Stimulation of rat erythrocyte P2X7 receptor induces the release of epoxyeicosatrienoic acids

The red blood cell (RBC) is an abundant source of epoxyeicosatrienoic acids (EETs) (Jiang H, et al. JBC 279 :; 2004). Spontaneous release of EETs, 0.66 ± 0.14 ng/10 9 RBCs, occurs during the incubation of rat RBCs in a salt buffer. Released EETs were analyzed by LC/MS: 14,15‐, 11,12‐, 8,9‐ and 5,6‐EET in a ratio of 1.3:1.0:0.9:0.8. EET release was dose‐dependently stimulated by the ATP analog BzATP and inhibited by P2X 2 antagonists, indicating stimulation of the ATP P2X 2 receptor. 1 mM ATP increased the release of EETs by ca . 40% to 0.98 ± 0.16 ng/10 9 RBC. 100 μM BzATP tripled the amount of EET release to 2.32 ± 0.26 ng/10 9 RBCs. Methyl arachidonyl fluorophosphonate, a Ca 2+ ‐dependent cytosolic PLA 2 inhibitor, blocked EET release. Glybenclamide, an inhibitor of the cystic fibrosis transmembrane regulator which is required for ATP release, also inhibited EET release. Stimulating the RBC P2X 2 receptor with either ATP or BzATP was not associated with hemolysis or arachidonic acid release into the buffer. Release of EETs from RBCs was inhibited by the epoxygenase inhibitor PPOH. Thus, the ATP mechanism that releases EETs from RBCs should participate in microcirculatory regulation. The vasodilatory, pro‐fibrinolytic, and antithrombotic activities of EETs indicate that their release from RBCs represents an important circulatory event. Supported by the Philip Morris USA Inc. and Philip Morris International (HJ) and by NIH HL‐25394 (JCM).