Nitroso - redox balance in control of coronary vasomotor tone 1 2 3

29 Reactive oxygen species (ROS) are essential in vascular homeostasis but may contribute to vascular 30 dysfunction when excessively produced. Superoxide anion (O2) can directly affect vascular tone by 31 reacting with K channels and indirectly by reacting with nitric oxide (NO) thereby scavenging NO and 32 causing nitroso-redox imbalance. After myocardial infarction (MI), oxidative stress increases, favoring 33 the imbalance and resulting in coronary vasoconstriction. Consequently, we hypothesized that ROS 34 scavenging results in coronary vasodilation, particularly after MI, and is enhanced after inhibition of NO35 production. 36 Chronically instrumented swine were studied at rest and during exercise before and after 37 scavenging of ROS with N-(2-Mercaptoproprionyl)-glycine (MPG, 20 mg/kg I.V.) in the presence or 38 absence of prior inhibition of endothelial NO synthase (eNOS) with N-nitro-L-arginine (LNNA, 20 39 mg/kg I.V.). 40 In normal swine, MPG resulted in coronary vasodilation as evidenced by an increased coronary 41 venous O2 tension, and trends towards increased coronary venous O2 saturation and decreased myocardial 42 O2 extraction. These effects were not altered by prior inhibition of eNOS. In MI swine, MPG showed a 43 significant vasodilator effect, which surprisingly, was abolished by prior inhibition of eNOS. Moreover, 44 eNOS dimer/monomer ratio was decreased after MI, reflecting eNOS uncoupling. 45 In conclusion, ROS exert a small coronary vasoconstrictor influence in normal swine, which does 46 not involve scavenging of NO. This vasoconstrictor influence of ROS is slightly enhanced after MI. Since 47 inhibition of eNOS abolished rather than augmented the vasoconstrictor influence of ROS in swine with 48 MI, while eNOS dimer/monomer ratio was decreased, our data imply that uncoupled eNOS may be a 49 significant source of O2 after MI. 50 51 by 10.2.32.246 on July 1, 2017 http://jaysiology.org/ D ow nladed fom Superoxide and coronary vasomotor tone 3 Introduction 52 Nitric oxide (NO) and superoxide (O2) are key players in cellular nitroso-redox balance and are required 53 for normal vascular homeostasis. The importance of the nitroso-redox balance in the cardiovascular 54 system is underlined by studies that show that oxidative stress, i.e. a disturbance of the nitroso-redox 55 balance contributes to the pathogenesis of diabetes, hypertension and atherosclerosis (1, 16, 38, 49, 50, 56 66). Although several studies have shown that oxidative stress is increased after a myocardial infarction 57 (MI), even in the remote myocardium (7, 9), and that the increased oxidative stress contributes to 58 endothelial dysfunction in isolated large coronary arteries (6), the influence of the increased oxidative 59 stress on the coronary microvasculature in the remote myocardium after MI in vivo has not been 60 investigated to date. 61 Under normal, physiological, conditions O2 is enzymatically produced by a variety of oxidases, 62 including xanthine oxidase and NADPH oxidase and as a byproduct of oxidative phosphorylation in the 63 mitochondria, which is presumed to be the major source of O2 production (18, 67). O2 can affect 64 vascular function either directly, by changing the opening probability of K channels or indirectly, by 65 quenching NO and forming ONOO, both leading to vasoconstriction (60). To prevent the deleterious 66 actions of high concentrations of O2, its concentration is tightly controlled and kept in the picomolar 67 range by superoxide dismutase (SOD) thereby creating H2O2, a membrane-permeable vasodilator (37, 41) 68 that has been suggested to be the factor that couples myocardial metabolism to coronary vasomotor tone 69 (56). 70 After MI, O2 is excessively produced resulting in oxidative stress even in the remote, non71 infarcted myocardium (7, 9), which may result in enhanced coronary vasoconstriction. Given the role of 72 the mitochondrial respiratory chain as a major source of O2, oxidative stress is likely to increase during 73 exercise. An increased O2-mediated vasoconstriction may therefore directly contribute to the relative 74 hypoperfusion of the remote non-infarcted myocardium that is particularly observed during exercise (29). 75 Thus, we hypothesize that the vasoconstrictor effect of O2in the remote, non-infarcted myocardium is 76 enhanced after MI, especially during exercise. To test this hypothesis, we investigated the effects of N-(277 by 10.2.32.246 on July 1, 2017 http://jaysiology.org/ D ow nladed fom Superoxide and coronary vasomotor tone 4 Mercaptoproprionyl)-glycine (MPG), a synthetic aminothiol antioxidant that acts primarily, though 78 perhaps not exclusively (2, 59), through scavenging of O2 (16), in chronically instrumented swine with 79 and without a recent MI. 80 In addition to its direct coronary vasoconstrictor effect, oxidative stress may result in a shift in the 81 nitroso-redox balance towards the formation of peroxinitrate (ONOO-), thereby reducing NO 82 bioavailability, and counteracting the physiological NO-mediated quiescent and dilated state of coronary 83 blood vessels (64). Moreover, O2 can lead to uncoupling of endothelial nitric oxide synthase (eNOS), 84 through oxidation of its cofactor tetrahydrobiopterin (BH4), thereby causing a shift from NO to O2 85 production (14, 31) and further aggravating microvascular dysfunction. We therefore also investigated 86 whether eNOS uncoupling occurred in the remote myocardium after MI, and if the contribution of eNOS87 dependent O2 was altered after MI. 88 89