Triphosphoinositide breakdown and dense body release as the earliest events in thrombin-induced activation of human platelets

The activation by thrombin of human platelets prelabelled with 32P induced a 30-40% decrease in 32P-triphosphoinositides (TPI) in the first 10 sec; the decrease in the other 32P-labelled phosphoinositides occurred by 20-30 sec. At 10 sec., the intensity of these effects was maximum with 0.2-0.4 U/ml thrombin. Under these conditions, 53, 20 and 15% of the dense granule, alpha-granule and lysosome constituents, respectively were released and thromboxane B2 synthesis reached only 10% of its maximum. Together with experiments carried out with chlorpromazine - or PGE1 - treated platelets, our results suggest the existence of a close relationship between TPI-breakdown and dense body release which appear to be the earliest events resulting from the activation of human platelets by thrombin.     

AGEPC (platelet activating factor) induced stimulation of rabbit platelets: effects on phosphatidylinositol, di- and triphosphoinositides and phosphatidic acid metabolism

Stimulation of washed rabbit platelets with AGEPC (1-O-alkyl-2-acetyl-$\text{sn}$-glyceryl-3-phosphorylcholine) caused a 15–20% decrease in their phosphatidylinositol level within 15 seconds without affecting other major classes of phospholipids. In the same time frame the level of phosphatidic acid (PA) increased dramatically some four fold. LysoGEPC, which is inactive in stimulating rabbit platelets, did not cause any change in PI or PA. When [³²Pi] was present during the stimulation of platelets by AGEPC, the incorporation of radiolabel into PI-4-phosphate (DPI), PI-4,5-bis phosphate (TPI) and PA was enhanced significantly within one minute while the incorporation into PI increased only after one minute. These results clearly established that AGEPC induced stimulation of rabbit platelets was associated with the metabolism of inositol phospholipids and phosphatidic acid. The relevance of these findings to the mode of action of AGEPC and Ca²⁺ mobilization is also discussed.     

400 MHz proton nuclear magnetic resonance studies of the polar headgroup conformation of inositolphospholipids

400 MHz ¹H-resonances of 1-phosphatidyl-myoinositol (PI), PI 4-phosphate (DPI) and PI 4,5-bisphosphate (TPI) in CD₃OD were assigned. Proton resonances in the inositol moiety shift downfield with the increase in the number of the phosphate from PI to TPI. From the ¹H-¹H and ¹H-³¹P vicinal coupling constants, rotamer populations around bonds in the polar headgroups were calculated. The H-C(5)O-P bond at position 5 of the inositol moiety tends to assume a gauche form. The H-C(1)O-P and the H-C(4)O-P bonds are not so strongly restricted to the gauche form as the H-C(5)O-P bond. The conformation of the glycerol moiety in PI, DPI and TPI is similar to that in phosphotidylcholine (PC) and phosphatidylethanolamine (PE). The CH-CH₂O-P bond in the glycerol moiety assumes a trans form. The acyl chains prefer a gauche arrangement to each other around the CHCH₂OCOR bond.