CALL FOR PAPERS Regulation of Cardiovascular Functions by Eicosanoids and Other Lipid Mediators Mechanism of rat mesenteric arterial KATP channel activation by 14,15-epoxyeicosatrienoic acid

Ye, Dan, Wei Zhou, Tong Lu, Setti G. Jagadeesh, John R. Falck, and Hon-Chi Lee. Mechanism of rat mesenteric arterial KATP channel activation by 14,15-epoxyeicosatrienoic acid. Am J Physiol Heart Circ Physiol 290: H1326–H1336, 2006; doi:10.1152/ajpheart.00318.2005.— Recently, we reported that 11,12-epoxyeicosatrienoic acid (11,12-EET) potently activates rat mesenteric arterial ATP-sensitive K (KATP) channels and produces significant vasodilation through protein kinase A-dependent mechanisms. In this study, we tried to further delineate the signaling steps involved in the activation of vascular KATP channels by EETs. Whole cell patch-clamp recordings [0.1 mM ATP in the pipette, holding potential (HP) 0 mV and testing potential (TP) 100 mV] in freshly isolated rat mesenteric smooth muscle cells showed small glibenclamide-sensitive KATP currents (19.0 7.9 pA, n 5) that increased 6.9-fold on exposure to 5 M 14,15-EET (132.0 29.0 pA, n 7, P 0.05 vs. control). With 1 mM ATP in the pipette solution, KATP currents (HP 0 mV and TP 100 mV) were increased 3.5-fold on exposure to 1 M 14,15-EET (57.5 14.3 pA, n 9, P 0.05 vs. baseline). In the presence of 100 nM iberiotoxin, 1 M 14,15-EET hyperpolarized the membrane potential from 20.5 0.9 mV at baseline to 27.1 3.0 mV (n 6 for both, P 0.05 vs. baseline), and the EET effects were significantly reversed by 10 M glibenclamide ( 21.8 1.4 mV, n 6, P 0.05 vs. EET). Incubation with 5 M 14,15-epoxyeicosa-5(Z)-enoic acid (14,15-EEZE), a 14,15-EET antagonist, abolished the 14,15-EET effects (31.0 11.8 pA, n 5, P 0.05 vs. 14,15-EET, P not significant vs. control). The 14,15-EET effects were inhibited by inclusion of anti-Gs antibody (1:500 dilution) but not by control IgG in the pipette solution. The effects of 14,15-EET were mimicked by cholera toxin (100 ng/ml), an exogenous ADP-ribosyltransferase. Treatment with the ADP-ribosyltransferase inhibitors 3-aminobenzamide (1 mM) or m-iodobenzylguanidine (100 M) abrogated the effects of 14,15-EET on KATP currents. These results were corroborated by vasodilation studies. 14,15-EET dose-dependently dilated isolated small mesenteric arteries, and this was significantly attenuated by treatment with 14,15-EEZE or 3-aminobenzamide. These results suggest that 14,15-EET activates vascular KATP channels through ADP-ribosylation of Gs .