Studies on the mechanism of the inhibition of platelet aggregation and release induced by high levels of arachidonate

Abstract

Abstract We have previously reported that arachidonic acid induced a biphasic pattern of platelet aggregation and the release of both dense and alpha- granule components. Low levels of arachidonate (0.025--0.1 mM) specifically induced aggregation and release, while high concentrations (0.15--0.35 mM) caused a progressive inhibition of these platelet responses in human gel-filtered platelets (GFP). We now report studies of the mechanism(s) responsible for this arachidonate-induced turn-off of platelet function. Electron micrographic studies demonstrated that there was no gross damage to the platelets during the turn-off. Active synthesis of malondialdehyde and thromboxane A2 was seen at the high arachidonate levels, despite the inhibition of aggregation. Furthermore, GFP inhibited by 0.25 mM arachidonate were capable of undergoing aggregation and serotonin release in response to other stimuli, such as collagen or thrombin. Thus, GFP appeared to be metabolically intact and functional during the inhibiton by high arachidonate levels. Thin-layer chromatographic studies revealed that prostaglandin metabolism was not changed at the high arachidonate levels. In addition, indomethacin (20 microM) did not abolish the arachidonate-induced inhibition of platelet function. Therefore, the inhibitory effect of high arachidonate did not depend on its conversion to other prostaglandin products. Platelet cyclic AMP levels increased twofold at the high arachidonate concentrations (1.3 +/- 0.3 pmole/10(8) platelets at peak aggregation, compared with 2.9 +/- 0.4 pmole/10(8) platelets at inhibition by 0.25 mM arachidonate, p less than 0.001). Prostaglandin-D2, a platelet inhibitor known to increase cyclic AMP, generated a similar rise (to 2.4 +/- 0.2 pmole/10(8) platelets). Thus, the magnitude of the arachidonate-induced increase in platelet cyclic AMP levels can account for the inhibition of aggregation and release.