Decreased Arteriolar Tetrahydrobiopterin is Linked to Superoxide Generation from Nitric Oxide Synthase in Mice Fed High Salt

Microcirculation (2010) 17 , 1–11. doi: 10.1111/j.1549‐8719.2009.00014.x Abstract Objective:  Impaired endothelium‐dependent arteriolar dilation in mice fed high salt (HS) is due to local oxidation of nitric oxide (NO) by superoxide anion (O 2 − ). We explored the possibility that “uncoupled” endothelial nitric oxide synthase (eNOS) is the source of this O 2 − . Methods:  Levels of L‐arginine (L‐Arg), tetrahydrobiopterin (BH 4 ), and O 2 − (hydroethidine oxidation) were measured in spinotrapezius muscle arterioles of mice fed normal salt (0.45%, NS) or (4%, HS) diets for 4 weeks, with or without dietary L‐Arg supplementation. The contribution of NO to endothelium‐dependent dilation was determined from the effect of N ω ‐nitro‐L‐arginine methyl ester (L‐NAME) on responses to acetylcholine (ACh). Results:  Arterioles in HS mice had lower [BH 4 ] and higher O 2 − levels than those in NS mice. ACh further increased arteriolar O 2 − in HS mice only. L‐Arg supplementation prevented the reduction in [BH 4 ] in arterioles of HS mice, and O 2 − was not elevated in these vessels. Compared to NS mice, arteriolar ACh responses were diminished and insensitive to L‐NAME in HS mice, but not in HS mice supplemented with L‐Arg. Conclusions:  These findings suggest that eNOS uncoupling due to low [BH 4 ] is responsible for O 2 − generation and reduced NO‐dependent dilation in arterioles of mice fed a HS diet.