Evidence That Activation of Phospholipase D can Mediate Secretion from Permeabilized Platelets

GTPγS and phorbol ester act synergistically to stimulate both Ca2+‐independent secretion and phospholipase D activity in permeabilized human platelets

“…Although BAPTA binds Ca2+ more rapidly than does EGTA [lo], which may account for the ability of the former to block neurotransmitter release [22], differences in the Ca"-binding kinetics of these chelating agents are unlikely to explain the present results. Thus, we have observed similar inhibitions of GTPyS-stimulated secretion and PLD activity by BAPTA in the presence of Ca2' buffered to a final pCa of 6 [2]. Moreover, dinitro-BAPTA, which has little or no Ca*'-binding capacity [21], was slightly more potent than BAPTA as an inhibitor of secretion and PLD activity in our experiments.…”

“…A similar role for PLD has also been suggested by others working with mast cells [19], HL-60 cells [20] and pancreatic islets [23]. However, in intact human platelets, PLD activity accounts for only l&20% of the total PA that accumulates after stimulation by thrombin [2,24]; most of the PA is formed by the sequential actions of phospholipase C and diacylglycerol kinase. Thus, a major role for PLD in secretion from intact platelets is only likely if the PA generated by this enzyme differs in its fatty acid composition or, more plausibly, in its subcellular localization from that formed as a result of phospholipase C activity.…”

“…A previous study from this laboratory [l] showed that GTPyS can induce the Ca"-independent secretion of both dense and a-granule constituents from electropermeabilized human platelets without causing stimulation of phosphoinositide-specific phospholipase C. This action of GTPyS was dependent on PKC activity [2] and was potentiated by phorbol ester (PMA) [1,2]. Similar studies with other permeabilized cells, particularly neutrophils [3], HL-60 cells [4] and RINmSF cells [5], in which Ca2'-independent secretion was observed, and with mast cells in which Ca2* was required [6,7], have led Gomperts and colleagues to propose that an unidentified G protein, designated G,, may mediate GTPySinduced exocytosis (reviewed in [S]).…”

Protein kinase C-dependent and Ca2+-dependent mechanisms of secretion from streptolysin O-permeabilized platelets: effects of leakage of cytosolic proteins

“…In addition, use of this experimental system has led to the recognition of at least two distinct roles for GTP-binding proteins in secretion from platelets [5], as found in experiments with other permeabilized cells [6]. Thus GTP or its more stable analogue, guanosine 5h-[γ-thio]-triphosphate (GTP [S]) is required for the G-protein-and Ca# + -dependent activation of phospholipase C in permeabilized platelets [7], which results in the stimulation of PKC by 1,2-diacylglycerol [8]. Secondly, experiments with electropermeabilized platelets have shown that GTP[S] acts synergistically with PKC when this enzyme is stimulated by phorbol 12-myristate 13-acetate (PMA), to induce a Ca# + -independent secretion of granule constituents [5] ; this effect is associated with the activation of phospholipase D, but not of phospholipase C [8,9].…”

Choline in acute coronary syndrome: an emerging biomarker with implications for the integrated assessment of plaque vulnerability

“…PLD activity is increased in specific inflammatory reactions in leukocytes [21] and is involved in platelet activation. It has been hypothesized that a major factor causing choline increase in whole blood and serum is the activation of platelet and leukocyte PLD by thrombin and collagen [22][23][24][25]. Moreover, it has been suggested that most of the generated choline remains intracellular and only a minor fraction is released.…”