Real time measurement of nitric oxide released from cultured endothelial cells

Direct detection of nitric oxide (NO) is essential for understanding the pr ecise mechanism of its production from endothelial cells, Previously, we de veloped an NO detection system based on the chemiluminescence reaction betw een NO and luminol-H2O2. Here, we have applied this system to cultured endo thelial cells for the direct and on-time measurement of NO. The perfusate f rom cultured endothelial cells was continuously mixed with luminol-H2O2. N- G-monomethyl-L-arginine (L-NMMA) (10(-4) M) decreased the chemiluminescence signal of SO, suggesting the existence of basal NO release. Bradykinin (10 (-8) M-10(-6) M) increased the NO signal (10(-6) M; 5.1+/-0.4 fmol/min, cor responding to 1.7 pM in the perfusate), and this was inhibited by 10(-4) M L-NMMA (1.8+/-0.3 fmol/min). These results corresponded to the changes in c GMP levels in RFL-6 cells, which provide an NO bioassay system. We conclude that the luminol-H2O2 system is useful for the direct and continuous measu rement of NO from cultured endothelial cells.