On the mechanism of NO release from sydnonimines.
The vasodilator and antiaggregatory properties of sydnonimines like SIN-1 are thought to be due to their marked stimulatory action on soluble guanylate cyclase. Enzyme activation and consecutive cyclic GMP accumulation is mediated by the liberation of nitric oxide (NO) from the open-ring A forms of sydnonimines. The purpose of the present study was to investigate the mechanism of NO release from sydnonimines in direct comparison to their stimulatory effect at the target enzyme, soluble guanylate cyclase. All sydnonimines tested were found to spontaneously liberate NO, the rate of which closely correlated with the extent of enzyme activation. NO release occurred nonlinearly with time and became maximal at high sydnonimine concentration. The in vitro stability of the A forms neither correlated with the measured rate of NO release nor with enzyme activation, indicating that a direct stimulation of guanylate cyclase by the A forms is rather unlikely. Besides NO, all sydnonimines generated NO2- and NO3- at a nearly equimolar rate. The addition of cysteine induced a marked shift from NO3- to NO2- with a small reduction in NO release, which is paralleled by a weak rightward shift of the EC50 at the guanylate cyclase. All tested sydnonimines were found to consume molecular oxygen at rates that closely corresponded to the measured rates of NO formation. By a molar comparison, the amounts of consumed oxygen are clearly higher, as would be expected for the oxidative conversion of NO to NO2- and NO3-. Oxygen seems to be additionally involved in the induction of NO formation while being converted to superoxide (O2-). In accordance with an autocatalytic process, O2- further enhances sydnonimine decomposition, since in the presence of superoxide dismutase (SOD) the rate of SIN-1C and NO2-/NO3- formation from SIN-1A was reduced, whereas the rate of NO liberation seemingly increased. O2- has, however, no influence on the rate of hydrolysis of SIN-1 to SIN-1A. At the level of guanylate cyclase, the presence of SOD induced a leftward shift of the concentration-response curve to SIN-1, in agreement with an enhancement of efficacy of NO by blocking the NO-scavenging effect of O2-. An additional O2- generation markedly enhanced SIN-1A decomposition to NO2-/NO3- and reduced the apparent rate of NO formation. We conclude from our results that oxygen plays a key role in the decomposition of sydnonimines and thus in the formation of NO as their pharmacodynamically active principle. Oxygen attack most probably occurs by one-electron abstraction from the A form of the respective sydnonimine compound.(ABSTRACT TRUNCATED AT 400 WORDS)
Feelisch
M
M
Institute of Pharmacology, Heinrich Heine University, Düsseldorf, F.R.G.
Ostrowski
J
J
Noack
E
E
D000818:Animals; D004789:Enzyme Activation; D006162:Guanylate Cyclase / Q000378:metabolism; D008099:Liver / Q000378:metabolism; D008981:Molsidomine / Q000031:analogs & derivatives / Q000493:pharmacokinetics*; D009566:Nitrates / Q000493:pharmacokinetics; D009569:Nitric Oxide / Q000493:pharmacokinetics*; D009573:Nitrites / Q000493:pharmacokinetics; D010101:Oxygen Consumption; D051381:Rats; D013481:Superoxides / Q000378:metabolism
D009566:Nitrates; D009573:Nitrites; D013481:Superoxides; D009569:Nitric Oxide; D008981:Molsidomine; D006162:Guanylate Cyclase