In eNOS knockout mice skeletal muscle arteriolar dilation to acetylcholine is mediated by EDHF

The mechanisms that account for acetylcholine (ACh)-induced responses of skeletal muscle arterioles of mice lacking endothelial nitric oxide (NO) synthase (eNOS-KO) were investigated. Isolated, cannulated, and pressurized arterioles of gracilis muscle from male eNOS-KO (74.1 ± 2.3 μm) and wild-type (WT, 87.2 ± 2.1 μm) mice developed spontaneous tone accounting for 63 and 61% of their passive diameter (116.8 ± 3.4 vs. 143.2 ± 2.8 μm, respectively) and dilated dose-dependently to ACh (10− 9-10− 7M). These dilations were significantly smaller in vessels of eNOS-KO compared with WT mice (29.2 ± 2.0 μm vs. 46.3 ± 2.1 μm, at maximum concentration) but responses to the NO donor, sodium nitrite (NaNO2, 10− 6-3 × 10− 5 M), were comparable in the vessels of the two strains. N G-nitro-l-arginine (l-NNA, 10− 4 M), an inhibitor of eNOS, inhibited ACh-induced dilations by 60–90% in arterioles of WT mice but did not affect responses in those of eNOS-KO mice. In arterioles of eNOS-KO mice, dilations to ACh were not affected by indomethacin but were essentially abolished by inhibitors of cytochrome P- 450, clotrimazole (CTZ, 2 × 10− 6 M) or miconazole (MCZ, 2 × 10− 6 M), as well as by either high K+ (40 mM) or iberiotoxin [10− 7 M, a blocker of Ca2+-dependent K+ channels (KCachannels)]. On the other hand, in WT arterioles CTZ or MCZ inhibited ACh-induced dilations only by ∼10% and only in the presence of l-NNA. These results indicate that in arterioles of eNOS-KO mice, endothelium-derived hyperpolarizing factor (EDHF), synthesized via cytochrome P -450, accounts entirely for the mediation of ACh-induced dilation via an increase in KCa-channel activity. In contrast, in arterioles of WT mice, endothelium-derived NO predominantly mediates ACh-induced dilation in which participation of EDHF becomes apparent only after inhibition of NO synthesis.