Cyclic Nucleotides and Phosphodiesterases in Platelets

Introduction

It is now almost 30 years since the discovery that prostaglandin E₁ (PGE₁) inhibits platelet responses to aggregating agents, together with finding that the effects of this compound are mediated by adenosine 3′, 5′-cyclic monophosphate (cAMP) initiated interest in the physiological and pharmacological regulation of platelet function by other agents that increase platelet cAMP, as reviewed elsewhere.¹ The most important agonists that stimulate cAMP formation in platelets have now been identified as prostacyclin (PGI₂), prostaglandin D₂ (PGD₂), and adenosine, which exert their effects through receptors of the serpentine or seven transmembrane segment class (IP, DP and A₂ receptors, respectively).² The latter then stimulate cAMP formation by adenylyl cyclase via the GTP-dependent activation of the G-protein, G_s (Fig. 1). In the classical view, cAMP exerts its effects solely by binding to the RI and RII regulatory subunits of type I and type II cAMP-dependent protein kinases (PKA). The catalytic subunits of the kinases then dissociate and phosphorylate selected serine and threonine residues on target proteins that prevent or reverse platelet activation.² A crucial role is played by cAMP phosphodiesterases, which degrade cAMP to 5′-AMP, thereby diminishing and terminating the effects of agonists that stimulate cAMP formation (Fig. 1). In early studies, this was demonstrated by the ability of first-generation inhibitors of cAMP phosphodiesterases, particularly the methylxanthines, to inhibit platelet aggregation and potentiate the inhibitory effects of activators of platelet adenylyl cyclase.¹ Such studies provided the rationale for the subsequent development of more potent and selective phosphodiesterase inhibitors as potential antithrombotic agents.

Interest in the role of guanosine 3′,5′-cyclic monophosphate (cGMP) in platelets closely followed the discovery of the inhibitory action of cAMP. An early hypothesis that cGMP might potentiate platelet aggregation was abandoned by 1978, after it was shown that some inhibitors of platelet aggregation, such as nitroprusside (NP), also increased platelet cGMP.¹ It soon emerged that all nitrovasodilators release nitric oxide and activate soluble guanylyl cyclase (GC) and that the cGMP formed stimulates cGMP-dependent protein kinases (PKG) in many cells and tissues (Fig. 1), including vascular smooth muscle and platelets.³ The crucial physiological importance of this pathway was established with the identification of endothelium-derived relaxing factor (EDRF) as nitric oxide.⁴ cGMP phosphodiesterases play an essential role by limiting increases in cellular cGMP, and inhibition of these enzymes was found to potentiate the effects of nitric oxide and nitric oxide donors on platelets and other cells.⁵

The ability of cAMP and cGMP to activate distinct protein kinases led to a persistent view that these two cyclic nucleotides operate in parallel and independent ways to inhibit platelet function, cAMP mediating the effects of agonists such as PGI₂, and cGMP mediating the effects of nitric oxide.^2,3 However, over the last 10 years, considerable evidence has accumulated to indicate that this is not the case in platelets (or in many other cells) and that cross-talk between the cAMP and cGMP systems may occur on at least two levels, affecting both cyclic nucleotide phosphodiesterase (PDE) and protein kinase activities (Fig. 1). One of the most significant of these interactions is through the effects of cGMP on the hydrolysis of cAMP by PDEs. It is the purpose of this chapter to describe platelet PDEs and to discuss how their individual characteristics and regulation may impact platelet function and the design of useful antithrombotic agents. In addition, evidence that both cGMP and cAMP may activate PKG and that these cyclic nucleotides may exert effects in platelets that do not involve either PKA or PKG will be discussed briefly.

• References

• 1 Haslam RJ, Davidson MML, Davies T, Lynham JA, McClenaghan MD. Regulation of blood platelet function by cyclic nucleotides. In: Advances in Cyclic Nucleotide Research. vol. 9. George WJ, Ignarro LJ. eds. New York: Raven Press; 1978: 533-552.
• 2 Blockmans D, Deckmyn H, Vermylen J. Platelet activation. Blood Rev 1995; 9: 143-156.
• 3 Schmidt HHHW, Lohmann SM, Walter U. The nitric oxide and cGMP signal transduction system: regulation and mechanism of action. Biochim Biophys Acta 1993; 1178: 153-175.
• 4 Palmer RMJ, Ferrige AG, Moncada S. Nitric oxide release accounts for the biological activity of endothelium-derived relaxing factor. Nature 1987; 327: 524-526.
