Angiotensin II as the link between nitric oxide and neuroadrenergic function

In recent years, endothelium has gained overwhelming relevance as an organ that is deeply involved in the pathophysiology of cardiovascular disease. Endothelial cells can produce different relaxing substances, including nitric oxide (NO) that is derived from the transformation of L-arginine into citrulline by the activity of the constitutive endothelial enzyme NO synthase (eNOS) [1]. NO plays a major role in the regulation of vascular homeostasis because NO availability is crucial not only to maintain vessels in a constant state of relaxation, but also in inhibiting platelet aggregation [2], vascular smooth muscle cell proliferation [3] and migration [4], monocyte adhesion [5] and adhesion molecule expression [6], thereby protecting vascular wall from the development of atherosclerosis and thrombosis. Major cardiovascular risk factors, including ageing, hypertension and hypercholesterolemia, can alter endothelial cell function and morphology [7]. One of the peculiar features of an altered endothelium is the so-called endothelial dysfunction, characterized by impaired NO availability, which should be distinguished from the endothelial activation, characterized by the acquisition of inflammatory activity. However, this differentiation can be regarded as semantic because reduced NO availability can activate several pathways involved in endothelial cell activation, whereas inflammation can contribute to the NO breakdown by the production of oxidative stress [8]. It is now universally accepted that endothelial dysfunction represents the leading mechanism promoting atherosclerosis and favouring, as a further step, the development of cardiovascular events [7,8]. Accumulating evidence from prospective studies indicates that endothelial dysfunction is independently and significantly associated with an increased incidence of cardiovascular mortality and morbidity [7]. Because endothelial dysfunction can be reversed by appropriate non-pharmacological and pharmacological treatment [9], it is conceivable that assessment of endothelium-dependent vasodilation might contribute to the determination of the global cardiovascular risk of the individual patient. Angiotensin II as a target of the NO–sympathetic interactions When considering the emerging role of endothelial dysfunction in cardiovascular medicine, two separate studies in this issue of the journal, reporting the findings of van der Linde et al. [10,11], provide an opportunity to further address some specific mechanisms of disease activated by an altered endothelium. In the first of the two studies, Linde et al. [10] evaluated the effect of an impaired NO availability, pharmacologically obtained by the systemic administration of the specific NO synthase antagonist NG-nitro-L-arginine-methyl ester (L-NAME), on the pressor response to different agonists, such as angiotensin II, noradrenaline and phenylephrine. At baseline, all these agonists elicited comparable blood pressure increase. However, when the administration of these vasoconstrictor agents was repeated again in presence of L-NAME, although the response to noradrenaline and phenylephrine remained unchanged, the blood pressure increase elicited by angiotensin II showed a marked and significant potentiation. The potentiating effect of L-NAME on angiotensin II mainly appears to be exerted at a peripheral level because the enhanced pressor responses are haemodynamically characterized by an increase in peripheral vascular resistance, indirectly calculated as the ratio between mean arterial pressure and cardiac output. The concept that angiotensin II represents a target for L-NAME activity appears to be strengthened by the data provided by the same group of investigators in the second study [11], which was aimed at more deeply exploring the complex crosstalks between NO, the sympathetic nervous system and the renin–angiotensin–aldosterone system. In this specific set of experiments, by assessing the pressor and renal vasoconstrictor effects of L-NAME administration before and after α-blockade alone or combined with an angiotensin II receptor antagonist, the authors were able to demonstrate that: (i) the L-NAME-induced systemic and renal vasoconstriction is potentiated by α-blockade and (ii) this potentiation is almost entirely abolished when an angiotensin II receptor antagonist is concomitantly administered. These results can be regarded as a further demonstration that angiotensin II exerts a central role in the NO–sympathetic interaction by causing a potentiation of the vascular response to adrenergic and NO activation. NO–angiotensin II interactions: new findings Taken together, the data of the two studies by van der Linde et al. [10,11] provide information of key relevance in an area previously characterized by conflicting results. One of the possible reasons for the discrepant results collected to date might be related to methodological problems. Studies assessing vascular reactivity require adequate methodologies and experience in the field to produce affordable results. In the present studies [10,11], the authors made use of a rigorous methodological approach, characterized in the first case by intra-arterial measurement of