Download PDF
Thromb Haemost 1982; 47(02): 090-095
DOI: 10.1055/s-0038-1657136
Original Article
Schattauer GmbH Stuttgart

Effect of cAMP Phosphodiesterase Inhibitors on ADP-Induced Shape Change, cAMP and Nucleoside Diphosphokinase Activity of Rabbit Platelets

S C-T Lam
The Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada and the Department of Pathology, McMaster University, Hamilton, Ontario, Canada
,
M A Guccione
The Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada and the Department of Pathology, McMaster University, Hamilton, Ontario, Canada
,
M A Packham
The Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada and the Department of Pathology, McMaster University, Hamilton, Ontario, Canada
,
J F Mustard
The Department of Biochemistry, University of Toronto, Toronto, Ontario, Canada and the Department of Pathology, McMaster University, Hamilton, Ontario, Canada
› Author Affiliations
Further Information

Publication History

Received 05 November 1981

Accepted 18 January 1982

Publication Date:
13 July 2018 (online)

Also available at
  PDF Download  Permissions and Reprints

Summary

The effects of cAMP phosphodiesterase inhibitors on ADP-induced shape change and cAMP concentrations have been studied. Caffeine (10 mM), theophylline (8 mM), dipyridamole (0.2 mM), or papaverine (0.05 mM) prevented the shape change of washed rabbit platelets induced by 0.4 μM ADP. At these concentrations, none of these cAMP phosphodiesterase inhibitors increased 14C-cAMP in platelets in which the cytoplasmic adenine nucleotides had been labelled with 14C-adenine. By a protein binding assay, only papaverine by itself increased platelet cAMP above its basal level. These results indicate that two pools of cAMP may exist in platelets. Both methods showed that stimulation of platelet adenylate cyclase with PGE1 (1 μM) resulted in an increase in platelet cAMP and all these cAMP phosphodiesterase inhibitors potentiated this increase caused by PGE1. By themselves, some of these compounds may act through mechanisms that do not involve platelet cAMP. The effects of these cAMP phosphodiesterase inhibitors on platelet nucleoside diphosphokinase (NDK) activity were also investigated. At concentrations that prevented ADP-induced shape change, papaverine and dipyridamole had no effect on the formation of 14C-ATP from 14C-ADP by washed rabbit platelets. The methylxanthines partially inhibited NDK activity of washed rabbit platelets and of isolated platelet membranes, probably due to the structural similarity between the adenine ring of ADP and these substances. However, adenine (8 mM) inhibited ADP-induced shape change and platelet NDK activity but was a less effective inhibitor of ADP-induced platelet aggregation. Thus it seems unlikely that interference with platelet NDK or the ADP receptor is the major mechanism by which the methylxanthines inhibit platelet functions.

Key words

Platelets - cAMP phosphodiesterase inhibitors - Dipyridamole - Adenine - Nucleoside diphosphokinase
 

