CELL-FREE HEMOGLOBIN POTENTIATES ACETYLCHOLINE-INDUCED CORONARY VASOCONSTRICTION IN RABBIT HEARTS

Cell-free hemoglobin (Hb) preparations have been shown to alter vascular tone in vitro and in vivo. The high affinity of Hb for nitric oxide, the putative endothelium-derived relaxing factor (EDRF), may be primarily responsible for this activity, but the contribution of tissue-damaging oxygen-derived free radicals has not been established. We investigated the effects of human Hb interdimerically cross-linked with bis-(3,5-dibromosalicyl)fumarate (alphalphaHb) on the coronary vasomotor response to acetylcholine (ACh) in isolated perfused rabbit hearts. Infusion of 0.1 g/dl alphaalphaHb altered the dose-dependent response to ACh, decreasing the calculated IC50 (ACh concn at which coronary pressure is 50% of its maximal value) from 3.96 +/- 0.34 to 0.85 +/- 0.06 muM (P < 0.01). This augmented sensitivity to ACh was only partially reversed upon washout of alphaalphaHb (IC50 1.93 +/- 0.13 muM). Simultaneous infusion of 60 muM deferoxamine mesylate with alphaalphaHb attenuated this response (IC50 decreased from 3.86 +/- 0.27 to 1.73 +/- 0.38 muM), which was completely reversed after removal of alphaalphaHb (IC50 3.41 +/- 0.17 muM). N(G)-nitro-L-arginine methyl ester (50 muM) and cross-linked cyanomethemoglobin (CNmetalphalphaHb, 0.1 g/dl) induced a significant (P < 0.05) increase in ACh-induced vasoconstriction accompanied by a reduction in myocardial functions in the same range as that caused by alphaalphaHb. Infusion of deferoxamine mesylate (60 muM) with CNmetalphalphaHb completely prevented the reduction in IC50 elicited by the infusion of CNmetalphaalphaHb alone. These data demonstrate that alphaalphaHb can alter coronary vasomotor responsiveness and suggest the involvement of at least two mechanisms, one that is related to an accessible ferrous heme and is reversible and another that does not require an open heme site and is irreversible.