Tubuloglomerular feedback-dependent modulation of renal myogenic autoregulation by nitric oxide

Nonselective inhibition of nitric oxide (NO) synthase (NOS) augments myogenic autoregulation, an action that implies enhancement of pressure-induced constriction and dilatation. This pattern is not explained solely by interaction with a vasoconstrictor pathway. To test involvement of the Rho-Rho kinase pathway in modulation of autoregulation by NO, the selective Rho kinase inhibitor Y-27632 and/or the NOS inhibitor N(omega)-nitro-l-arginine methyl ester (l-NAME) were infused into the left renal artery of anesthetized rats. Y-27632 and l-NAME were also infused into isolated, perfused hydronephrotic kidneys to assess myogenic autoregulation over a wide range of perfusion pressure. In vivo, l-NAME reduced renal vascular conductance and augmented myogenic autoregulation, as shown by increased slope of gain reduction and associated phase peak in the pressure-flow transfer function. Y-27632 (10 mumol/l) strongly dilated the renal vasculature and profoundly inhibited autoregulation in the absence or presence of l-NAME in vivo and in vitro. Afferent arteriolar constriction induced by 30 mmol/l KCl was reversed (-92 +/- 3%) by Y-27632. Phenylephrine caused strong renal vasoconstriction but did not affect autoregulation. Inhibition of neuronal NOS by N(5)-(1-imino-3-butenyl)-l-ornithine (l-VNIO) did not cause significant vasoconstriction but did augment myogenic autoregulation. Thus vasoconstriction is neither necessary (l-VNIO) nor sufficient (phenylephrine) to explain the augmented myogenic autoregulation induced by l-NAME. The effect of l-VNIO implicates tubuloglomerular feedback (TGF) and neuronal NOS at the macula densa in regulation of the myogenic mechanism. This conclusion was confirmed by the demonstration that systemic furosemide removed the TGF signature from the pressure-flow transfer function and significantly inhibited myogenic autoregulation. In the presence of furosemide, augmentation of myogenic autoregulation by l-NAME was significantly reduced. These results provide a potential mechanism to explain interaction between myogenic and TGF-mediated autoregulation.     

Insulin sensitivity and big ET-1 conversion to ET-1 after ETA- or ETB-receptor blockade in humans.

Cardiovascular diseases are characterized by insulin resistance and elevated endothelin (ET)-1 levels. Furthermore, ET-1 induces insulin resistance. To elucidate this mechanism, six healthy subjects were studied during a hyperinsulinemic euglycemic clamp during infusion of (the ET-1 precursor) big ET-1 alone or after ET(A)- or ET(B)-receptor blockade. Insulin levels rose after big ET-1 with or without the ET(B) antagonist BQ-788 (P < 0.05) but were unchanged after the ET(A) antagonist BQ-123 + big ET-1. Infused glucose divided by insulin fell after big ET-1 with or without BQ-788 (P < 0.05). Insulin and infused glucose divided by insulin values were normalized by ET(A) blockade. Mean arterial blood pressure rose during big ET-1 with or without BQ-788 (P < 0.001) but was unchanged after BQ-123. Skeletal muscle, splanchnic, and renal blood flow responses to big ET-1 were abolished by BQ-123. ET-1 levels rose after big ET-1 (P < 0.01) in a similar way after BQ-123 or BQ-788, despite higher elimination capacity after ET(A) blockade. In conclusion, ET-1-induced reduction in insulin sensitivity and clearance as well as splanchnic and renal vasoconstriction are ET(A) mediated. ET(A)-receptor stimulation seems to inhibit the conversion of big ET-1 to ET-1.     

Altered role of smooth muscle endothelin receptors in coronary endothelin-1 and alpha1-adrenoceptor-mediated vasoconstriction in Type 2 diabetes

Regulation of vascular tone and blood flow involves interactions between numerous local and systemic vascular control signals, many of which are altered by Type 2 diabetes (T2D). Vascular responses to endothelin-1 (ET-1) are mediated by endothelin type A (ET(A)) and type B (ET(B)) receptors that have been implicated in cross talk with alpha(1)-adrenoceptors (alpha(1)-AR). ET(A) and ET(B) receptor expression and plasma ET-1 levels are elevated in T2D; however, whether this influences coronary alpha(1)-AR function has not been examined. Therefore, we examined the effect of ET(A) and ET(B) receptor inhibition on coronary vasoconstriction to ET-1 and alpha(1)-AR activation in a mouse model of T2D. Coronary vascular responses were examined in isolated mouse hearts from control and diet-induced T2D C57BL/6J mice. Responses to ET-1 and the selective alpha(1)-AR agonist phenylephrine (PE) were examined alone and in the presence of the nitric oxide synthase inhibitor N(omega)-nitro-l-arginine methyl ester (l-NAME) alone or in combination with selective ET(A) or ET(B) receptor inhibitors BQ-123 and BQ-788, respectively. Vasoconstriction to ET-1 was enhanced, whereas ET(B), but not ET(A), receptor blockade reduced basal coronary tone in T2D hearts. In the presence of l-NAME, ET(A) receptor inhibition attenuated ET-1 vasoconstriction in both groups, whereas ET(B) inhibition abolished this response only in control hearts. In addition, ET(A) inhibition enhanced alpha(1)-AR-mediated vasoconstriction in T2D, but not control, hearts following l-NAME treatment. Therefore, in this model, enhanced coronary ET-1 responsiveness is mediated primarily through smooth muscle ET(B) receptors, whereas the interaction with alpha(1)-ARs is mediated solely through the ET(A) receptor subtype.     