  • References

  • 1 Mills DCB, Smith JB. The influence on platelet aggregation of drugs that affect the accumulation of adenosine 3':5'-cyclic monophosphate in platelets. Biochem J 1971; 121: 185-196
    CrossrefPubMedGoogle Scholar
  • 2 Salzman EW. Cyclic AMP and platelet function. N Engl J Med 1972; 286: 358-363
    CrossrefPubMedGoogle Scholar
  • 3 Haslam RJ. Roles of cyclic nucleotides in platelet function. In: Biochemistry and Pharmacology of Platelets. Ciba Foundation Symposium 35. 121-151 (new series) Elsevier Excerpta Medica; North Holland: 1975
    Google Scholar
  • 4 Holmsen H. Classification and possible mechanisms of action of some drugs that inhibit platelet aggregation. Ser Haematol 1976; 8: 50-80
    PubMedGoogle Scholar
  • 5 Haslam RJ, Davidson MML, Davies T, Lynham JA, McClenaghan MD. Regulation of blood platelet function by cyclic nucleotides. Adv Cyclic Nucleotide Res 1978; 9: 533-552
    PubMedGoogle Scholar
  • 6 Marquis NR, Vigdahl RL, Tavormina PA. Platelet aggregation. I. Regulation by cyclic AMP and prostaglandin E1 . Biochem Biophys Res Comm 1969; 36: 965-972
    CrossrefPubMedGoogle Scholar
  • 7 Wolfe SM, Shulman NR. Adenyl cyclase activity in human platelets. Biochem Biophys Res Comm 1969; 35: 265-272
    CrossrefPubMedGoogle Scholar
  • 8 Haslam RJ. Interactions of the pharmacological receptors of blood platelets with adenylate cyclase. Ser Haematol 1973; 6: 333-350
    PubMedGoogle Scholar
  • 9 McDonald JWD, Stuart RK. Regulation of cyclic-AMP levels and aggregation in human platelets by prostaglandin E1 . J Lab Clin Med 1973; 81: 838-849
    PubMedGoogle Scholar
  • 10 Salzman EW. Prostaglandins, cyclic AMP, and platelet function. Thrombos Diathes Haemorrh 1974; (Suppl. 60) 311-319
    PubMedGoogle Scholar
  • 11 Michel H, Caen JP, Born GVR, Miller R, D’Auriac GA, Meyer P. Relation between the inhibition of aggregation and the concentration of cAMP in human and rat platelets. Br J Haematol 1976; 33: 27-38
    CrossrefPubMedGoogle Scholar
  • 12 Best LC, Martin TJ, Russel RGG, Preston FE. Prostacyclin increases cyclic AMP levels and adenylate cyclase activity in platelets. Nature 1977; 267: 850-852
    CrossrefPubMedGoogle Scholar
  • 13 Gorman RR, Bunting S, Miller OV. Modulation of human platelet adenylate cyclase by prostacyclin (PGX). Prostaglandins 1977; 13: 377-388
    CrossrefPubMedGoogle Scholar
  • 14 Tateson JE, Moncada S, Vane JR. Effects of prostacyclin (PGX) on cyclic AMP concentrations in human platelets. Prostaglandins 1977; 13: 389-397
    CrossrefPubMedGoogle Scholar
  • 15 Ardlie NG, Glew G, Schultz BG, Schwartz CJ. Inhibition and reversal of platelet aggregation by methyl xanthines. Thrombos Diathes Haemorrh 1967; 18: 670-673
    PubMedGoogle Scholar
  • 16 Ball G, Brereton GG, Fulwood M, Ireland DM, Yates P. Effect of prostaglandin E1 alone and in combination with theophylline or aspirin on collagen-induced platelet aggregation and on platelet nucleotides including adenosine 3' 5'-cyclic monophosphate. Biochem J 1970; 120: 709-718
    CrossrefPubMedGoogle Scholar
  • 17 Cucuianu MP, Nishizawa EE, Mustard JF. Effect of pyrimido-pyrimidine compounds on platelet function. J Lab Clin Med 1971; 77: 958-974
    PubMedGoogle Scholar
  • 18 Niewiarowski S, Lukasiewicz H, Nath N, Sha AT. Inhibition of human platelet aggregation by dipyridamole and two related compounds and its modification by add glycoproteins of human plasma. J Lab Clin Med 1975; 86: 64-76
