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In catalyzing the reversible hydration of CO2 to bicarbonate and protons, the ubiquitous enzyme carbonic anhydrase (CA) plays a crucial role in CO2 transport, in acid-base balance, and in linking local acidosis to O2 unloading from hemoglobin. Considering the structural similarity between bicarbonate and nitrite, we hypothesized that CA uses nitrite as a substrate to produce the potent vasodilator nitric oxide (NO) to increase local blood flow to metabolically active tissues. Here we show that CA readily reacts with nitrite to generate NO, particularly at low pH, and that the NO produced in the reaction induces vasodilation in aortic rings. This reaction occurs under normoxic and hypoxic conditions and in various tissues at physiological levels of CA and nitrite. Furthermore, two specific inhibitors of the CO2 hydration, dorzolamide and acetazolamide, increase the CA-catalyzed production of vasoactive NO from nitrite. This enhancing effect may explain the known vasodilating effects of these drugs and indicates that CO2 and nitrite bind differently to the enzyme active site. Kinetic analyses show a higher reaction rate at high pH, suggesting that anionic nitrite participates more effectively in catalysis. Taken together, our results reveal a novel nitrous anhydrase enzymatic activity of CA that would function to link the in vivo main end products of energy metabolism (CO2/H+) to the generation of vasoactive NO. The CA-mediated NO production may be important to the correlation between blood flow and metabolic activity in tissues, as occurring for instance in active areas of the brain.

Acetylcholine stimulates endothelial cells leading to relaxation of the underlying smooth muscle cells either through gap junction transmission or release of various diffusible substances. The identification of nitric oxide (NO) as an endothelium-derived relaxing factor (EDRF) rests on observations interfering with the ¬_arginine-NO pathway, as well as the physiological similarities between the endogenous substance and authentic NO. Thus pharmacological inhibition of the ¬_arginine-NO pathway in vivo and knockout of the gene for the endothelial cell constitutive nitric oxide synthase (NOS) raises the blood pressure (Huang et al. 1995), reduces acetylcholine-induced relaxations in vitro (Cohen & Vanhoutte, 1995;Huang et al. 1995), and reduces the production of the end-products of NO metabolism, nitrite (NOµ¦) and nitrate (NOצ) (Ignarro et al. 1993). EDRF and NO appear to cause comparable relaxations in bioassays (Palmer et al. 1987;Feelisch et al. 1994). Moreover, NO release has been measured with chemiluminescence (Palmer et al. 1987), by conversion of oxyhaemoglobin to methaemoglobin (Kelm & Schrader, 1990), and recently by the use of either membrane-covered electrodes (Goligorsky et al. 1994) or polarographic electrodes (Shibuki & Okada, 1991), which allow direct measurements of NO released from cell cultures (Malinski & Taha, 1992) and unmounted arteries (Cohen et al. 1997). Thus the evidence appears incontrovertible that NO participates in endothelium-dependent relaxations, but other endothelium-derived relaxing factors could be more important, since evidence for a direct relationship between endothelium-derived NO and acetylcholine-induced relaxation is lacking. Endothelium-dependent relaxation of vascular smooth muscle involves in some cases hyperpolarization of the cell membrane, and has been attributed to a diffusible endothelium-derived hyperpolarizing factor (EDHF) (Murphy

Introduction Multiple regulatory systems are involved in normal erectile function. Disruption of psychological, neurological, hormonal, vascular, and cavernosal factors, individually, or in combination, can induced erectile dysfunction (ED). The contribution of neurogenic, vascular, and cavernosal factors was thoroughly reviewed by our committee, while psychological and hormonal factors contributing to ED were evaluated by other committees. Aim To provide state of the art knowledge on the physiology of ED. Methods An international consultation in collaboration with the major urology and sexual medicine associations assembled over 200 multidisciplinary experts from 60 countries into 17 committees. Committee members established specific objectives and scopes for various male and female sexual medicine topics. The recommendations concerning state-of-the-art knowledge in the respective sexual medicine topic represent the opinion of experts from five different continents developed in a process over a 2-year period. Concerning the pathophysiology of ED committee, there were seven experts from five different countries. Main Outcome Measure Expert opinion was based on the grading of evidence-based medical literature, widespread internal committee discussion, public presentation, and debate. Results The epidemiology and classification of neurogenic ED was reviewed. The evidence for the association between vascular ED and atherosclerosis/hypercholesterolemia, hypertension and diabetes was evaluated. In addition, the pathophysiological mechanisms implicated in vascular ED were defined, including: arterial remodeling, increased vasoconstriction, impaired neurogenic vasodilatation, and impaired endothelium-dependent vasodilatation. The possible mechanisms underlying the association between chronic renal failure and ED were also evaluated as well as the evidence supporting the association of ED with various classes of medications. Conclusions A better understanding of how diseases interfere with the physiological mechanisms that regulate penile erection has been achieved over the last few years, which helps establish a strategy for the prevention and treatment of ED.

Erectile dysfunction (ED) and coronary artery disease (CAD) overlap in risk factors, aetiology and clinical outcomes. It has become clear that ED is an important marker of vascular disease throughout the arterial tree--including CAD, stroke and diabetes. Epidemiological studies have demonstrated a close association between ED and vascular disease. The shared aetiological factor is endothelial dysfunction. The fact that ED tends to precede the onset of symptoms of other vascular diseases--because blood vessels in the penis are narrower in diameter than elsewhere in the body so blood flow is restricted sooner by atherosclerosis--means that it can be used as a 'window' on vascular health. There is growing evidence that patients presenting with ED should be investigated for cardiovascular disease (CVD), including diabetes, even if they have no symptoms. Early detection could facilitate prompt intervention and a reduction in long-term complications. Treatments that reduce endothelial dysfunction offer the potential of improving the functioning of the entire vascular system, improving outcomes in CVD and diabetes, as well as providing effective treatment for ED.

