Received for publication June 14, 2006.
Revised August 1, 2006.
Accepted for publication August 1, 2006.

ELECTRON SPIN RESONANCE AND CHEMILUMINESCENCE ANALYSES TO ELUCIDATE THE VASODILATING MECHANISM OF SODIUM NITROPRUSSIDE

Abstract

Aim of the study was to elucidate the vasodilating mechanism of sodium nitroprusside (SNP). To do this, SNP was i.v. infused in pigs (1.67 µmoles/Kg) and the following paramagnetic metabolites identified by ESR: i) nitrosylhemoglobin (HbFe(II)NO) as an index of the bioconservative pathway; ii) transferrin; iii) [Fe(II)(CN)₅NO]^3- and [Fe(II)(CN)₄NO]^2-, the reduced penta- and tetra-coordinated intermediates of SNP; iii) methemoglobin (Met-Hb). The results indicate that: a) 17 % of the dose is converted to HbFe(II)NO at the end of infusion; b) NO administered as SNP does not undergo bioinactivation (oxidative metabolism) since no significant increase of met-Hb was observed b) the equilibrium involving the paramagnetic species of SNP is shifted towards HbFe(II)NO, since a significant increase of transferrin but no detection of the reduced paramagnetic intermediates of SNP was observed. The results obtained indicate that the hemodynamic effect induced by SNP is not mediated by HbFe(II)NO, at least under physiological conditions; hence a direct release of NO form SNP in the vascular target should be considered. To demonstrate this mechanism, endothelial cells were incubated with SNP and the release of NO determined by a novel chemiluminescence method. The results indicate that the endothelium is able to metabolize SNP, with the formation of stoichiometric amounts of NO. In conclusion, SNP is rapidly metabolized to HbFe(II)NO but the pharmacological response is mediated by a direct mechanism of NO release of the parent compound at the cellular target.

Key words: Nitric oxide, Structure/function/mechanism