Platelet-activating factor-induced calcium mobilization in human platelets and neutrophils: effects of PAF-acether...

Since one of the first measurable events that occurs as a consequence of receptor-mediated cell activation is an increase in the cytosolic free calcium concentration, the calcium-selective fluorescent indicator fura-2 was employed to monitor increases in cytosolic calcium following PAF-acether stimulation of human neutrophils and platelets. In neutrophils, approximately 70% of the increase in cytosolic calcium was attributed to release from an intracellular source and 30% could be attributed to enhanced influx through the plasma membrane. In platelets, only 30% was released from an intracellular pool and the remainder reflected enhanced influx of calcium. In both cell types, the intracellular source of calcium was a non-mitochondrial, vesicular compartment, probably endoplasmic reticulum. The ability of different compounds to antagonize the PAF-acether-induced increase in cytosolic calcium was investigated using quin2 and fura-2 as calcium indicators. In decreasing order of potency, the following PAF-acether antagonists inhibited the change in platelet cytosolic free calcium elicited by 10 nM PAF-acether: L-652,731, kadsurenone, triazolam, diltiazem and alprazolam. L-652,731, kadsurenone, triazolam and diltiazem were also tested in neutrophils stimulated by 10 nM PAF-acether. While the antagonists were 7-20 times less active in neutrophils as compared to platelets, they all inhibited the rise in free calcium with the same order of potency in both cell types. Each of the antagonists was PAF-acether-specific, inhibited both the rate and magnitude of calcium mobilization and appeared to exhibit competitive antagonism. These data demonstrate that calcium mobilization can provide a rapid, sensitive and quantitative method by which to evaluate agonists such as PAF-acether and PAF-acether antagonists such as kadsurenone in different cell types.