{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2026,10,21]],"date-time":"2026-10-21T10:55:02Z","timestamp":1792580102980,"version":"4.5.0"},"reference-count":32,"publisher":"American Society of Hematology","issue":"7","content-domain":{"domain":["ashpublications.org"],"crossmark-restriction":true},"short-container-title":[],"published-print":{"date-parts":[[2002,10,1]]},"abstract":"<jats:p>Adenosine triphosphate (ATP) and its stable analog, \u03b1,\u03b2-methylene ATP, activate the platelet P2X1 ion channel, causing a rapid Ca++ influx. Here, we show that, in washed apyrase-treated platelets, \u03b1,\u03b2-methylene ATP elicits reversible extracellular signal-regulated kinase 2 (ERK2) phosphorylation through a Ca++- and protein kinase C\u2013dependent pathway. In contrast, high-performance liquid chromatography-purified adenosine diphosphate (ADP) did not trigger ERK2 phosphorylation. \u03b1,\u03b2-Methylene ATP also activated the ERK2 pathway in P2X1-transfected HEK293 cells but not in cells expressing mutated P2X1delL nonfunctional channels. Because ATP released from the dense granules during platelet activation contributes to platelet aggregation elicited by low doses of collagen, and because collagen causes ERK2 phosphorylation, we have investigated the role of P2X1-mediated ERK2 activation in these platelet responses. We found that the antagonism of P2X1 with ADP or desensitization of this ion channel with \u03b1,\u03b2-methylene ATP both resulted in impaired ERK2 phosphorylation, ATP secretion, and platelet aggregation induced by low concentrations of collagen (\u2264\u20091 \u03bcg\/mL) without affecting the minor early dense granule release. Selective MEK1\/2 inhibition by U-0126 and Ca++ chelation with EGTA (ethyleneglycoltetraacetic acid) behaved similarly, whereas the PKC inhibitor GF109203-X totally prevented collagen-induced secretion and ERK2 activation. In contrast, when elicited by high collagen concentrations (2 \u03bcg\/mL), platelet aggregation and secretion no longer depended on P2X1 or ERK2 activation, as shown by the lack of their inhibition by \u03b1,\u03b2-methylene ATP or U-0126. We thus conclude that mild platelet stimulation with collagen rapidly releases ATP, which activates the P2X1-PKC-ERK2 pathway. This process enhances further degranulation of the collagen-primed granules allowing platelet aggregation to be completed.<\/jats:p>","DOI":"10.1182\/blood-2002-03-0812","type":"journal-article","created":{"date-parts":[[2002,9,18]],"date-time":"2002-09-18T12:26:06Z","timestamp":1032351966000},"page":"2499-2505","update-policy":"https:\/\/doi.org\/10.1182\/blood.2019cm0000","source":"Crossref","is-referenced-by-count":77,"title":["P2X1-mediated activation of extracellular signal-regulated kinase 2 contributes to platelet secretion and aggregation induced by collagen"],"prefix":"10.1182","volume":"100","author":[{"given":"Ce\u0301cile","family":"Oury","sequence":"first","affiliation":[{"name":"From the Center for Molecular and Vascular Biology, University of Leuven, Belgium."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Emese","family":"Toth-Zsamboki","sequence":"additional","affiliation":[{"name":"From the Center for Molecular and Vascular Biology, University of Leuven, Belgium."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Jos","family":"Vermylen","sequence":"additional","affiliation":[{"name":"From the Center for Molecular and Vascular Biology, University of Leuven, Belgium."}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Marc F.","family":"Hoylaerts","sequence":"additional","affiliation":[{"name":"From the Center for Molecular and Vascular Biology, University of Leuven, Belgium."