Endothelial nitric oxide production is tightly coupled to the citrulline-NO cycle

Nitric oxide (NO) is an important vasorelaxant produced along With L-Citrulline from L-arginine in a reaction catalyzed by endothelial nitric oxide synthase (eNOS). Previous Studies suggested that the recycling Of L-Citrulline to L-arginine is essential for NO production in endothelial cells. However, there is no direct evidence demonstrating the degree to which the recycling Of L-citrulline to L-arginine is coupled to NO production. We hypothesized that the amount of NO formed would be significantly higher than the amount of L-Citrulline formed due to the efficiency Of L-Citrulline recycling via the citrulline-NO cycle. To test this hypothesis, endothelial cells were incubated with [C-14]-L-arginine and stimulated by various agents to produce NO. The extent of NO and [C-14]-L-citrulline formation were simultaneously determined. NO production exceeded apparent L-Citrulline formation of the order of 8 to 1, under both basal and stimulated conditions. As further support, alpha-methyl-DL-aspartate, an inhibitor of argininosuccinate synthase (AS), a component of the citrulline-NO cycle, inhibited NO production in a dose-dependent manner. The results of this study provide evidence for the essential and efficient coupling Of L-citrulline recycling, via the citrulline-NO cycle, to endothelial NO production. (c) 2007 Elsevier Inc. All rights reserved.