    PubMedGoogle Scholar
  • 19 Subbarao K, Rucinski B, Niewiarowski S. Effect of dipyridamole and its monoglucuronide derivative on adenosine uptake by human platelets and ADP-induced platelet aggregation. Biochem Pharmacol 1977; 26: 906-907
    CrossrefPubMedGoogle Scholar
  • 20 Best LC, McGuire MB, Jones PBB, Holland TK, Martin TJ, Preston FE, Segal DS, Russell RGG. Mode of action of dipyridamole on human platelets. Thromb Res 1979; 16: 367-379
    CrossrefPubMedGoogle Scholar
  • 21 Tang SS, Frojmovic MM. Inhibition of platelet function by antithrombotic agents which selectively inhibit low-Km cyclic 3', 5'-adenosine monophosphate phosphodiesterase. J Lab Clin Med 1980; 95: 241-257
    PubMedGoogle Scholar
  • 22 Guccione MA, Packham MA, Kinlough-Rathbone RL, Mustard JF. Reactions of 14C-ADP and 14C-ATP with washed platelets from rabbits. Blood 1971; 37: 542-555
    PubMedGoogle Scholar
  • 23 Feinberg H, Belamarich F, Born GVR. Transphosphorylation in human and rabbit platelet suspensions. Fed Proc 1973; 32: 220
    PubMedGoogle Scholar
  • 24 Mustard JF, Packham MA, Perry DW, Guccione MA, Kinlough-Rathbone RL. Enzyme activities on the platelet surface in relation to the action of adenosine diphosphate. In: Biochemistry and Pharmacology of Platelets. Ciba Foundation Symposium 35. 47-75 (new series) Elsevier Excerpta Medica; North Holland: 1975
    Google Scholar
  • 25 Packham MA, Guccione MA, Perry DW, Kinlough-Rathbone RL, Mustard JF. AMP inhibition of reactions of ADP with washed platelets from human and rabbits. Am J Physiol 1972; 223: 419-424
    CrossrefPubMedGoogle Scholar
  • 26 Lips JPM, Sixma JJ, Schiphorst ME. Binding of adenosine diphosphate to human blood platelets and to isolated blood platelet membranes. Biochim Biophys Acta 1980; 628: 451-467
    CrossrefPubMedGoogle Scholar
  • 27 Kinlough-Rathbone RL, Packham MA, Mustard JF. The effect of prostaglandin E1 on platelet function in vitro and in vivo. Br J Haematol 1970; 19: 559-571
    CrossrefPubMedGoogle Scholar
  • 28 Aster RH, Jandl JH. Platelet sequestration in man. I. Methods. J Clin Invest 1964; 43: 843-855
    CrossrefPubMedGoogle Scholar
  • 29 Ardlie NG, Packham MA, Mustard JF. Adenosine diphosphate-induced platelet aggregation in suspensions of washed rabbit platelets. Br J Haematol 1970; 19: 7-17
    CrossrefPubMedGoogle Scholar
  • 30 Ardlie NG, Perry DW, Packham MA, Mustard JF. Influence of apyrase on stability of suspensions of washed rabbit platelets. Proc Soc Exptl Biol Med 1971; 136: 1021-1023
    CrossrefPubMedGoogle Scholar
  • 31 Molnar J, Lorand L. Studies on apyrases. Arch Biochem Biophys 1961; 93: 353-363
    CrossrefPubMedGoogle Scholar
  • 32 Nurden AT, Caen JP. Specific roles for platelet surface glycoproteins in platelet function. Nature 1975; 255: 720-722
    CrossrefPubMedGoogle Scholar
  • 33 Lowry OH, Rosebrough NJ, Farr AL, Randall RJ. Protein measurement with the Folin phenol reagent. J Biol Chem 1951; 193: 265-275
    PubMedGoogle Scholar
  • 34 Mustard JF, Packham MA. Platelet aggregation and the platelet release reaction in thromboembolism. Can Med Assoc J 1970; 103: 859-863
    PubMedGoogle Scholar
  • 35 Lam SC-T, Harfenist EJ, Packham MA, Mustard JF. Investigation of possible mechanisms of pyridoxal 5'-phosphate inhibition of platelet reactions. Thromb Res 1980; 20: 633-645
