THROMBIN RECEPTORS ON HUMAN PLATELETS - INITIAL LOCALIZATION AND SUBSEQUENT REDISTRIBUTION DURING PLATELET ACTIVATION

Platelet responses to thrombin are at least partly mediated by a G-pro tein-coupled receptor whose NH2 terminus is a substrate for thrombin, In the present studies we have examined the location of thrombin recep tors in resting platelets and followed their redistribution during pla telet activation, The results reveal several new aspects of thrombin r eceptor biology, 1) On resting platelets, approximately two-thirds of the receptors were located in the plasma membrane, The remainder were present in the membranes of the surface connecting system, 2) When pla telets were activated by ADP or a thromboxane analog, thrombin recepto rs that were initially in the surface connecting system were exposed o n the platelet surface, increasing the number of detectable receptors by 40% and presumably making them available for subsequent activation by thrombin, 3) Platelet activation by thrombin rapidly abolished the binding of the antibodies whose epitopes are sensitive to receptor cle avage and left the platelets in a state refractory to both thrombin an d the agonist peptide, SFLLRN, This was accompanied by a 60% decrease in the binding of receptor antibodies directed COOH-terminal to the cl eavage site irrespective of whether the receptors were activated prote olytically by thrombin or nonproteolytically by SFLLRN, 4) The loss of antibody binding sites caused by thrombin was due in part to receptor internalization and in part to the shedding of thrombin receptors int o membrane microparticles, especially under conditions in which aggreg ation was allowed to occur, However, at least 40% of the cleaved recep tors remained on the platelet surface, 5) Lacking the ability to synth esize new receptors and lacking an intracellular reserve of preformed receptors comparable to that found in endothelial cells, platelets wer e unable to repopulate their surface with intact receptors following e xposure to thrombin, This difference underlies the ability of endothel ial cells to recover responsiveness to thrombin rapidly while platelet s do not, despite the presence on both of the same receptor for thromb in.