Mediation of EDHF-induced reduction of smooth muscle [Ca2+](i) and arteriolar dilation by K+ channels, 5,6-EET, and gap junctions

Objective: To characterize the role of K+ channels, the cytochrome P-450 (C YP) metabolite 5.6-EET, and gap junctions in modulation of arteriolar myoge nic tone by a non-nitric oxide nonprostaglandin mediator, termed "endotheli m-independent hyperpolarizing factor" (EDHF). released to acetylcholine (AC h) in skeletal muscle arterioles. Methods: In isolated rat gracilis arterioles. simultaneous changes in smoot h muscle (aSM) [Ca2+](i) (assessed by changes in fura-2 ratiometric signal, R-Ca) and diameter were measured in response to ACh in the presence of ind omethacin and L-NAME. Results: ACh, the K-ATP channel opener pinacidil, and the Ca2+ channel inhi bitor verapamil elicited comparable decreases in aSM [Ca2+](i) (max.: -32 /- 3%. 29 +/- 3%. and -30 +/- 3%, respectively) and arteriolar dilations (m ax.: 90 +/- 4%. 96 +/- 2%. and 95 +/- 2%. respectively). ACh-induced respon ses were inhibited by KCl-depolarization K-Ca channel blockers (TEA. charyb dotoxin), or gap junction inhibitors (18 alpha -glycyrrhetinic acid, hypero smolar sucrose). The channel inhibitor glibenclamide, the K-IR channel inhi bitor barium chloride. or the CYP inhibitor 17-octadecynoic acid (ODYA) wer e without effect. Tbe Putative EDHF analogue 5,6-EET elicited constrictions in the Presence of the endothelium that could be prevented by indomethacin or a TxA(2) receptor antagonist. whereas in the absence of the endothelium , EDHF elicited only small, charybdotoxin-insensitive decreases in aSM R-Ca and dilations (max.: -8 +/- 2% and 27 +/- 4% respectively). Conclusions: In skeletal muscle arterioles. EDHF 1) substantially and rapid ly reduces myogenic tone by decreasing aSM [Ca2+](i) via opening K-Ca chann els, 2) it is unlikely to be 5.6-EET or other CYP metabolites. but 3) requi res functional gap junctions.