmacological Selectivity of the Human P2Y1-Purinoceptor. Br J Pharmacol 1996; 118: 167-73.
    CrossrefPubMedGoogle Scholar
  • 21 Grynkiewicz G, Poenie M, Tsien RY. A New Generation of Ca2+ Indicators with Greatly Improved Fluorescence Properties. J Biol Chem 1985; 260: 3440-50.
    PubMedGoogle Scholar
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    Google Scholar
  • 23 Michal F, Maguire MH, Gough G. 2-Methylthioadenosine-5’-phosphate: a Specific Inhibitor of Platelet Aggregation. Nature 1969; 222: 1073-4.
    CrossrefPubMedGoogle Scholar
  • 24 Cusack NJ, Hourani SMO. Specific but noncompetitive inhibition by 2-alkylthio analogues of adenosine 5’-monophosphate and adenosine 5’-triphosphate of human platelet aggregation induced by adenosine 5’-diphosphate. Br J Pharmacol 1982; 75: 397-400.
    CrossrefPubMedGoogle Scholar
  • 25 Boyer JL, Romero-Avila T, Schachter JB, Harden TK. Identification of Competitive Antagonists of the P2Y1 Receptor. Mol Pharmacol 1996; 50: 1323-9.
    PubMedGoogle Scholar
  • 26 Ohlmann P, Laugwitz KL, Nurnberg B. et al The human platelet ADP receptor activates Gi2 proteins. Biochem J 1995; 312: 775-9.
    CrossrefPubMedGoogle Scholar
  • 27 Fagura MS, Dainty IA, McKay GD, Kirk IP, Humphries RG, Robertson MJ, Dougall IG, Leff P. P2Y1-receptors in human platelets which are pharmacologically distinct from P2YADP-receptors. Br J Pharmacol 1998; 124: 157-64.
    CrossrefPubMedGoogle Scholar
  • 28 Offermanns S, Toombs CF, Hu YH, Simon I. M. Defective Platelet Activation in Gαq-deficient Mice. Nature 1997; 389: 183-6.
    CrossrefPubMedGoogle Scholar
  • 29 Schachter JB, Boyer JL, Li Q, Nicholas RA, Harden TK. Fidelity in Functional Coupling of the Rat P2Y1 Receptor to Phospholipase C. Br J Pharma-col 1997; 122: 1021-4.
    CrossrefPubMedGoogle Scholar
  • 30 Savi P, Beauverger P, Labouret C. et al Role of P2Y1 purinoceptor in ADP-induced platelet activation. FEBS Lett 1998; 422: 291-5.
    CrossrefPubMedGoogle Scholar
  • 31 McKenzie F. Basic techniques to study G-protein function. In: Signal Transduction: A Practical Approach. Milligan G. ed Oxford: IRL Press; 1992: 31-56.
    Google Scholar
  • 32 Savi P, Pflieger AM, Herbert JM. cAMP is not an Important Messenger for ADP-induced Platelet Aggregation. Blood Coagul Fibrinol 1996; 7: 249-52.
    CrossrefPubMedGoogle Scholar
  • 33 MacKenzie AB, Mahaut-Smith MP, Sage SO. Activation of Receptor-Operated Cation Channels via P2x1 Not P2T Purinoceptors in Human Platelets. J Biol Chem 1996; 271: 2879-81.
    CrossrefPubMedGoogle Scholar
  • 34 Scase TJ, Heath MF, Allen JM, Sage SO. Identification of a P2X1 Purinoceptor Expressed on Human Platelets. Biochem. Biophys Res Commun 1998; 242: 525-8.
    CrossrefPubMedGoogle Scholar
  • 35 Humphries RG, Robertson MJ, Leff P. A Novel Series of P2T Purinoceptor Antagonists: Definition of the Role of ADP in Arterial Thrombosis. Trends Pharmacol Sci 1995; 16: 179-81.
    CrossrefPubMedGoogle Scholar
  • 36 Cattaneo M, Lecchi A, Randi AM, McGregor JL, Mannucci PM. Identification of a New Congenital Defect of Platelet Function Characterized by Severe Impairment of Platelet Responses to Adenosine Diphosphate. Blood 1992; 80: 2787-96.
    PubMedGoogle Scholar
  • 37 Nurden P, Savi P, Heilmann E. et al An Inherited Bleeding Disorder Linked to a Defective Interaction between ADP and Its Receptor on Platelets. J Clin Invest 1995; 95: 1612-22.
    CrossrefPubMedGoogle Scholar
  • 38 Jin J, Kunapuli SP. Coactivation of two different G protein-coupled receptors is essential for ADP-induced platelet aggregation. Proc Natl Acad Sci USA 1998; 95: 8070-4.
    CrossrefPubMedGoogle Scholar
  • 39 Hechler B, Leon C, Vial C, Vigne P, Frelin C, Cazenave JP, Gachet C. The P2Y1 Receptor Is Necessary for Adenosine 5’-Diphosphate-Induced Platelet Aggregation. Blood 1998; 92: 152-9.
    PubMedGoogle Scholar

Correspondence to:

