CONTRIBUTION OF PHOSPHOLIPASES A(2) AND D TO ARACHIDONIC-ACID LIBERATION AND PROSTAGLANDIN D-2 FORMATION WITH INCREASE IN INTRACELLULAR CA2+ CONCENTRATION IN RAT PERITONEAL MAST-CELLS

The contribution of phospholipases A(2) (PLA(2)) and D (PLD) activation to arachidonic acid liberation and prostaglandin D-2 (PGD(2)) formation was studied in stimulated rat peritoneal mast cells. Stimulation of the cells with ionomycin induced time-dependent and Ca2+-concentration-dependent increase in arachidonic acid liberation and PGD(2) formation, and the Ca2+-dependent increase was especially remarkable at extracellular Ca2+ concentration higher than 200 mu M. Staurosporine did not induce any effect on the arachidonic acid liberation, indicating that protein kinase C is not involved in the liberation. Addition of ethanol to the cells decreased the ionomycin-stimulated arachidonic acid liberation to 40% of the control, while it decreased the PGD(2) formation almost completely, with the increase in phosphatidylethanol formation. Propranolol, a phosphatidate phosphohydrolase inhibitor, caused similar effects. p-Bromophenacyl bromide, a PLA(2) inhibitor, inhibited partially the arachidonic acid liberation. The inhibition of the liberation by combination of p-bromophenacyl bromide and ethanol was additive and reached approximately 90%. Under the conditions used phromophenacyl bromide did not influence significantly the PLD activity assessed by the phosphatidylethanol formation. Histamine release was decreased by ethanol treatment to 35% of the control. These results suggest that more than half of the total arachidonic acid liberation is mediated by the sequential pathway of PLD/phosphatidate phosphohydrolase/diacylglycerol Lipase and more than half of histamine release is also dependent on PLD activation, while the PGD(2) formation is fully mediated by the pathway. PLA(2) also contributes to arachidonic acid liberation but to a lower extent.