Determinants of terminal mesenteric artery resistance during the first postnatal month

Experiments were conducted to delineate the vascular effector systems that contribute to setting mesenteric vascular tone in swine during the first postnatal month. Terminal mesenteric arteries (TMA), which function as resistance vessels, were studied in vitro with a microvascular perfusion system allowing independent pressure and flow manipulation. When pressure was varied 0-100 mmHg in the absence of flow, TMA from 1-day-old animals demonstrated myogenic vasoconstriction, whereas TMA from 40-day-old animals did not. In 1- but not 40-day-old TMA, the endothelin A (ET(A)) receptor antagonist BQ-610 shifted the pressure-diameter curve upward, whereas the ET(B) receptor antagonist BQ-788 and the L-arginine analog N(G)-monomethyl-L-arginine (L-NMMA) shifted the curve downward; in all instances, myogenic vasoconstriction was preserved. Flow eliminated myogenic vasoconstriction in 1-day-old TMA, i.e., diameter increased as a function of pressure. The effect of BQ-610 was lost under flow conditions; however, BQ-788 and N-acyl-L-Trp-3,5-bis-(trifluoromethyl) benzyl ester, an antagonist specific to the substance P neurokinin-1 (NK(1)) receptor, shifted the pressure-diameter curve downward in the presence of flow, whereas L-NMMA restored myogenic vasoconstriction. Adding flow had no effect on the pressure-diameter relationship in 40-day-old TMA. Other blocking agents, including prazosin, losartan, indomethacin, and charybdotoxin, had no effect on the pressure-diameter relationship in either age group under flow or no-flow conditions. Constitutive production of nitric oxide (NO) and endothelin-1 participates in setting resistance in 1-day-old TMA, and important stimulants to NO production include flow and activation of ET(B) and NK(1) receptors. In contrast, 40-day-old TMA act as passive conduits in which the elastic properties of the vessel are the primary determinant of diameter.     

Impaired endothelin-induced vasoconstriction in coronary arteries of Zucker obese rats is associated with uncoupling of [Ca2+]i signaling

Although insulin resistance (IR) is a major risk factor for coronary artery disease, little is known about the regulation of coronary vascular tone in IR by endothelin-1 (ET-1). We examined ET-1 and PGF(2alpha)-induced vasoconstriction in isolated small coronary arteries (SCAs; approximately 250 microM) of Zucker obese (ZO) rats and control Zucker lean (ZL) rats. ET-1 response was assessed in the absence and presence of endothelin type A (ET(A); BQ-123), type B (ET(B); BQ-788), or both receptor inhibitors. ZO arteries displayed reduced contraction to ET-1 compared with ZL arteries. In contrast, PGF(2alpha) elicited similar vasoconstriction in both groups. ET(A) inhibition diminished the ET-1 response in both groups. ET(B) inhibition alone or in combination with ET(A) blockade, however, restored the ET-1 response in ZO arteries to the level of ZL arteries. Similarly, inhibition of endothelial nitric oxide (NO) synthase with N(omega)-nitro-l-arginine methyl ester (l-NAME) enhanced the contraction to ET-1 and abolished the difference between ZO and ZL arteries. In vascular smooth muscle cells from ZO, ET-1-induced elevation of myoplasmic intracellular free calcium concentration ([Ca2+]i) (measured by fluo-4 AM fluorescence), and maximal contractions were diminished compared with ZL, both in the presence and absence of l-NAME. However, increases in [Ca2+]i elicited similar contractions of the vascular smooth muscle cells in both groups. Analysis of protein and total RNA from SCA of ZO and ZL revealed equal expression of ET-1 and the ET(A) and ET(B) receptors. Thus coronary arteries from ZO rats exhibit reduced ET-1-induced vasoconstriction resulting from increased ET(B)-mediated generation of NO and diminished elevation of myoplasmic [Ca2+]i.     

