Adenosine and Forskolin Inhibit Platelet Aggregation by Collagen but not the Proximal Signalling Events

Joanne C. Clark, Deirdre M. Kavanagh, Stephanie Watson, Jeremy A. Pike, Robert K. Andrews, Elizabeth E. Gardiner, Natalie S. Poulter, Stephen J. Hill, Steve P. Watson

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Abstract

Background The G protein-coupled receptor, adenosine A 2A, signals through the stimulatory G protein, G s, in platelets leading to activation of adenylyl cyclase and elevation of cyclic adenosine monophosphate (cAMP) and inhibition of platelet activation. Objective This article investigates the effect of A 2A receptor activation on signalling by the collagen receptor glycoprotein (GP) VI in platelets. Methods Washed human platelets were stimulated by collagen or the GPVI-specific agonist collagen-related peptide (CRP) in the presence of the adenosine receptor agonist, 5′-N-ethylcarboxamidoadenosine (NECA) or the adenylyl cyclase activator, forskolin and analysed for aggregation, adenosine triphosphate secretion, protein phosphorylation, spreading, Ca 2+ mobilisation, GPVI receptor clustering, cAMP, thromboxane B 2 (TxB 2) and P-selectin exposure. Results NECA, a bioactive adenosine analogue, partially inhibits aggregation and secretion to collagen or CRP in the absence or presence of the P2Y 12 receptor antagonist, cangrelor and the cyclooxygenase inhibitor, indomethacin. The inhibitory effect in the presence of the three inhibitors is largely overcome at higher concentrations of collagen but not CRP. Neither NECA nor forskolin altered clustering of GPVI, elevation of Ca 2+ or spreading of platelets on a collagen surface. Further, neither NECA nor forskolin, altered collagen-induced tyrosine phosphorylation of Syk, LAT nor PLCγ2. However, NECA and forskolin inhibited platelet activation by the TxA 2 mimetic, U46619, but not the combination of adenosine diphosphate and collagen. Conclusion NECA and forskolin have no effect on the proximal signalling events by collagen. They inhibit platelet activation in a response-specific manner in part through inhibition of the feedback action of TxA 2.

Original languageEnglish
Pages (from-to)1124-1137
Number of pages14
JournalThrombosis and Haemostasis
Volume119
Issue number7
DOIs
Publication statusPublished - 1 Jul 2019

Keywords

  • adenosine
  • cAMP
  • collagen
  • glycoprotein VI (GPVI) receptor
  • platelets

Cite this

Clark, J. C., Kavanagh, D. M., Watson, S., Pike, J. A., Andrews, R. K., Gardiner, E. E., Poulter, N. S., Hill, S. J., & Watson, S. P. (2019). Adenosine and Forskolin Inhibit Platelet Aggregation by Collagen but not the Proximal Signalling Events. Thrombosis and Haemostasis, 119(7), 1124-1137. https://doi.org/10.1055/s-0039-1688788