}],"role":[{"vocabulary":"crossref","role":"author"}]}],"member":"234","reference":[{"key":"2019111902364508500_B1","first-page":"85","article-title":"Adenine nucleotides in blood platelets.","volume-title":"The Platelets. Physiology and Pharmacology.","author":"Reimers","year":"1985"},{"key":"2019111902364508500_B2","doi-asserted-by":"crossref","first-page":"835","DOI":"10.1055\/s-0038-1650674","article-title":"ADP receptors on platelets.","volume":"76","author":"Mills","year":"1996","journal-title":"Thromb Haemost"},{"key":"2019111902364508500_B3","doi-asserted-by":"crossref","first-page":"222","DOI":"10.1055\/s-0037-1616220","article-title":"ADP receptors of platelets and their inhibition.","volume":"86","author":"Gachet","year":"2001","journal-title":"Thromb Haemost"},{"key":"2019111902364508500_B4","doi-asserted-by":"crossref","first-page":"202","DOI":"10.1038\/35051599","article-title":"Identification of the platelet ADP receptor targeted by antithrombotic drugs.","volume":"409","author":"Hollopeter","year":"2001","journal-title":"Nature."},{"key":"2019111902364508500_B5","doi-asserted-by":"crossref","first-page":"2879","DOI":"10.1074\/jbc.271.6.2879","article-title":"Activation of receptor-operated cation channels via P2X1 not P2T purinoceptors in human platelets.","volume":"271","author":"MacKenzie","year":"1996","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B6","doi-asserted-by":"crossref","first-page":"108","DOI":"10.1038\/sj.bjp.0703517","article-title":"ADP is not an agonist at P2X1 receptors: evidence for separate receptors stimulated by ATP and ADP on human platelets.","volume":"131","author":"Mahaut-Smith","year":"2000","journal-title":"Br J Pharmacol."},{"key":"2019111902364508500_B7","first-page":"178","article-title":"Pharmacological characterization of heterologously expressed ATP-gated cation channels (P2X purinoceptors).","volume":"48","author":"Evans","year":"1995","journal-title":"Mol Pharmacol."},{"key":"2019111902364508500_B8","doi-asserted-by":"crossref","first-page":"303","DOI":"10.1055\/s-0037-1615684","article-title":"Platelet shape change evoked by selective activation of P2X1 purinoceptors with \u03b1,\u03b2-methylene ATP.","volume":"85","author":"Rolf","year":"2001","journal-title":"Thromb Haemost."},{"key":"2019111902364508500_B9","doi-asserted-by":"crossref","first-page":"1264","DOI":"10.1055\/s-0037-1616060","article-title":"The ATP-gated P2X1 ion channel acts as a positive regulator of platelet responses to collagen.","volume":"86","author":"Oury","year":"2001","journal-title":"Thromb Haemost."},{"key":"2019111902364508500_B10","doi-asserted-by":"crossref","first-page":"716","DOI":"10.1111\/j.1749-6632.1999.tb11351.x","article-title":"Functional and molecular diversity of purinergic ion channel receptors.","volume":"868","author":"MacKenzie","year":"1999","journal-title":"Ann N Y Acad Sci."},{"key":"2019111902364508500_B11","doi-asserted-by":"crossref","first-page":"19965","DOI":"10.1074\/jbc.273.32.19965","article-title":"ATP-stimulated activation of the mitogen-activated protein kinases through ionotrophic P2X2 purinoreceptors in PC12 cells. difference in purinoreceptor sensitivity in two PC12 cell lines.","volume":"273","author":"Swanson","year":"1998","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B12","doi-asserted-by":"crossref","first-page":"211","DOI":"10.1016\/S0955-0674(99)80028-3","article-title":"Organization and regulation of mitogen-activated protein kinase signaling pathways.","volume":"11","author":"Garrington","year":"1999","journal-title":"Curr Opin Cell Biol."