H.-M. Jantzen
COR Therapeutics, Inc., 256 E.
Grand Ave.
South San Francisco CA 94080 USA
Phone: +16502446831   
Fax: +16502449208   

  • References

  • 1 Savi P, Herbert JM. ADP Receptors on Platelets and ADP-Selective Antiaggregating Agents. Med Res Rev 1996; 16: 159-79.
    CrossrefPubMedGoogle Scholar
  • 2 Hourani SMO, Hall DA. Receptors for ADP on Human Blood Platelets. Trends Pharmacol Sci 1994; 15: 103-8.
    CrossrefPubMedGoogle Scholar
  • 3 Mills DCB. ADP Receptors on Platelets. Thromb Haemost 1996; 76: 835-6.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 4 Puri RN, Colman RW. ADP-Induced Platelet Activation. Crit Rev Biochem Mol Biol 1997; 3: 437-502.
    PubMedGoogle Scholar
  • 5 Boarder MR, Hourani SMO. The regulation of vascular function by P2 receptors: multiple sites and multiple receptors. Trends Pharmacol Sci 1998; 19: 99-107.
    CrossrefPubMedGoogle Scholar
  • 6 Sage SO. Calcium Entry Mechanisms in Human Platelets. Exp Physiol 1997; 82: 807-23.
    CrossrefPubMedGoogle Scholar
  • 7 Greco NJ, Yamamoto N, Jackson BW, Tandon NN, Moos M, Jamieson GA. Identification of a nucleotide-binding site on glycoprotein IIb. J Biol Chem 1991; 266: 13627-33.
    PubMedGoogle Scholar
  • 8 Harden TK, Boyer JL, Nicholas RA. P2 Purinergic Receptors: Subtype-Associated Signaling Responses and Structure. Annu Rev Pharmacol Toxicol 1995; 35: 541-79.
    CrossrefPubMedGoogle Scholar
  • 9 North RA, Barnard EA. Nucleotide receptors. Curr Opin Neurobiol 1997; 7: 346-57.
    CrossrefPubMedGoogle Scholar
  • 10 Fredholm BB, Abbracchio MP, Burnstock G, Dubyak GR, Harden TK, Jacobson KA, Schwabe U, Williams M. Towards a revised nomenclature for P1 and P2 receptors. Trends Pharmacol Sci 1997; 18: 79-82.
    PubMedGoogle Scholar
  • 11 Leon C, Hechler B, Vial C, Leray C, Cazenave JP, Gachet C. The P2Y1 Receptor is an ADP Receptor Antagonized by ATP and Expressed in Platelets and Megakaryoblastic Cells. FEBS Lett 1997; 403: 26-30.
    CrossrefPubMedGoogle Scholar
  • 12 Gachet C, Hechler B, Leon C, Vial C, Leray C, Ohlmann P, Cazenave JP. Activation of ADP Receptors and Platelet Function. Thromb Haemost 1997; 78: 271-5.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 13 Hechler B, Vigne P, Leon C, Breittmeyer J-P, Gachet C, Frelin C. ATP Derivatives Are Antagonists of the P2Y1 Receptor: Similarities to the Platelet ADP Receptor. Mol Pharmacol 1998; 53: 727-33.
    CrossrefPubMedGoogle Scholar
  • 14 Daniel JL, Dangelmaier C, Jin J, Ashby B, Smith JB, Kunapuli SP. Molecular Basis for ADP-induced Platelet Activation. J Biol Chem 1998; 273: 2024-9.
    CrossrefPubMedGoogle Scholar
  • 15 Jin J, Daniel JL, Kunapuli SP. Molecular Basis for ADP-induced Platelet Activation. J Biol Chem 1998; 273: 2030-4
    CrossrefPubMedGoogle Scholar
  • 16 Savi P, Bornia J, Sabel V, Delfaud M, Herbert JM. Characterization of P2x1 Purinoceptors on Rat Platelets: Effect of Clopidogrel. Br J Haematol 1997; 98: 880-6.
    CrossrefPubMedGoogle Scholar
  • 17 Vial C, Hechler B, Leon C, Cazenave JP, Gachet C. Presence of P2X1 Purinoceptors in Human Platelets and Megakaryoblastic Cell Lines. Thromb Haemost 1997; 78: 1500-4.
    Article in Thieme ConnectPubMedGoogle Scholar
  • 18 Fratantoni JC, Poindexter BJ. Measuring Platelet Aggregation with Micro-plate Reader. Am J Clin Pathol 1990; 94: 613-7.
    CrossrefPubMedGoogle Scholar
  • 19 Gachet C, Cazenave JP, Ohlmann P, Hilf G, Wieland T, Jakobs KH. ADP receptor-induced activation of guanine-nucleotide binding proteins in human platelet membranes. Eur J Biochem 1992; 207: 259-63.
    CrossrefPubMedGoogle Scholar
  • 20 Schachter JB, Li Q, Boyer JL, Nicholas RA, Harden TK. Second Messenger Cascade Specificity and Pharmacological Selectivity of the Human P2Y1-Purinoceptor. Br J Pharmacol 1996; 118: 167-73.
    CrossrefPubMedGoogle Scholar
