Rapid resensitization of purinergic receptor function in human platelets.
BACKGROUND
Adenosine diphosphate (ADP) is a critical regulator of platelet activation, mediating its actions through two G protein-coupled receptors (GPCRs), the P2Y(1) and P2Y(12) purinergic receptors. Recently, we demonstrated that both receptors desensitize and internalize in human platelets by differential kinase-dependent mechanisms.


OBJECTIVE
To demonstrate whether responses to P2Y(1) and P2Y(12) purinergic receptors resensitize in human platelets and determine the role of receptor traffic in this process.


METHODS
These studies were undertaken either in human platelets or in cells stably expressing epitope-tagged P2Y(1) and P2Y(12) purinergic receptor constructs.


RESULTS
In this study we show for the first time that responses to both of these receptors can rapidly resensitize following agonist-dependent desensitization in human platelets. Further, we show that in human platelets or in 1321N1 cells stably expressing receptor constructs, the disruption of receptor internalization, dephosphorylation or subsequent receptor recycling is sufficient to block resensitization of purinergic receptor responses. We also show that, in platelets, internalization of both these receptors is dependent upon dynamin, and that this process is required for resensitization of responses.


CONCLUSIONS
This study is therefore the first to show that both P2Y(1) and P2Y(12) receptor activities are rapidly and reversibly modulated in human platelets, and it reveals that the underlying mechanism requires receptor trafficking as an essential part of this process.
Department of Physiology and Pharmacology, School of Medical Sciences, University Walk, Bristol, UK.
S J
SJ
Mundell
J F
JF
Barton
M B
MB
Mayo-Martin
A R
AR
Hardy
A W
AW
Poole
D000244:Adenosine Diphosphate / Q000097:blood / Q000494:pharmacology; D001792:Blood Platelets / Q000187:drug effects / Q000378:metabolism*; D002118:Calcium / Q000097:blood; D002460:Cell Line; D000242:Cyclic AMP / Q000097:blood; D006801:Humans; D066298:In Vitro Techniques; D008985:Monensin / Q000494:pharmacology; D015539:Platelet Activation / Q000187:drug effects / Q000502:physiology; D058913:Purinergic Agonists; D011983:Receptors, Purinergic / Q000097:blood*; D018048:Receptors, Purinergic P2 / Q000097:blood* / Q000235:genetics / Q000378:metabolism; D058472:Receptors, Purinergic P2Y1; D058925:Receptors, Purinergic P2Y12; D011994:Recombinant Proteins / Q000235:genetics / Q000378:metabolism; D014162:Transfection
C546382:P2RY1 protein, human; C548576:P2RY12 protein, human; D058913:Purinergic Agonists; D011983:Receptors, Purinergic; D018048:Receptors, Purinergic P2; D058472:Receptors, Purinergic P2Y1; D058925:Receptors, Purinergic P2Y12; D011994:Recombinant Proteins; D000244:Adenosine Diphosphate; D008985:Monensin; D000242:Cyclic AMP; D002118:Calcium