blood pressure and utilization of an NO clamp and, in the second case, by the use of the Finapres approach to quantify systolic and diastolic blood pressure on a beat-to-beat basis. Needless to say that studies evaluating the acute pressor response to any kind of manoeuvre require an accurate blood pressure measurement, with intra-arterial as well as finger blood pressure recording presently representing the ‘gold standard’. In addition, the utilization of the NO clamp is of particular relevance [12]. Assessment of NO availability in humans is usually achieved by evaluating the effect of NO-synthase blockade by specific inhibitors including L-NAME or NG-monomethyl-L-arginine (L-NMMA). However, the administration of these substances (either systemically or intra-arterially) triggers a vasoconstrictive response which per se may affect vascular reactivity. Therefore, it is sometimes difficult to compare the results obtained in the same patients but with different baseline haemodynamic values (in terms of difference in local vascular tone or different blood pressure values). This problem can be overcome by the administration of sodium nitroprusside at a very low dosage, which allows baseline haemodynamic values to be restored and thus an ideal experimental setting. Moreover nitrates, which are exogenous NO donors, restore basal NO availability, allowing a genuine evaluation of the vascular effects of stimulated NO release. Apart from these important methodological considerations, the present studies also provide useful information concerning the interaction between NO and angiotensin II. The fact that, in the presence of reduced NO availability, the same amount of exogenous angiotensin II exerts an increased biological activity suggests that certain pathways may switch, from a physiological to a pathological role, depending on the integrity of the other systems. In other words, angiotensin II, which is a physiological peptide, can assume a pathological role not as a result of any change in its tissue or plasma concentrations, but because of the greater or lesser NO vascular availability. This interaction is specific, at least in these experimental conditions, because it is not seen with adrenergic agonists. As discussed by van der Linde et al. [10,11], several pathophysiological mechanisms may be responsible for the interaction between angiotensin II and NO. However, it is likely that angiotensin II exerts a negative effect on endothelial function through the release of ET-1 from vascular cells [13] and production of vasoconstrictor prostanoids, PGH2, from the endothelium [14]. Both of these effects could be partially blunted by a preserved NO availability and, conversely, potentiated when NO activity is depressed. Moreover, both in cultured vascular smooth muscle cells and in rats, exposure to angiotensin II increases oxygen free radical production via membrane-bound NADH/NADPH-driven oxidases [15,16]. Increased oxidative stress usually causes NO breakdown [17] but this effect has no role in the experimental conditions of the authors’ study, as characterized by decreased NO production because of NO synthase inhibition by L-NAME. However, it is important to underline that oxidative stress can induce a prohypertensive effect because vitamin C administration can lower blood pressure values in patients with essential hypertension [18]. Finally, AT2 receptors might play a crucial role. This possibility is in line with experimental evidence demonstrating that the stimulation of this receptor subtype can stimulate NO synthesis in endothelial cells [19] and in isolated coronary microvessels [20]. Another hypothesis raised by the authors is that impaired NO availability might determine an upregulation of AT1 receptors, therefore determining an increased sensitivity to exogenous angiotensin II. Although fascinating, this hypothesis does not appear to be supported by the short-term mechanism involved in this specific interaction between the NO pathway and angiotensin II. However, the possibility that chronic impaired NO availability could determine AT1 receptor upregulation might represent an interesting field of future research. The results obtained by Linde et al. [10,11], although suggestive of a possible mechanism of pathology, remain preliminary. This is because they were obtained in healthy individuals and, in addition, in artificial conditions in which an impairment in NO availability has been achieved via a pharmacological approach. Further caution is warranted because sympathetic activity was inferred by indirect markers of adrenergic tone, with those based on an evaluation of haemodynamic responses to reflex stimuli (e,g, the cold pressor test) being largely criticized by several investigators in recent years [21–24]. Despite these limitations, the results of the studies by the Dutch investigators open the door to a new concept with respect to the crosstalk between the endothelium, the sympathetic nervous system and the renin–angiotensin–aldosterone system. Future studies should aim to determine whether the same mechanisms contribute to high blood pressure values and/or the pathogenesis of atherosclerosis in patients with essential hypertension, who are usually characterized by impaired NO availability.