Biochemical and Ultrastructural Studies Suggest that the Effects of Thapsigargin on Human Platelets Are Mediated by Changes in Intracellular Calcium but not by Intracellular Histamine

Summary

The involvement of intracellular histamine in thapsigargin (Tg)-induced platelet aggregation was studied. Platelet aggregation induced by 0.25 and 0.5 µM Tg was not accompained by a rise in intracellular histamine but a significant (p <0.01) increase in the level of intracellular histamine was observed at 1 µM Tg. Preincubation of platelets with inhibitors of histamine metabolizing enzymes had little effect on intracellular histamine levels in platelets stimulated by 0.5 µM Tg. In addition, the inhibitors of histidine decarboxylase (HDC), α-methyl histidine (α-MH) and α-fluoromethyl histidine (α-FMH) failed to inhibit Tg-induced aggregation. The intracellular histamine receptor antagonist, N,N-diethyl-2-[4-(phenylmethyl)phenoxy] ethanamine. HC1 (DPPE), inhibited Tg-induced aggregation but with IC₅₀ values dependent on the concentration of agonist used. The inhibitory effects of DPPE on Tg-induced aggregation were not reversed by the addition of histamine to saponin-permeabilized platelets suggesting non-histamine mediated effects of DPPE on Tg-induced aggregation. Tg stimulated an increase in the cytosolic free calcium concentration which was unaffected by DPPE indicating that the effects of DPPE are also not due to the inhibition of mobilization of cytosolic calcium. The ultrastructural studies suggest that the major Tg-induced changes (pseudopod formation and granule centralization) are consistent with a primary role for Tg to mobilize calcium; DPPE had very little effect on these ultrastructural changes. The results indicate that the effects of Tg on human platelets are mediated by an increase in cytosolic calcium but not by intracellular histamine.

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