<b>AUGMENTATION OF MALONDIALDEHYDE PRODUCTION IN RESERPINIZED RABBIT PLATELETS BY ENHANCEMENT OF INTRACELLULAR CA²⁺-MOBILIZATION IN RESPONSE TO </b><b>THROMBIN </b>

Abstract

Blood platelets from reserpinized rabbits produced a larger amount of malondialdehyde (M DA) in response to thrombin than the platelets from control rabbits. The MDA production was almost completely inhibited by aspirin and indomethacin, indicating that the MDA was produced through the cyclooxygenase pathway. When intracellular free Ca" was diminished by tetracaine or dibutyryl cyclic AMP, thrombin could not induce MDA production in the reserpinized or in the control platelets. When intracellular Ca" was raised by ionophore A23l87 plus Ca", MDA was produced in the absence of thrornbin. No difference in MDA production, however, was observed between the reserpinized and the control platelets. The measurement of intracellular bound calcium with chlortetracycline showed that the fluorescence after thrombin addition in the reserpinized platelets was decreased more than in the controls, suggesting an enhancement of intracellular liberation of Ca" from the binding sites of the reserpinized platelets. The augmentation of MDA production in the reserpinized platelets can be explained as a result of a stimulation of the cyclooxygenase pathway. The stimulation might be due to an excess supply of the substrates caused by Ca"-dependent degradation of platelet phospholipids, which is enhanced by intracellular Ca" mobilization in response to thrombin.It IS thought that thrombin-induced platelet responses are mediated by intracellular calcium ions liberated from specific storage sites (7, 9, ll, l2, 24, 38). An increased cytoplasmic level of calcium ion increases the activity of phospholipases (27, 29, 30, 32), which liberate fatty acids, arachidonic acid among others (1, 4), from platelet phospholipids. The fatty acid is transformed into MDA and other products through biologically active intermediates in the cyclooxygenase pathway (3,16, 17). In order to estimate the cyclooxygenase pathway, measurement of MDA by thiobarbiturate (TBA) has been utilized as a simple method (33). Recently the production of TBA-reactive substances through the lipoxygenase pathway was demonstrated in human platelets (2, 34). We have demonstrated that serotonin-depleted blood platelets from reserpinized rabbits aggregated more rapidly and produced a larger amount of TBApositive substances than the control in response to thrombin, although no difference in the production was observed in response to arachidonic acid (36). The enhancement of thrombin-induced degradation of phospholipids and subsequent production of fatty acids and their oxygenated metabolites in the reserpinized platelets were also demonstrated (37). It is

Enhancement of the Release Reaction Not Accompanied with Enhanced Phosphatidylinositol Turnover in the Reserpinized Rabbit Blood Platelets

“…T he blood platelets from reserpine-treated rabbits contain a larger am ount of calcium (1), and aggregate m ore rapidly with production of a larger am ount of m alondialdehyde in response to throm bin than the control platelets (2). It is conceivable th at the increased sensitivity of the reserpinized platelets is attributed to an augm entation of intracellular Ca2+-m obilization which causes an activation of phospholipase A 2 and an increase in arachidonate release from platelet phospholipids.…”

“…It is conceivable th at the increased sensitivity of the reserpinized platelets is attributed to an augm entation of intracellular Ca2+-m obilization which causes an activation of phospholipase A 2 and an increase in arachidonate release from platelet phospholipids. Subsequent increase in arachidonate m etabolites produced through the cyclooxygenase system, P G G 2 and throm boxane A 2 etc., will induce the rapid and m arked aggregation (1,3). Calcium plays an im portant role also in regulation of platelet secretion of granular com ponents.…”