},{"key":"2019111902364508500_B13","doi-asserted-by":"crossref","first-page":"14816","DOI":"10.1074\/jbc.270.24.14816","article-title":"Differential activation of cytosolic phospholipase A2 (cPLA2) by thrombin and thrombin receptor agonist peptide in human platelets. Evidence for activation of cPLA2 independent of the mitogen-activated protein kinases ERK1\/2.","volume":"270","author":"Kramer","year":"1995","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B14","doi-asserted-by":"crossref","first-page":"207","DOI":"10.1042\/bj3180207","article-title":"Inhibition of mitogen-activated protein kinase kinase does not impair primary activation of human platelets.","volume":"318","author":"Borsch-Haubold","year":"1996","journal-title":"Biochem J."},{"key":"2019111902364508500_B15","doi-asserted-by":"crossref","first-page":"16494","DOI":"10.1074\/jbc.271.28.16494","article-title":"Stimulation of mitogen-activated protein kinase and Na+\/H+ exchanger in human platelets. Differential effect of phorbol ester and vasopressin.","volume":"271","author":"Aharonovitz","year":"1998","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B16","doi-asserted-by":"crossref","first-page":"15659","DOI":"10.1074\/jbc.M009218200","article-title":"Role of the ERK pathway in the activation of store-mediated calcium entry in human platelets.","volume":"276","author":"Rosado","year":"2001","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B17","doi-asserted-by":"crossref","first-page":"22381","DOI":"10.1074\/jbc.272.36.22381","article-title":"Negative regulation of mitogen-activated protein kinase activation by integrin \u03b1IIb\u03b23 in platelets.","volume":"272","author":"Nadal","year":"1997","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B18","doi-asserted-by":"crossref","first-page":"42226","DOI":"10.1074\/jbc.M106129200","article-title":"A mitogen-activated protein kinase-dependent signaling pathway in the activation of platelet integrin \u03b1IIb\u03b23.","volume":"276","author":"Li","year":"2001","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B19","doi-asserted-by":"crossref","first-page":"213","DOI":"10.1021\/jm981072s","article-title":"Antagonists of the platelet P2T receptor: a novel approach to antithrombotic therapy.","volume":"42","author":"Ingall","year":"1999","journal-title":"J Med Chem."},{"key":"2019111902364508500_B20","doi-asserted-by":"crossref","first-page":"22611","DOI":"10.1074\/jbc.C000305200","article-title":"A natural dominant negative P2X1 receptor due to deletion of a single amino acid residue.","volume":"275","author":"Oury","year":"2000","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B21","doi-asserted-by":"crossref","first-page":"28766","DOI":"10.1074\/jbc.273.44.28766","article-title":"Direct inhibition of cyclooxygenase-1 and -2 by the kinase inhibitors SB 203580 and PD 98059. SB 203580 also inhibits thromboxane synthase.","volume":"273","author":"Borsch-Haubold","year":"1998","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B22","doi-asserted-by":"crossref","first-page":"445","DOI":"10.1016\/0049-3848(86)90089-7","article-title":"ATP secretion occurs as an initial response in collagen-induced platelet activation.","volume":"43","author":"Malmgren","year":"1986","journal-title":"Thromb Res."},{"key":"2019111902364508500_B23","doi-asserted-by":"crossref","first-page":"2283","DOI":"10.1046\/j.1432-1327.2000.01235.x","article-title":"The P2Y1 receptor mediates ADP-induced p38 kinase-activating factor generation in human platelets.","volume":"267","author":"Dangelmaier","year":"2000","journal-title":"Eur J Biochem."},{"key":"2019111902364508500_B24","doi-asserted-by":"crossref","first-page":"1313","DOI":"10.1182\/blood.V94.4.1313","article-title":"A critical role for N-ethylmaleimide-sensitive fusion protein (NSF) in platelet granule secretion.","volume":"94","author":"Polgar","year":"1999","journal-title":"Blood."