DIFFERENT ABILITIES OF THROMBIN RECEPTOR ACTIVATING PEPTIDE AND THROMBIN TO INDUCE PLATELET CALCIUM RISE AND FULL RELEASE REACTION

Synthetic peptides (TRAP or Thrombin Receptor Activating Peptide) corr esponding to at least the first five aminoacids of the new N-terminal tail generated after thrombin proteolysis of its receptor are effectiv e to mimic thrombin. We have studied two different TRAPs (SFLLR, and S FLLRN) in their effectiveness to induce the different platelet respons es in comparison with thrombin. Using Indo-1/AM-labelled platelets, th e maximum rise in cytoplasmic ionized calcium was lower with TRAPs tha n with thrombin. At threshold concentrations allowing maximal aggregat ion (50 mu M SFLLR, 5 mu M SFLLRN and 1 nM thrombin) the TRAPs-induced release reaction was about the same level as with thrombin, except wh en external calcium was removed by addition of 1 mM EDTA. In these con ditions, the dense granule release induced by TRAPs was reduced by ove r 60%, that of lysosome release by 75%, compared to only 15% of reduct ion in the presence of thrombin. Thus calcium influx was more importan t for TRAPs-induced release than for thrombin-induced release. At stro ng concentrations giving maximal aggregation and release in the absenc e of secondary mediators (by pretreatment with ADP scavengers plus asp irin), SFLLRN mobilized less calcium, with a fast return towards the b asal level and induced smaller lysosome release than did thrombin. The results further demonstrate the essential role of external Calcium in triggering sustained and full platelet responses, and emphasize the m ajor difference between TRAP and thrombin in mobilizing [Ca2+](i). Thu s, apart from the proteolysis of the seven transmembrane receptor, ano ther thrombin binding site or thrombin receptor interaction is require d to obtain full and complete responses.