• 5 Thompson WJ. Cyclic nucleotide phosphodiesterases: pharmacology, biochemistry and function. Pharmacol Ther 1991; 51: 13-33.
• 6 Beavo JA. Cyclic nucleotide phosphodiesterases: functional implications of multiple isoforms. Physiol Rev 1995; 75: 725-748.
• 7 Perry MJ, Higgs GA. Chemotherapeutic potential of phosphodiesterase inhibitors. Curr Opin Chem Biol 1998; 2: 472-481.
• 8 Simpson AWM, Reeves ML, Rink TJ. Effects of SK&F 94120, an inhibitor of cyclic nucleotide phosphodiesterase type III, on human platelets. Biochem Pharmacol 1988; 37: 2315-2320.
• 9 Weishaar RE, Burrows SD, Kobylarz DC, Quade MM, Evans DB. Multiple molecular forms of cyclic nucleotide phosphodiesterase in cardiac and smooth muscle and in platelets. Isolation, characterization, and effects of various reference phosphodiesterase inhibitors and cardiotonic agents. Biochem Pharmacol 1986; 35: 787-800.
• 10 Ito M, Nishikawa M, Fujioka M, Miyahara M, Isaka N, Shiku H, Nakano T. Characterization of the isoenzymes of cyclic nucleotide phosphodiesterase in human platelets and the effects of E4021. Cell Signalling 1996; 8: 575-581.
• 11 Martins TJ, Mumby MC, Beavo JA. Purification and characterization of a cyclic GMP-stimulated cyclic nucleotide phosphodiesterase from bovine tissues. J Biol Chem 1982; 257: 1973-1979.
• 12 Stroop SD, Beavo JA. Structure and function studies of the cGMP-stimulated phosphodiesterase. J Biol Chem 1991; 266: 23802-23809.
• 13 MacFarland RT, Zelus BD, Beavo JA. High concentrations of a cGMP-stimulated phosphodiesterase mediate ANP-induced decreases in cAMP and steroidogenesis in adrenal glomerulosa cells. J Biol Chem 1991; 266: 136-142.
• 14 Méry P-F, Pavoine C, Belhassen L, Pecker F, Fischmeister R. Nitric oxide regulates cardiac Ca²⁺ current. Involvement of cGMP-inhibited and cGMP-stimulated phosphodiesterases through guanylyl cyclase activation. J Biol Chem 1993; 268: 26286-26295.
• 15 Yang Q, Paskind M, Bolger G, Thompson WJ, Repaske DR, Cutler LS, Epstein PM. A novel cyclic GMP stimulated phosphodiesterase from rat brain. Biochem Biophys Res Commun 1994; 205: 1850-1858.
• 16 Podzuweit T, Nennstiel P, Müller A. Isozyme selective inhibition of cGMP-stimulated cyclic nucleotide phosphodiesterases by erythro-9-(2-hydroxy-3-nonyl)adenine. Cell Signaling 1995; 7: 733-738.
• 17 Degerman E, Belfrage P, Manganiello VC. Structure, localization, and regulation of cGMP-inhibited phosphodiesterase (PDE3). J Biol Chem 1997; 272: 6823-6826.
• 18 Maurice DH, Haslam RJ. Molecular basis of the synergistic inhibition of platelet function by nitrovasodilators and activators of adenylate cyclase: inhibition of cyclic AMP breakdown by cyclic GMP. Mol Pharmacol 1990; 37: 671-681.
• 19 Maurice DH, Haslam RJ. Nitroprusside enhances isoprenaline-induced increases in cAMP in rat aortic smooth muscle. Eur J Pharmacol 1990; 191: 471-475.
• 20 Meacci E, Taira M, Moos M, Smith CJ, Movsesian MA, Degerman E, Belfrage P, Manganiello V. Molecular cloning and expression of human myocardial cGMP-inhibited cAMP phosphodiesterase. Proc Natl Acad Sci USA 1992; 89: 3721-3725.
• 21 Liu H, Maurice DH. Expression of cyclic GMP-inhibited phosphodiesterases 3A and 3B (PDE3A and PDE3B) in rat tissues: Differential subcellular localization and regulated expression by cyclic AMP. Br J Pharmacol 1998; 125: 1501-1510.
• 22 Kasuya J, Goko H, Fujita-Yamaguchi Y. Multiple transcripts for the human cardiac form of the cGMP-inhibited cAMP phosphodiesterase. J Biol Chem 1995; 270: 14305-14312.
