Ex Vivo Measurement of Brain Tissue Nitrite and Nitrate Accurately Reflects Nitric Oxide Synthase Activity In Vivo

Abstract: The ex vivo tissue concentration of nitrite and nitrate (NO_x) was found to correlate closely with the activity of nitric oxide synthase (NOS; EC 1.14.13.39) in various brain regions. Systemic administration of the nonselective NOS inhibitor N ^ω-nitro- l -arginine ( l -NA) at doses that completely inhibited both central and peripheral NOS, depleted whole-brain and CSF NO_x by up to 75% but had no effect on plasma NO_x. Selective inhibition of central NOS by intracerebroventricular administration of l -NA methyl ester produced similar decreases in levels of whole-brain NO_x. A residual concentration of NO_x of 10–15 µ M remained in all brain regions even after complete inhibition of brain NOS. Brain NO_x content decreased rapidly and in parallel with the inhibition of brain NOS. The ex vivo measurement of levels of brain NO_x was found to reflect the in vivo efficacy of several different types of NOS inhibitor: l -NA, N ^ω-monomethyl- l -arginine, and 7-nitroindazole. Intraperitoneal administration of the NOS substrate l -arginine increased brain NO_x concentrations by up to 150% of control values. These results demonstrate that the ex vivo measurement of levels of brain tissue NO_x is a rapid, reliable, and straightforward technique to determine NOS activity in vivo. This method can be used to assess both the regional distribution and the degree of inhibition of NOS activity in vivo.