The activation by Ca 2+ of platelet phospholipase A 2: Effects of dibutyryl cyclic adenosine monophosphate and 8-( N, N-diethylamino)-octyl-3,4,5-trimethoxybenzoate

Abstract

Thrombin-induced release of arachidonic acid from human platelet phosphatidylcholine is found to be more than 90% impaired by incubation of platelets with 1 mM dibutyryl cyclic adenosine monophosphate (Bt 2 cyclic AMP) or with 0.6 mM 8-( N, N-diethylamino)-octyl-3,4,5-trimethoxybenzoate (TMB-8), an intracellular calcium antagonist. Incorporation of arachidonic acid into platelet phospholipids is not enhanced by Bt 2 cyclic AMP. The addition of external Ca 2+ to thrombin-treated platelets incubated with Bt 2 cyclic AMP or TMB-8 does not counteract the observed inhibition. However, when divalent cation ionophore A23187 is employed as an activating agent, much less inhibition is produced by Bt 2 cyclic AMP or TMB-8. The inhibition which does result can be overcome by added Ca 2+. Inhibition of arachidonic acid liberation by Bt 2 cyclic AMP, but not by TMB-8, can be overcome by high concentrations of A23187. When Mg 2+ is substituted for Ca 2+, ionophore-induced release of arachidonic acid from phosphatidylcholine of inhibitor-free controls is depressed and inhibition by Bt 2 cyclic AMP is slightly enhanced. The phospholipase A 2 activity of platelet lysates is increased by the presence of added Ca 2+, however, the addition of either A23187 or Bt 2 cyclic AMP is without effect on this activity. We suggest that Bt 2 cyclic AMP may promote a compartmentalization of Ca 2+, thereby inhibiting phospholipase A activity. The compartmentalization may be overcome by ionophore. By contrast, TMB-8 may immobilize platelet Ca 2+ stores in situ or restrict access of Ca 2+ to phospholipase A in a manner not susceptible to reversal by high concentrations of ionophore.