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1. Nitric oxide (NO) relaxes vascular smooth muscle (VSM) by mechanisms which are not fully understood. One possibility is that NO hyperpolarizes membranes, thereby diminishing Ca2+ entry through voltage-dependent Ca2+ channels. In the current study, the effects of NO on membrane potential of rabbit mesenteric arteries were recorded using intracellular microelectrodes. 2. NO, released by 3-morpholinosydnonimine (SIN-1, 3/M), reversibly hyperpolarized arteries by -9-5 + 4 0 mV (means + S.D., n = 97) from a resting membrane potential of -53-1 + 5-7 mV. The hyperpolarization was blocked by oxyhaemoglobin (20 /M), and only occurred in arteries pre-treated with NW-nitro-L-arginine (100 /M) or denuded of endothelium. 3. The effect of SIN-1 was concentration dependent (EC50 ; 0 4/uM), and its dose response was shifted to the left by zaprinast (100 uM), an inhibitor of cGMP-specific phosphodiesterases.4. The hyperpolarization due to SIN-1 was modified by changes in extracellular K+ concentration, but not by changes in Ca2+ Nae or C1-. The hyperpolarization was blocked by glibenclamide (IC50 ; 015 ,M), but not by apamin (3-300 nM), barium (5-150 uM), tetraethylammonium (01-10 mM), or 4-aminopyridine (5-500 uM). The hyperpolarization due to lemakalim (0-03-3juM), an activator of ATP-sensitive potassium channels (KATP), displayed the same sensitivities to these K+ channel blocking agents, whereas the endothelium-derived hyperpolarizing factor, triggered by the addition of acetylcholine (3 ,M), caused a hyperpolarization (-15-3 + 6-2 mV) that was blocked by apamin, but not by any other agent. 5. These results suggest that NO hyperpolarizes VSM in rabbit mesenteric arteries by activating KATP channels, with the accumulation of cGMP as an intermediate step.

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The aim of this study was to examine whether extreme endurance stress of trained athletes can influence lipid peroxidation and muscle enzymes. A randomized and placebo-controlled study was carried out on 24 trained long-distance runners who were substituted with alpha-tocopherol (400 I.U. d-1) and ascorbic acid (200 mg d-1) during 4.5 weeks prior to a marathon race. The serum concentrations of retinol, ascorbic acid, beta-carotene, alpha-tocopherol, malondialdehyde (TBARS) and uric acid as well as glutathione peroxidase (GSH Px) and catalase were measured 4.5 weeks before (A), immediately before (B), immediately after (C) and 24 h after (D) the course. After competition (C) TBARS serum concentrations of the athletes (n = 22) decreased in both groups (P < 0.0001). The ascorbic acid serum concentration increased significantly in the supplemented group from (A) to (B) (P < 0.01), from (B) to (C) (P < 0.001) and in the placebo group a significant increase from (B) to (C) (P < 0.01) was observed. The alpha-tocopherol serum concentration increased significantly in the supplemented group from (A) to (B) (P < 0.001) and from (B) to (C) (P < 0.05). The enzymes glutathione peroxidase (GSH Px) and catalase measured in erythrocytes as well as the serum selenium levels did not show significant differences at any time. A significant increase of CK concentration was observed from (C) to (D) in the supplemented group (P < 0.01) and in the placebo group (P < 0.001). The increase of CK serum concentration is remarkably lower in the supplemented group compared with the placebo group (P < 0.01). It is concluded that endurance training coupled with antioxidant vitamin supplementation reduces blood CK increase under exercise stress.

Superoxide dismutase (SOD) rapidly scavenges superoxide (O°) and also prolongs the vasorelaxant effects of nitric oxide (NO), thought to be the endotheliumderived relaxing factor. This prolongation has been ascribed to prevention of the reaction between°2-with NO. We report that SOD supports a reversible reduction of NO to NO-. When cyanamide and catalase were used to generate NO-in the presence of SOD, NO was measured by the conversion of HbO2 to MetHb. When SOD[Cu(I)] was exposed to NO anaerobicafly, NO-was trapped by MetHb forming nitrosylmyoglobin. When NO was generated by 3-morpholinosydnonimine hydrochloride in the presence of SOD, NO-or a similar reductant was formed, which reduced catalase compound II and promoted the formation of the catalase[Fe(H)J-NO complex. It is, therefore, conceivable that SOD may protect NO and endothelium-derived relaxing factor by a mechanism in addition toO°scavenging and that NO-may be a physiologically important form of endothelium-derived relaxing factor.Nitric oxide is thought to be the endothelium-derived relaxing factor (EDRF), a vasodilator produced from arginine. The similar chemical properties of NO and EDRF, including their apparent reactivity with°2, support this proposal (1-4). The reaction of NO with O2 (reactions la and b) has been directly observed by using pulse-radiolysis (5), but evidence of the reaction of EDRF with°2 is indirect. Namely, superoxide dismutase (SOD) prolongs the effects of both EDRF and exogenous NO, whether or not there is a simultaneous addition of compounds thought to generate O°. This consistent effect of SOD has been attributed (1-4) to its known ability to catalyze 0°dismutation (6).One report (5) suggests the rate constant for reaction la may be much slower than diffusion limited (-56 x 106 M-1 s-1 at 370C). Also, the intracellular concentration of°2 is estimated to be quite low [e.g., in liver <66 pM (7) Analyses of the metabolites of NO in vivo also fail to prove that reaction la is significant. Although peroxynitrite (ONOO-) can decompose to NO2 and 0OH in the absence of SOD (9) or may convert to NO' in its presence (10), nitrate (NO-) is still a major product of reaction la (5, 9). Therefore, when reaction la is significant, SOD should not only protect NO but should also decrease NO production in favor of NO and NO-. However, product analyses have failed to meet the latter expectation (11, 12) and led us to examine reactions involving SOD and NO in more detail. METHODS 3-Morpholinosydnonimine hydrochloride (SIN-1) was from Cassella AG (Frankfurt), CuZn-SOD (from bovine erythrocytes), cyanamide, and hydroxylamine were from Sigma, and catalase (from bovine liver) was from Boehringer Mannheim. HbO2 was prepared by reduction of MetHb (Aldrich) with dithionite, oxygenation, and purification on a Sephadex G-25 column (in Krebs/Hepes buffer containing 140mM NaCl, 2.7 mM KCl, 0.42 mM NaH2PO4, 1 mM MgCl2, 1.8 mM CaCI2, 20 mM Hepes, adjusted to pH 7.4 at 37°C by adding NaOH) and measured at 415 nm (e = 131 mM-1-cm-1) (13). Metmyoglob...