DIACYLGLYCEROL ELEVATIONS IN CONTROL PLATELETS ARE UNACCOMPANIED BY PLECKSTRIN PHOSPHORYLATION - IMPLICATIONS FOR THE ROLE OF DIACYLGLYCEROL IN PLATELET ACTIVATION

Several laboratories have reported that diacylglycerol levels in human platelets (approximate to 100 pmol/10(9) platelets) increased severalfold in response to 0.5-1 U/ml thrombin. We report here fluctuations in diacylglycerol mass in control platelets, the magnitude of which were 60-90% of that measured in platelets treated with 0.2-0.5 U/ml of thrombin. These control platelets were not activated by such criteria as absence of aggregation, secretion, phosphatidic acid production and phosphorylation of the protein kinase C substrate, pleckstrin. Thrombin treatment evoked all of the above responses. Analysis of the diacylglycerol molecular species by reverse-phase HPLC of the dimethylated, phosphorylated derivatives showed that all of the molecular species that were present in control platelets were also present in thrombin-treated platelets. Most of the species appeared to fluctuate at random in control platelets with the exception of 1-stearoyl-2-arachidonoyl-sn-glycerol which was more or less stable and increased severalfold over control values only upon thrombin treatment. Furthermore, only this species accumulated as [P-32]phosphorylated PtdOH in thrombin-treated platelets prelabelled with [P-32]P-i. Our findings show that, in platelets elevation of diacylglycerol molecular species other than the 1-stearoyl-2-arachidonoyl species occurs, but these changes are not necessarily linked to activation of protein kinase C as measured by pleckstrin phosphorylation which was observed only upon elevation of 1-stearoyl-2-arachidonoyl-sn-glycerol.