  • 21 Grynkiewicz G, Poenie M, Tsien RY. A New Generation of Ca2+ Indicators with Greatly Improved Fluorescence Properties. J Biol Chem 1985; 260: 3440-50.
    PubMedGoogle Scholar
  • 22 Harlow E, Lane D. eds. Antibodies: a laboratory manual. Cold Spring Harbor NY: Cold Spring Harbor Laboratory; 1988
    Google Scholar
  • 23 Michal F, Maguire MH, Gough G. 2-Methylthioadenosine-5’-phosphate: a Specific Inhibitor of Platelet Aggregation. Nature 1969; 222: 1073-4.
    CrossrefPubMedGoogle Scholar
  • 24 Cusack NJ, Hourani SMO. Specific but noncompetitive inhibition by 2-alkylthio analogues of adenosine 5’-monophosphate and adenosine 5’-triphosphate of human platelet aggregation induced by adenosine 5’-diphosphate. Br J Pharmacol 1982; 75: 397-400.
    CrossrefPubMedGoogle Scholar
  • 25 Boyer JL, Romero-Avila T, Schachter JB, Harden TK. Identification of Competitive Antagonists of the P2Y1 Receptor. Mol Pharmacol 1996; 50: 1323-9.
    PubMedGoogle Scholar
  • 26 Ohlmann P, Laugwitz KL, Nurnberg B. et al The human platelet ADP receptor activates Gi2 proteins. Biochem J 1995; 312: 775-9.
    CrossrefPubMedGoogle Scholar
  • 27 Fagura MS, Dainty IA, McKay GD, Kirk IP, Humphries RG, Robertson MJ, Dougall IG, Leff P. P2Y1-receptors in human platelets which are pharmacologically distinct from P2YADP-receptors. Br J Pharmacol 1998; 124: 157-64.
    CrossrefPubMedGoogle Scholar
  • 28 Offermanns S, Toombs CF, Hu YH, Simon I. M. Defective Platelet Activation in Gαq-deficient Mice. Nature 1997; 389: 183-6.
    CrossrefPubMedGoogle Scholar
  • 29 Schachter JB, Boyer JL, Li Q, Nicholas RA, Harden TK. Fidelity in Functional Coupling of the Rat P2Y1 Receptor to Phospholipase C. Br J Pharma-col 1997; 122: 1021-4.
    CrossrefPubMedGoogle Scholar
  • 30 Savi P, Beauverger P, Labouret C. et al Role of P2Y1 purinoceptor in ADP-induced platelet activation. FEBS Lett 1998; 422: 291-5.
    CrossrefPubMedGoogle Scholar
  • 31 McKenzie F. Basic techniques to study G-protein function. In: Signal Transduction: A Practical Approach. Milligan G. ed Oxford: IRL Press; 1992: 31-56.
    Google Scholar
  • 32 Savi P, Pflieger AM, Herbert JM. cAMP is not an Important Messenger for ADP-induced Platelet Aggregation. Blood Coagul Fibrinol 1996; 7: 249-52.
    CrossrefPubMedGoogle Scholar
  • 33 MacKenzie AB, Mahaut-Smith MP, Sage SO. Activation of Receptor-Operated Cation Channels via P2x1 Not P2T Purinoceptors in Human Platelets. J Biol Chem 1996; 271: 2879-81.
    CrossrefPubMedGoogle Scholar
  • 34 Scase TJ, Heath MF, Allen JM, Sage SO. Identification of a P2X1 Purinoceptor Expressed on Human Platelets. Biochem. Biophys Res Commun 1998; 242: 525-8.
    CrossrefPubMedGoogle Scholar
  • 35 Humphries RG, Robertson MJ, Leff P. A Novel Series of P2T Purinoceptor Antagonists: Definition of the Role of ADP in Arterial Thrombosis. Trends Pharmacol Sci 1995; 16: 179-81.
    CrossrefPubMedGoogle Scholar
  • 36 Cattaneo M, Lecchi A, Randi AM, McGregor JL, Mannucci PM. Identification of a New Congenital Defect of Platelet Function Characterized by Severe Impairment of Platelet Responses to Adenosine Diphosphate. Blood 1992; 80: 2787-96.
    PubMedGoogle Scholar
  • 37 Nurden P, Savi P, Heilmann E. et al An Inherited Bleeding Disorder Linked to a Defective Interaction between ADP and Its Receptor on Platelets. J Clin Invest 1995; 95: 1612-22.
    CrossrefPubMedGoogle Scholar
  • 38 Jin J, Kunapuli SP. Coactivation of two different G protein-coupled receptors is essential for ADP-induced platelet aggregation. Proc Natl Acad Sci USA 1998; 95: 8070-4.
    CrossrefPubMedGoogle Scholar
  • 39 Hechler B, Leon C, Vial C, Vigne P, Frelin C, Cazenave JP, Gachet C. The P2Y1 Receptor Is Necessary for Adenosine 5’-Diphosphate-Induced Platelet Aggregation. Blood 1998; 92: 152-9.
    PubMedGoogle Scholar

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