Mechanism of hypoxic pulmonary vasoconstriction involves ET(A) receptor-mediated inhibition of K(ATP) channel

There is controversy on the role of endothelin (ET)-1 in the mechanism of hypoxic pulmonary vasoconstriction (HPV). Although HPV is inhibited by ET-1 subtype A (ET(A))-receptor antagonists in animals, it has been reported that ET(A)-receptor blockade does not affect HPV in isolated lungs. Thus we reassessed the role of ET-1 in HPV in both rats and isolated blood- and physiological salt solution (PSS)-perfused rat lungs. In rats, the ET(A)-receptor antagonist BQ-123 and the nonselective ET(A)- and ET(B)-receptor antagonist PD-145065, but not the ET(B)-receptor antagonist BQ-788, inhibited HPV. Similarly, BQ-123, but not BQ-788, attenuated HPV in blood-perfused lungs. In PSS-perfused lungs, either BQ-123, BQ-788, or the combination of both attenuated HPV equally. Inhibition of HPV by combined BQ-123 and BQ-788 in PSS-perfused lungs was prevented by costimulation with angiotensin II. The ATP-sensitive K(+) (K(ATP))-channel blocker glibenclamide also prevented inhibition of HPV by BQ-123 in both lungs and rats. These results suggest that ET-1 contributes to HPV in both isolated lungs and intact animals through ET(A) receptor-mediated suppression of K(ATP)-channel activity.     

Hyperinsulinemia fails to augment ET-1 action in the skeletal muscle vascular bed in vivo in humans

Endogenous endothelin action is augmented in human obesity and type 2 diabetes and contributes to endothelial dysfunction and impairs insulin-mediated vasodilation in humans. We hypothesized that insulin resistance-associated hyperinsulinemia could preferentially drive endothelin-mediated vasoconstriction. We applied hyperinsulinemic-euglycemic clamps with higher insulin dosing in obese subjects than lean subjects (30 vs. 10 mU.m(-2).min(-1), respectively), with the goal of matching insulin's nitric oxide (NO)-mediated vascular effects. We predicted that, under these circumstances, insulin-stimulated endothelin-1 (ET-1) action (assessed with the type A endothelin receptor antagonist BQ-123) would be augmented in proportion to hyperinsulinemia. NO bioactivity was assessed using the nitric oxide synthase inhibitor N(G)-monomethyl-l-arginine. Insulin-mediated vasodilation and insulin-stimulated NO bioavailability were well matched across groups by this approach. As expected, steady-state insulin levels were approximately threefold higher in obese than lean subjects (109.2 +/- 10.2 pmol/l vs. 518.4 +/- 84.0, P = 0.03). Despite this, the augmentation of insulin-mediated vasodilation by BQ-123 was not different between groups. ET-1 flux across the leg was not augmented by insulin alone but was increased with the addition of BQ-123 to insulin (P = 0.01 BQ-123 effect, P = not significant comparing groups). Endothelin antagonism augmented insulin-stimulated NO bioavailability and NOx flux, but not differently between groups and not proportional to hyperinsulinemia. These findings do not support the hypothesis that insulin resistance-associated hyperinsulinemia preferentially drives endothelin-mediated vasoconstriction.     

Responses to neither exogenous nor endogenous endothelin-1 are altered in patients with hypercholesterolemia

There is some controversy regarding whether vascular responses to endothelin are altered in hypercholesterolemia. Studies performed to date have been compromised by the use of endothelin antagonists at inappropriate concentrations. In the current study, we examine the role of endothelin-1 in hypercholesterolemic patients using lower, more selective doses of specific endothelin antagonists. Twenty-two patients with hypercholesterolemia (total plasma cholesterol > 6.0 mmol/l) and 17 healthy controls were recruited. Forearm vascular responses to endothelin-1 (5 pmol/min), the endothelin A antagonist BQ-123 (10 nmol/min), and the endothelin B antagonist BQ-788 (1 nmol/min) were obtained. Endothelin-1 caused a significant vasoconstriction in both hypercholesterolemic and control subjects, an effect that was not significantly different between the two groups (P = 0.784). BQ-123 caused a significant vasodilatation that was not significantly different between the two groups (P = 0.899). Similarly, responses to BQ-788 (P = 0.774) and mean plasma endothelin-1 levels were not different (control vs. hypercholesterolemia, 1.16 +/- 0.18 vs. 1.06 +/- 0.15 fmol/ml; P = 0.64). Responses to neither exogenous nor endogenous endothelin are influenced by plasma cholesterol levels in humans. It is thus unlikely that the endothelin system contributes to early vascular disease pathology in patients with hypercholesterolemia.     

The renal and systemic hemodynamic effects of a nitric oxide-synthase inhibitor are reversed by a selective endothelin(a) receptor antagonist in men.

There is evidence for an interaction between nitric oxide (NO) and endothelin (ET) at the level of the renal vasculature. We hypothesized that acute renal effects of systemic NO synthase inhibition (NG-monomethyl-l-arginine, L-NMMA) may be blunted by coadministration of a specific ET(A) receptor antagonist (BQ-123) in healthy humans. Fifteen healthy young male subjects participated in this randomized, double-blind, placebo-controlled 3-way crossover study. These sodium-repleted volunteers received L-NMMA alone, or BQ-123 alone, or L-NMMA with a subsequent coinfusion of BQ-123. