Endothelial nitric oxide synthase uncoupling: Is it a physiological mechanism of endothelium-dependent relaxation in cerebral artery?

Nitric oxide (NO) is generated from the conversion of L -arginine to L-citrulline by endothelial nitric oxide synthase (eNOS), which requires Ca/calmodulin, FAD, FMN, and tetrahydrobiopterin (BH4) as cofactors. Chemical studies in vitro demonstrated that the catalytic mechanisms of NOS involve flavin-mediated electron transport from a flavincontaining reductase domain to a heme-containing oxygenase domain. Here, oxygen is reduced and incorporated into the guanidine group of L-arginine, giving rise to NO and Lcitrulline. Calcium/calmodulin binding to NOS increases the rate of reduction of both flavins and the heme iron. Reduction of iron (III) to iron (II) facilitates oxygen binding to the heme group to form a transient ferrous–dioxygen complex. NOS can also produce superoxide under certain conditions. Superoxide is generated from the oxygenase domain by dissociation of the ferrous–dioxygen complex [1,2]. This state is referred to as the “uncoupled state of eNOS” (eNOS uncoupling). BH4 couples L-arginine oxidation to NADPH consumption and prevents dissociation of the ferrous–dioxygen complex. Although in vitro data using purified eNOS demonstrate that modulation of the BH4 concentration may regulate the ratio of superoxide to NO generated by eNOS, it is as yet unclear how the endothelium regulates the production of these radicals in vivo. Although there are some methods to detect and quantify free radicals superoxide as well as NO in chemical systems, their detection in vivo has potential limitations. NO has a variety of functions, but its action as the endothelium-derived relaxing factor (EDRF) is the most important for the