},{"key":"2019111902364508500_B25","doi-asserted-by":"crossref","first-page":"33844","DOI":"10.1074\/jbc.M002834200","article-title":"Small GTPase Rab4 regulates Ca2+-induced alpha-granule secretion in platelets.","volume":"275","author":"Shirakawa","year":"2000","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B26","doi-asserted-by":"crossref","first-page":"307","DOI":"10.1016\/S0197-0186(01)00093-6","article-title":"Bisperoxovanadium complex promotes dopamine exocytosis in PC12 cells.","volume":"40","author":"Bieger","year":"2002","journal-title":"Neurochem Int."},{"key":"2019111902364508500_B27","doi-asserted-by":"crossref","first-page":"605","DOI":"10.1124\/jpet.300.2.605","article-title":"EctoNucleotidase in cardiac sympathetic nerve endings modulates ATP-mediated feedback of norepinephrine release.","volume":"300","author":"Sesti","year":"2002","journal-title":"J Pharmacol Exp Ther."},{"key":"2019111902364508500_B28","doi-asserted-by":"crossref","first-page":"8","DOI":"10.1002\/glia.10004","article-title":"Activation of P2X(7) receptors induced [(3)H]GABA release from the RBA-2 type-2 astrocyte cell line through a Cl(-)\/HCO(3)(-)-dependent mechanism.","volume":"37","author":"Wang","year":"2002","journal-title":"Glia."},{"key":"2019111902364508500_B29","doi-asserted-by":"crossref","first-page":"951","DOI":"10.1023\/A:1012336601854","article-title":"Distinct mechanisms underlying alpha1-adrenoceptor and P2x purinoceptor operated ATP release and contraction in the guinea-pig vas deferens.","volume":"26","author":"Sperlagh","year":"2001","journal-title":"Neurochem Res."},{"key":"2019111902364508500_B30","doi-asserted-by":"crossref","first-page":"6522","DOI":"10.1523\/JNEUROSCI.21-17-06522.2001","article-title":"ATP P2X receptor-mediated enhancement of glutamate release and evoked EPSCs in dorsal horn neurons of the rat spinal cord.","volume":"21","author":"Nakatsuka","year":"2001","journal-title":"J Neurosci."},{"key":"2019111902364508500_B31","doi-asserted-by":"crossref","first-page":"39379","DOI":"10.1074\/jbc.M102933200","article-title":"Identification of protein kinase Calpha as an essential, but not sufficient, cytosolic factor for Ca2+-induced alpha- and dense-core granule secretion in platelets.","volume":"276","author":"Yoshioka","year":"2001","journal-title":"J Biol Chem."},{"key":"2019111902364508500_B32","doi-asserted-by":"crossref","first-page":"863","DOI":"10.1038\/287863a0","article-title":"Ca2+-calmodulin-dependent phosphorylation and platelet secretion.","volume":"287","author":"Nishikawa","year":"1980","journal-title":"Nature."}],"container-title":["Blood"],"original-title":[],"language":"en","link":[{"URL":"http:\/\/ashpublications.org\/blood\/article-pdf\/100\/7\/2499\/1254377\/h81902002499.pdf","content-type":"application\/pdf","content-version":"vor","intended-application":"syndication"},{"URL":"http:\/\/ashpublications.org\/blood\/article-pdf\/100\/7\/2499\/1254377\/h81902002499.pdf","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2019,11,19]],"date-time":"2019-11-19T02:38:55Z","timestamp":1574131135000},"score":1,"resource":{"primary":{"URL":"https:\/\/ashpublications.org\/blood\/article\/100\/7\/2499\/106173\/P2X1mediated-activation-of-extracellular"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2002,10,1]]},"references-count":32,"journal-issue":{"issue":"7","published-print":{"date-parts":[[2002,10,1]]}},"URL":"https:\/\/doi.org\/10.1182\/blood-2002-03-0812","relation":{},"ISSN":["1528-0020","0006-4971"],"issn-type":[{"value":"1528-0020","type":"electronic"},{"value":"0006-4971","type":"print"}],"subject":[],"published":{"date-parts":[[2002,10,1]]}}}