    CrossrefPubMedGoogle Scholar
  • 36 Haslam RJ, Rosson GM. Effects of adenosine on levels of adenosine cyclic 3',5'-monophosphate in human blood platelets in relation to adenosine incorporation and platelet aggregation. Mol Pharmacol 1975; 11: 528-544
    PubMedGoogle Scholar
  • 37 Brown BL, Albano JDM, Ekins RP, Sgherzi AM. A simple and sensitive saturation assay method for the measurement of adenosine 3':5'-cyclic monophosphate. Biochem J 1971; 121: 561-562
    CrossrefPubMedGoogle Scholar
  • 38 McElroy FA, Philp RB. Relative potendes of dipyridamole and related agents as inhibitors of cyclic nucleotide phosphodiesterases: Possible explanation of mechanism of inhibition of platelet function. Life Sci 1975; 17: 1479-1494
    CrossrefPubMedGoogle Scholar
  • 39 Radomski M, Swies J, Gryglewski RJ. Disaggregatory action of phosphodiesterase inhibitors. Pharmacol Res Comm 1981; 13: 41-47
    CrossrefPubMedGoogle Scholar
  • 40 Adler JR, Handin RI. Solubilization and characterization of a platelet membrane ADP-binding protein. J Biol Chem 1979; 254: 3866-3872
    PubMedGoogle Scholar
  • 41 Cooper DMF, Rodbell M. ADP is a potent inhibitor of human platelet plasma membrane adenylate cyclase. Nature 1979; 282: 517-518
    CrossrefPubMedGoogle Scholar
  • 42 Markwardt F, Hoffmann A. Effects of papaverine derivatives on cyclic AMP phosphodiesterase of human platelets. Biochem Pharmacol 1970; 19: 2519-2520
    CrossrefPubMedGoogle Scholar
  • 43 Vigdahl RL, Mongin J Jr, Marquis NR. Platelet aggregation. IV. Platelet phosphodiesterase and its inhibition by vasodilators. Biochem Biophys Res Comm 1971; 42: 1088-1094
    CrossrefPubMedGoogle Scholar
  • 44 Rozenberg MC, Walker CM. The effect of pyrimidine compounds on the potentiation of adenosine inhibition of aggregation, on adenosine phosphorylation and phosphodiesterase activity of blood platelets. Br J Haematol 1973; 24: 409-418
    CrossrefPubMedGoogle Scholar
  • 45 Asano T, Ochiai Y, Hidaka A. Selective inhibition of separated forms of human platelet cyclic nucleotide phosphodiesterase by platelet aggregation inhibitors. Mol Pharmacol 1977; 13: 400-406
    PubMedGoogle Scholar
  • 46 Tsien WH, Sheppard H. The lack of correlation between inhibition of platelet aggregation and cAMP levels. Fed Proc 1981; 40: 809
    PubMedGoogle Scholar
  • 47 Zahavi M, Kakkar VV. Adenosine 3',5'-cyclic monophosphate (c-AMP) and inhibitors of platelet aggregation: A possibility of a double c-AMP pool in human platelets. Thromb Haemostas 1981; 46: 415 (Abstr)
    PubMedGoogle Scholar
  • 48 Amer MS, Mayol RF. Studies with phosphodiesterase. III. Two forms of the enzyme from human blood platelets. Biochim Biophys Acta 1973; 309: 149-156
    CrossrefPubMedGoogle Scholar
  • 49 Pichard A-L, Hanoune J, Kaplan J-C. Multiple forms of cyclic adenosine 3',5'-monophosphate phosphodiesterase from human blood platelets. I. Kinetic and electrophoretic characterization of two molecular spedes. Biochim Biophys Acta 1973; 315: 370-377
    CrossrefPubMedGoogle Scholar
  • 50 Hidaka H, Asano T. Human blood platelet 3',5'-cyclic nucleotide phosphodiesterase. Isolation of low-Km and high-Km phosphodiesterase. Biochim Biophys Acta 1976; 429: 485-497
    CrossrefPubMedGoogle Scholar
  • 51 Lecomte M-C, Galand C, Boivin P. Inhibition of human erythrocyte casein kinase by methylxanthines. FEBS Lett 1980; 116: 45-47
    CrossrefPubMedGoogle Scholar