Development and Biological Application of Highly Sensitive Detection Method for Endothelial Relaxing Factor : NO

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Nitric oxide (NO) reacts with H_2O_2 to produce a rise in potent luminol dependent chemiluminescence. NO at pM levels was detectable by this newly developed aquachemiluminescence system. The chemiluminescence is based on the formation of peroxynitrite in this reaction. I applied this reaction to the continuous detection of NO released from the rat isolated perfused kidney with simultaneous pressure changes. In Wister rats, basel NO release was 85* 11 fmol/min/g kidney weight and acetylcholine (ACh) increased NO release dose-dependently with the pressure reduction. An NO synthesis inhibitor, N^G-monomethyl-L-arginine, abolished the chemiluminescence. This inhibition was reversed by the addition of L-arginine. These changes in the NO release were always associated with mirror image changes in the perfusion pressure. Chemial endothelial denudation diminished the responses to ACh, however, infusion of authentic NO solution caused an increase in the chemiluminescence and a concomitant decrease in the pressure. the infusion of SOD caused no significant change in either perfusion pressure or chemiluminescence. Papaverine reduced the pressure but did not influence the chemiluminescence. NO derivatives like nitrite and nitrate showed no emission. thio-nitroso compounds exhibited emission 104 times* less than NO.Compounds which are biological active on the endothelium or endothelium derived compounds exhibited no emmision. These results suggest that this method is highly sensitive and useful for detection of NO and that this system may provide a new method of evaluating the pathophysiological roles of NO.