• 23 Meanwell NA. Inhibitors of platelet phosphodiesterase: Potential antithrombotic agents. Drug News Perspect 1991; 4: 400-413.
• 24 Hidaka H, Hayashi H, Kohri H, Kimura Y, Hosokawa T, Igawa T, Saitoh Y. Selective inhibitor of platelet cyclic adenosine monophosphate phosphodiesterase, cilostamide, inhibits platelet aggregation. J Pharmacol Exp Ther 1979; 211: 26-30.
• 25 Ochiai M, Isshiki T, Takeshita S, Eto K, Toyoizumi H, Sato T, Miyashita H. Use of cilostazol, a novel antiplatelet agent, in a post-Palmaz-Schatz stenting regimen. Am J Cardiol 1997; 79: 1471-1474.
• 26 Dawson DL, Cutler BS, Meissner MH, Strandness DE. Cilostazol has beneficial effects in treatment of intermittent claudication. Circulation 1998; 98: 678-686.
• 27 Fischer TA, Erbel R, Treese N. Current status of phosphodiesterase inhibitors in the treatment of congestive heart failure. Drugs 1992; 44: 928-945.
• 28 Packer M, Carver JR, Rodeheffer RJ, Ivanhoe RJ, DiBianco R, Zeldis SM, Hendrix GH, Bommer WJ, Elkayam U, Kukin ML. et al. for the PROMISE Study Research Group. Effects of oral milrinone on mortality in severe chronic heart failure. N Engl J Med 1991; 325: 1468-1475.
• 29 Feldman AM, Bristow MR, Parmley WW, Carson PE, Pepine CJ, Gilbert EM, Strobeck JE, Hendrix GH, Powers ER, Bain RP, White BG. Effects of vesnarinone on morbidity and mortality in patients with heart failure. N Eng J Med 1993; 329: 149-155.
• 30 Alvarez R, Banerjee GL, Bruno JJ, Jones GL, Liittschwager K, Strosberg AM, Venuti MC. A potent and selective inhibitor of cyclic inhibitor of cyclic AMP phosphodiesterase with potential cardiotonic and antithrombotic properties. Mol Pharmacol 1986; 29: 554-560.
• 31 Thomas MK, Francis SH, Corbin JD. Characterization of a purified bovine lung cGMP-binding cGMP phosphodiesterase. J Biol Chem 1990; 265: 14964-14970.
• 32 Burns F, Rodger IW, Pyne NJ. The catalytic subunit of protein kinase A triggers activation of the type V cyclic GMP-specific phosphodiesterase from guinea-pig lung. Biochem J 1992; 283: 487-491.
• 33 Loughney K, Hill TR, Florio VA, Uher L, Rosman GJ, Wolda SL, Jones BA, Howard ML, McAllister-Lucas LM, Sonnenburg WK, Francis SH, Corbin JD, Beavo JA, Ferguson K. Isolation and characterization of cDNAs encoding PDE5A, a human cGMP-binding, cGMP-specific 3′,5′-cyclic nucleotide phosphodiesterase. Gene 1998; 216: 139-147.
• 34 Turko IV, Francis SH, Corbin JD. Binding of cGMP to both allosteric sites of cGMP-binding cGMP-specific phosphodiesterase (PDE5) is required for its phosphorylation. Biochem J 1998; 329: 505-510.
• 35 Lochhead A, Nekrasova E, Arshavsky VY, Pyne NJ. The regulation of the cGMP-binding cGMP phosphodiesterase by proteins that are immunologically related to γ subunit of the photoreceptor cGMP phosphodiesterase. J Biol Chem 1997; 272: 18397-18403.
• 36 Han P, Zhu X, Michaeli T. Alternative splicing of the high affinity cAMP-specific phosphodiesterase (PDE7A) mRNA in human skeletal muscle and heart. J Biol Chem 1997; 272: 16152-16157.
• 37 Soderling SH, Bayuga SJ, Beavo JA. Cloning and characterization of a cAMP-specific cyclic nucleotide phosphodiesterase. Proc Natl Acad Sci USA 1998; 95: 8991-8996.
• 38 Soderling SH, Bayuga SJ, Beavo JA. Identification and characterization of a novel family of cyclic nucleotide phosphodiesterases. J Biol Chem 1998; 273: 15553-15558.
• 39 Fisher DA, Smith JF, Pillar St JS, Denis SH, Cheng JB. Isolation and characterization of PDE9A, a novel human cGMP-specific phosphodiesterase. J Biol Chem 1998; 273: 15559-15564.
