The vascular actions of endothelin-1 (ET-1) reflect the combination of vasoconstrictor ETA and ETB receptors on smooth muscle cells and vasodilator ETB receptors on endothelial cells. The present study investigated the contribution of ET receptor subtypes using a comprehensive battery of agonists and antagonists infused directly into the renal artery of anesthetized rats to evaluate the actions of each receptor class alone and their interactions. ET-1 (5 pmol) reduced renal blood flow (RBF) 25 Ϯ 1%. ETA antagonist BQ-123 attenuated this response to a 15 Ϯ 1% decrease in RBF (P Ͻ 0.01), indicating net constriction by ETB receptors. Combined receptor blockade (BQ-123ϩBQ-788) resulted in a renal vasoconstriction of 7 Ϯ 1% (P ϭ 0.001 vs. BQ-123), supporting a constrictor action of ET B receptors. In marked contrast, the ETB antagonist BQ-788 enhanced the ET-1 RBF response to 60 Ϯ 5% (P Ͻ 0.001), suggesting ET B-mediated net dilation. Consistent with ET A blockade, the ETB agonist sarafotoxin 6C (S6C) produced vasoconstriction, reducing RBF by 23 Ϯ 5%. Dose-response curves for ET-1 and S6C showed similar degrees of constriction between 0.2 and 100 pmol. Both antagonists (BQ-123, BQ-788) were equally effective at threefold lower than the standard doses, suggesting complete inhibition. We conclude that ET B receptors alone exert a net constrictor effect but cause a net dilator influence when costimulated with ET A receptors. Such opposing actions indicate more complex than additive interaction between receptor subtypes. Model analysis suggests ET A-mediated constriction is appreciably greater without than with costimulation of ET B receptors. Possible explanations include ET-1 clearance by ET B receptors and/or a dilator ETB receptor function that counteracts constriction. vascular smooth muscle cells; endothelial cells; renal vascular resistance; nitric oxide ENDOTHELIN-1 (ET-1) PLAYS an important role in the regulation of renal hemodynamics and salt and water excretion (38). Under physiological conditions the vascular endothelium constitutively produces ET-1. Although endogenous ET-1 normally has a minor influence on renal hemodynamics (37), it gains more importance in disease states such as hypertension, acute and chronic renal failure, and congestive heart failure (38). ET-1 is capable of constricting or dilating the renal vasculature depending on the relative distribution and influence of ET A and ET B receptors. In the vascular wall, ET A receptors are limited to smooth muscle cells, whereas ET B receptors are expressed in both endothelial and smooth muscle cells (38). Both receptor types are coupled to Gq proteins, thereby activating several signaling pathways including phospholipase C and intracellular calcium concentration (38), which leads to constriction of smooth muscle cells. Activation of endothelial ET B receptors by the same signaling pathways produces vasodilation mediated by nitric oxide and possibly eicosanoids (14).There is general agreement that administration of ET-1 produces renal vasoconstrictio...

NAD(P)H oxidases (NOX) and reactive oxygen species (ROS) are involved in vasoconstriction and vascular remodeling during hypertension produced by chronic angiotensin II (ANG II) infusion. These effects are thought to be mediated largely through superoxide anion (O(2)(-)) scavenging of nitric oxide (NO). Little is known about the role of ROS in acute vasoconstrictor responses to agonists. We investigated renal blood flow (RBF) reactivity to ANG II (4 ng), norepinephrine (NE, 20 ng), and alpha(1)-adrenergic agonist phenylephrine (PE, 200 ng) injected into the renal artery (ira) of anesthetized Sprague-Dawley rats. The NOX inhibitor apocynin (1-4 mg.kg(-1).min(-1) ira, 2 min) or the superoxide dismutase mimetic Tempol (1.5-5 mg.kg(-1).min(-1) ira, 2 min) rapidly increased resting RBF by 8 +/- 1% (P < 0.001) or 3 +/- 1% (P < 0.05), respectively. During NO synthase (NOS) inhibition (N(omega)-nitro-l-arginine methyl ester, 25 mg/kg iv), the vasodilation tended to increase (apocynin 13 +/- 4%, Tempol 10 +/- 1%). During control conditions, both ANG II and NE reduced RBF by 24 +/- 4%. Apocynin dose dependently reduced the constriction by up to 44% (P < 0.05). Similarly, Tempol blocked the acute actions of ANG II and NE by up to 48-49% (P < 0.05). In other animals, apocynin (4 mg.kg(-1).min(-1) ira) attenuated vasoconstriction to ANG II, NE, and PE by 46-62% (P < 0.01). During NOS inhibition, apocynin reduced the reactivity to ANG II and NE by 60-72% (P < 0.01), and Tempol reduced it by 58-66% (P < 0.001). We conclude that NOX-derived ROS substantially contribute to basal RBF as well as to signaling of acute renal vasoconstrictor responses to ANG II, NE, and PE in normal rats. These effects are due to O(2)(-) rather than H(2)O(2), occur rapidly, and are independent of scavenging of NO.

Vascular endothelin (ET) type B (ET(B)) receptors exert dilator and constrictor actions in a complex interaction with ET(A) receptors. We aimed to clarify the presence and relative importance of nitric oxide (NO) and other mechanisms underlying the dilator effects of ET(B) receptors in rat kidneys. Complete inhibition of NO production with Nomega-nitro-L-arginine methyl ester (L-NAME, 25 mg/kg iv) enhanced the renal vasoconstriction elicited by ET-1 injected into the renal artery from -15 to -30%. Additional infusion of the NO donor nitroprusside (NP) into the renal artery did not reverse this effect (-29%) but effectively buffered ANG II-mediated vasoconstriction. Similarly, ET-1 responses were enhanced after a smaller intrarenal dose of L-NAME (-22 vs. -15%) and were unaffected by subsequent NP infusion (-21%). These results indicate that the responsiveness to ET-1 is buffered by ET(B) receptor-stimulated phasic release of NO, rather than its static mean level. Infusion of the ET(B) receptor antagonist BQ-788 into the renal artery further enhanced the ET-1 constrictor response to NP+L-NAME (-92 vs. -49%), revealing an NO-independent dilator component. In controls, vasoconstriction to ET-1 was unaffected by vehicle (-27 vs. -20%) and markedly enhanced by BQ-788 (-70%). The same pattern was observed when indomethacin (Indo) was used to inhibit cyclooxygenase (-20% for control, -22% with Indo, and -56% with ET(B) antagonist) or methylsulfonyl-6-(2-propargyloxyphenyl)-hexanamide (MS-PPOH) or miconazole+Indo was used to inhibit epoxygenase alone (-10% for control, -11% with MS-PPOH, and -35% with ET(B) antagonist) or in combination (-14% for control, -20% with Indo + miconazole, and -43% with ET(B) antagonist). We conclude that phasic release of NO, but not its static level, mediates part of the dilator effect of ET(B) receptors and that an NO-independent mechanism, distinct from prostanoids and epoxyeicosatetraenoic acids, perhaps ET(B) receptor clearance of ET-1, plays a major buffering role.

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