The reduction of circulating nitrite to nitric oxide (NO) has emerged as an important physiological reaction aimed to increase vasodilation during tissue hypoxia. Although hemoglobin, xanthine oxidase, endothelial NO synthase, and the bc(1) complex of the mitochondria are known to reduce nitrite anaerobically in vitro, their relative contribution to the hypoxic vasodilatory response has remained unsolved. Using a wire myograph, we have investigated how the nitrite-dependent vasodilation in rat aortic rings is controlled by oxygen tension, norepinephrine concentration, soluble guanylate cyclase (the target for vasoactive NO), and known nitrite reductase activities under hypoxia. Vasodilation followed overall first-order dependency on nitrite concentration and, at low oxygenation and norepinephrine levels, was induced by low-nitrite concentrations, comparable to those found in vivo. The vasoactive effect of nitrite during hypoxia was abolished on inhibition of soluble guanylate cyclase and was unaffected by removal of the endothelium or by inhibition of xanthine oxidase and of the mitochondrial bc(1) complex. In the presence of hemoglobin and inositol hexaphosphate (which increases the fraction of deoxygenated heme), the effect of nitrite was not different from that observed with inositol hexaphosphate alone, indicating that under the conditions investigated here deoxygenated hemoglobin did not enhance nitrite vasoactivity. Together, our results indicate that the mechanism for nitrite vasorelaxation is largely intrinsic to the vessel and that under hypoxia physiological nitrite concentrations are sufficient to induce NO-mediated vasodilation independently of the nitrite reductase activities investigated here. Possible reaction mechanisms for nitrite vasoactivity, including formation of S-nitrosothiols within the arterial smooth muscle, are discussed.

1 The purpose of this study was to investigate whether a membrane-permeable superoxide dismutase mimetic, tempol, added either alone or in combination with the nitric oxide (NO) donor molsidomine, prevents the development of pulmonary hypertension (PH) in chronic hypoxic rats. 2 Chronic hypobaric hypoxia (10% oxygen) for 2 weeks increased the right ventricular systolic pressure (RVSP), right ventricle and lung wet weight. Relaxations evoked by acetylcholine (ACh) and the molsidomine metabolite SIN-1 were impaired in isolated proximal, but not distal pulmonary arteries, from chronic hypoxic rats. 3 Treatment with tempol (86 mg kg À1 day À1 in drinking water) normalized RVSP and reduced right ventricular hypertrophy, while systemic blood pressure, lung and liver weights, and blunted ACh relaxation of pulmonary arteries were unchanged. 4 Treatment with molsidomine (15 mg kg À1 day À1 in drinking water) had the same effects as tempol, except that liver weight was reduced, and potassium and U46619-evoked vasoconstrictions in pulmonary arteries were increased. Combining tempol and molsidomine did not have additional effects compared to tempol alone. ACh relaxation in pulmonary arteries was not normalized by these treatments. 5 The media to lumen diameter ratio of the pulmonary arteries was greater for the hypoxic rats compared to the normoxic rats, and was not reversed by treatment with tempol, molsidomine, or the combination of tempol and molsidomine. 6 We conclude that tempol, like molsidomine, is able to correct RVSP and reduce right ventricular weight in the rat hypoxic model. Functional and structural properties of pulmonary small arteries were little affected. The results support the possibility that superoxide dismutase mimetics may be a useful means for the treatment of PH.

BACKGROUND AND PURPOSE In rodents, the endothelial KCa channels, KCa3.1 and KCa2.3, have been shown to play a crucial role in initiating endothelium‐derived hyperpolarizing factor (EDHF) vasodilator responses. However, it is not known to what extent these channels are involved in blood pressure regulation in large mammals, which would also allow us to address safety issues. We therefore characterized canine endothelial KCa3.1 and KCa2.3 functions and evaluated the effect of the KCa3.1/KCa2.3 activator SKA‐31 on blood pressure and heart rate in dogs. EXPERIMENTAL APPROACH Canine endothelial KCa3.1/KCa2.3 functions were studied by patch‐clamp electrophysiology and wire myography in mesenteric arteries. Systemic cardiovascular actions of acute SKA‐31 administration were monitored in conscious, unstressed beagle dogs. KEY RESULTS Mesenteric endothelial cells expressed functional KCa3.1 and KCa2.3 channels that were strongly activated by SKA‐31. SKA‐31 hyperpolarized the endothelial membrane and doubled endothelial hyperpolarization‐dependent vasodilator responses in mesenteric arteries. SKA‐31 (2 mg·kg−1, i.v.) rapidly decreased the MAP by 28 ± 6 mmHg; this response was transient (8 ± 1 s), and the initial drop was followed by a fast and pronounced increase in HR (+109 ± 7 beats min−1) reflecting baroreceptor activation. SKA‐31 significantly augmented similar transient depressor responses elicited by ACh (20 ng·kg−1) and doubled the magnitude of the response over time. CONCLUSIONS AND IMPLICATIONS Activation of endothelial KCa3.1 and KCa2.3 lowers arterial blood pressure in dogs by an immediate electrical vasodilator mechanism. The results support the concept that pharmacological activation of these channels may represent a potential unique endothelium‐specific antihypertensive therapy.