• 40 Hidaka H, Asano T. Human blood platelet 3′:5′-cyclic nucleotide phosphodiesterase. Isolation of low-K _m and high-K _m phosphodiesterase. Biochim Biophys Acta 1976; 429: 485-497.
• 41 Hagiwara M, Endo T, Kanayama T, Hidaka H. Effect of 1-(3-chloroanilino)-4-phenylphthalazine (MY-5445), a specific inhibitor of cyclic GMP phosphodiesterase, on human platelet aggregation. J Pharmacol Exp Ther 1984; 228: 467-471.
• 42 Dickinson NT, Jang EK, Haslam RJ. Activation of cGMP-stimulated phosphodiesterase by nitroprusside limits cAMP accumulation in human platelets: effects on platelet aggregation. Biochem J 1997; 323: 371-377.
• 43 Grant PG, Colman RW. Purification and characterization of a human platelet cyclic nucleotide phosphodiesterase. Biochemistry 1984; 23: 1801-1807.
• 44 Harrison SA, Reifsnyder DH, Gallis B, Cadd GG, Beavo JA. Isolation and characterization of bovine cardiac muscle cGMP-inhibited phosphodiesterase: A a receptor for new cardiotonic drugs. Mol Pharmacol 1986; 29: 506-514.
• 45 Macphee CH, Harrison SA, Beavo JA. Immunological identification of the major platelet low-K _m cAMP phosphodiesterase: probable target for anti-thrombotic agents. Proc Natl Acad Sci USA 1986; 83: 6660-6663.
• 46 Degerman E, Moos M, Rascón A, Vasta V, Meacci E, Smith CJ, Lindgren S, Andersson K-E, Belfrage P, Manganiello V. Single-step affinity purification, partial structure and properties of human platelet cGMP inhibited cAMP phosphodiesterase. Biochim Biophys Acta 1994; 1205: 189-198.
• 47 Leroy M-J, Degerman E, Taira M, Murata T, Wang LH, Movsesian MA, Meacci E, Manganiello VC. Characterization of two recombinant PDE3 (cGMP-inhibited cyclic nucleotide phosphodiesterase) isoforms, RcGIP1 and HcGIP2, expressed in NIH 3006 murine fibroblasts and Sf9 insect cells. Biochemistry 1996; 35: 10194-10202.
• 48 Cheung PP, Xu H, McLaughlin MM, Ghazaleh FA, Livi GP, Colman RW. Human platelet cGI-PDE: Expression in yeast and localization of the catalytic domain by deletion mutagenesis. Blood 1996; 88: 1321-1329.
• 49 Tang KM, Jang EK, Haslam RJ. Expression and mutagenesis of the catalytic domain cGMP-inhibited phosphodiesterase (PDE3) cloned from human platelets. Biochem J 1997; 323: 217-224.
• 50 Pillai R, Staub SF, Colicelli J. Mutational mapping of kinetic and pharmacological properties of a human cardiac cAMP phosphodiesterase. J Biol Chem 1994; 269: 30676-30681.
• 51 Masuoka H, Ito M, Sugioka M, Kozeki H, Konishi T, Tanaka T, Nakano T. Two isoforms of cGMP-inhibited cyclic nucleotide phosphodiesterases in human tissues distinguished by their responses to vesnarinone, a new cardiotonic agent. Biochem Biophys Res Commun 1993; 190: 412-417.
• 52 Omburo GA, Brickus T, Ghazaleh FA, Colman RW. Divalent metal cation requirement and possible classification of cGMP-inhibited phosphodiesterase as a metallohydrolase. Arch Biochem Biophys 1995; 323: 1-5.
• 53 Ghazaleh FA, Omburo GA, Colman RW. Evidence for the presence of essential histidine and cysteine residues in platelet cGMP-inhibited phosphodiesterase. Biochem J 1996; 317: 495-501.
• 54 Cheung PP, Yu L, Zhang H, Colman RW. Partial characterization of the active site of human platelet cAMP phosphodiesterase, PDE3A, by site-directed mutagenesis. Arch Biochem Biophys 1998; 360: 99-104.
• 55 Butt E, Beltman J, Becker DE, Jensen GS, Rybalkin SD, Jastorff B, Beavo JA. Characterization of cyclic nucleotide phosphodiesterases with cyclic AMP analogs: Topology of the catalytic sites and comparison with other cyclic AMP-binding proteins. Mol Pharmacol 1995; 47: 340-347.
