Relative contribution of eNOS and nNOS to endothelium-dependent vasodilation in the mouse aorta

Abstract

In large vessels, endothelium-dependent vasodilation is mainly attributed to endothelial nitric oxide synthase (eNOS)-derived NO production. However, we have recently shown that neuronal nitric oxide synthase (nNOS)-derived H(2)O(2) is also an endothelium-dependent relaxing factor in the mouse aorta. The relative contribution of nNOS/eNOS, H(2)O(2)/NO remains to be characterized. This work was undertaken to determine the relative contribution of NO versus H(2)O(2), and eNOS versus nNOS to endothelium-dependent vasodilation in the mouse aorta. We used carbon microsensors placed next to the lumen of the vessels to simultaneously measure NO, H(2)O(2) and vascular tone. Acetylcholine produced a concentration-dependent increase in NO and H(2)O(2) production with a good coefficient of linearity with acetylcholine-induced relaxation (R(2)=0.93 and 0.96 for NO and H(2)O(2), respectively). L-NAME, a non-selective inhibitor of nitric oxide synthase, abolished NO and H(2)O(2) production, and impaired vasodilation. Selective pharmacological inhibition of nNOS with L-Arg(NO2)-L-Dbu-NH(2) 2TFA and specific knock-down of nNOS abrogated H(2)O(2) and decreased by half acetylcholine-induced vasodilation. Catalase, which specifically decomposes H(2)O(2), did not interfere with NO, but impaired H(2)O(2) and decreased vasodilation to the same level as those obtained with nNOS inhibition or knocking down. Specific knocking down of eNOS had no effect on H(2)O(2) production but greatly reduced NO and decreased vasodilation to levels similar to those found with nNOS inhibition. In eNOS knocked-down mice, pharmacological nNOS inhibition dramatically reduced H(2)O(2) production and further reduced the residual acetylcholine-induced vasodilation. It is concluded that nNOS/eNOS and H(2)O(2)/NO both contribute in a significant way to relaxation in the mouse aorta.