• 56 Erhardt PW, Hagedorn AA, Sabio M. Cardiotonic agents. 3. A topographical model of the cardiac cAMP phosphodiesterase receptor. Mol Pharmacol 1988; 33: 1-13.
• 57 Venuti MC, Stephenson RA, Alvarez R, Bruno JJ, Strosberg AM. Inhibitors of cyclic AMP phosphodiesterase. 3. Synthesis and biological evaluation of pyrido and imidazolyl analogues of 1,2,3,5-tetrahydro-2-oxoimidazo[2,1-b]quinazoline. J Med Chem 1988; 31: 2136-2145.
• 58 Macphee CH, Reifsnyder DH, Moore TA, Lerea KM, Beavo JA. Phosphorylation results in activation of a cAMP phosphodiesterase in human platelets. J Biol Chem 1988; 263: 10353-10358.
• 59 Grant PG, Mannarino AF, Colman RW. cAMP-mediated phosphorylation of the low-K _m cAMP phosphodiesterase markedly stimulates its catalytic activity. Proc Natl Acad Sci USA 1988; 85: 9071-9075.
• 60 Sheth SB, Brennan KJ, Biradavolu R, Colman RW. Isolation and regulation of the cGMP-inhibited cAMP phosphodiesterase in human erythroleukemia cells. Thromb Haemost 1997; 77: 155-162.
• 61 López-Aparicio P, Rascón A, Manganiello VC, Andersson K-E, Belfrage P, Degerman E. Insulin induced phosphorylation and activation of the cGMP-inhibited cAMP phosphodiesterase in human platelets. Biochem Biophys Res Commun 1992; 186: 517-523.
• 62 Wijkander J, Landström TR, Manganiello V, Belfrage P, Degerman E. Insulin-induced phosphorylation and activation of phosphodiesterase 3B in rat adipocytes: Possible role for protein kinase B but not mitogen-activated protein kinase or p70 S6 kinase. Endocrinology 1998; 139: 219-227.
• 63 Grant PG, Mannarino AF, Colman RW. Purification and characterization of a cyclic GMP-stimulated cyclic nucleotide phosphodiesterase from the cytosol of human platelets. Thromb Res 1990; 59: 105-119.
• 64 Michie AM, Lobban M, Müller T, Harnett MM, Houslay MD. Rapid regulation of PDE-2 and PDE-4 cyclic AMP phosphodiesterase activity following ligation of the T cell antigen receptor on thymocytes: Analysis using the selective inhibitors erythro-9-(2-hydroxy-3-nonyl)-adenine (EHNA) and rolipram. Cell Signalling 1996; 8: 97-110.
• 65 Méry P-F, Pavoine C, Pecker F, Fischmeister R. Erythro-9-(2-hydroxy-3-nonyl)adenine inhibits cyclic GMP-stimulated phosphodiesterase in isolated cardiac myocytes. Mol Pharmacol 1995; 48: 121-130.
• 66 Yamamoto T, Yamamoto S, Osborne JC, Manganiello VC, Vaughan M, Hidaka H. Complex effects of inhibitors on cyclic GMP-stimulated cyclic nucleotide phosphodiesterase. J Biol Chem 1983; 258: 14173-14177.
• 67 Coquil J-F, Franks DJ, Wells JN, Dupuis M, Hamet P. Characteristics of a new binding protein distinct from the kinase for guanosine 3’:5’-monophosphate in rat platelets. Biochim Biophys Acta 1980; 631: 148-165.
• 68 Francis SH, Lincoln TM, Corbin JD. Characterization of a novel cGMP binding protein from rat lung. J Biol Chem 1980; 255: 620-626.
• 69 Robichon A. A new cGMP phosphodiesterase isolated from bovine platelets is substrate for cAMP- and cGMP-dependent protein kinases: evidence for a key role in the process of platelet activation. J Cell Biochem 1991; 47: 147-157.
• 70 Bowen R, Haslam RJ. Effects of nitrovasodilators on platelet cyclic nucleotide levels in rabbit blood; role for cyclic AMP in synergistic inhibition of platelet function by SIN-1 and prostaglandin E₁ . J Cardiovasc Pharmacol 1991; 17: 424-433.
• 71 Levin RI, Weksler BB, Jaffe EA. The interaction of sodium nitroprusside with human endothelial cells and platelets: nitroprusside and prostracyclin synergistically inhibit platelet function. Circulation 1982; 66: 1299-1307.
• 72 Radomski MW, Palmer RMJ, Moncada S. The anti-aggregating properties of vascular endothelium: interactions between prostacyclin and nitric oxide. Br J Pharmacol 1987; 92: 639-646.
• 73 Andersson TLG, Vinge E. Interactions between isoprenaline, sodium nitroprusside, and isozyme-selective phosphodiesterase inhibitors on ADP-induced aggregation and cyclic nucleotide levels in human platelets. J Cardiovasc Pharmacol 1991; 18: 237-242.
• 74 Anfossi G, Massucco P, Piretto V, Mularoni E, Cavalot F, Mattiello L, Trovati M. Glyceryl trinitrate enhances the adenosine-induced inhibition of platelet responses: A mechanism potentially involved in the in vivo anti-aggregating effects of organic nitrates. Clin Exp Pharmacol Physiol 1995; 22: 803-811.
• 75 Grünberg B, Negrescu E, Siess W. Synergistic phosphorylation of platelet rap1B by SIN-1 and iloprost. Eur J Pharmacol 1995; 288: 329-333.
• 76 Fisch A, Michael-Hepp J, Meyer J, Darius H. Synergistic interaction of adenylate cyclase activators and nitric oxide donor SIN-1 on platelet cyclic AMP. Eur J Pharmacol 1995; 289: 455-461.
• 77 Jiang H, Shabb JB, Corbin JD. Cross-activation: overriding cAMP/cGMP selectivities of protein kinases in tissues. Biochem Cell Biol 1992; 70: 1283-1289.
• 78 Jiang H, Colbran JL, Francis SH, Corbin JD. Direct evidence for cross-activation of cGMP-dependent protein kinase by cAMP in pig coronary arteries. J Biol Chem 1992; 267: 1015-1019.
• 79 Landgraf W, Hullin R, Göbel C, Hofmann F. Phosphorylation of cGMP-dependent protein kinase increases the affinity for cyclic AMP. Eur J Biochem 1986; 154: 113-117.
• 80 Smith JA, Francis SH, Walsh KA, Kumar S, Corbin JD. Autophosphorylation of Type Iβ cGMP-dependent protein kinase increases basal catalytic activity and enhances allosteric activation by cGMP or cAMP. J Biol Chem 1996; 271: 20756-20762.
• 81 Geiger J, Nolte C, Butt E, Sage SO, Walter U. Role of cGMP and cGMP-dependent protein kinase in nitrovasodilator inhibition of agonist-evoked calcium elevation in human platelets. Proc Natl Acad Sci USA 1992; 89: 1031-1035.
• 82 Butt E, Nolte C, Schulz S, Beltman J, Beavo JA, Jastorff B, Walter U. Analysis of the functional role of cGMP-dependent protein kinase in intact human platelets using a specific activator 8-parachlorophenylthio-cGMP. Biochem Pharmacol 1992; 43: 2591-2600.
• 83 Butt E, Pöhler D, Genieser H-G, Huggins JP, Bucher B. Inhibition of cyclic GMP-dependent protein kinase-mediated effects by (Rp)-8-bromo-PET-cyclic GMPS. Br J. Pharmacol 1995; 116: 3110-3116.
• 84 Pfeifer A, Klatt P, Massberg S, Ny L, Sausbier M, Hirneiss C, Wang G-X, Korth M, Aszódi A, Andersson K-E, Krombach F, Mayerhofer A, Ruth P, Fässler R, Hofmann F. Defective smooth muscle regulation in cGMP kinase I-deficient mice. EMBO J 1998; 17: 3045-3051.
• 85 Massberg S, Sausbier M, Klatt P, Bauer M, Pfeifer A, Siess W, Fässler R, Ruth P, Krombach F, Hofmann F. Increased adhesion and aggregation of platelets lacking cyclic guanosine 3′, 5′-monphosphate kinase I. J Exp Med 1999; 189: 1255-1263.
• 86 Tang KM, Sherwood JL, Haslam RJ. Photoaffinity labelling of cyclic GMP-binding proteins in human platelets. Biochem J 1993; 294: 329-333.
• 87 Surks HK, Sun B, Zhu Y, Baur W, Hartwig J, Mendelsohn ME. Identification and cloning of a cGMP-gated ion channel from human platelets and megakaryocytes. Circulation 1995; 92: I-36 (Abstr).
• 88 de Rooij J, Zwartkruis FJT, Verheijen MHG, Cool RH, Nijman SMB, Wittinghofer A, Bos JL. Epac is a Rap1 guanine-nucleotide-exchange factor directly activated by cyclic AMP. Nature 1998; 396: 474-477.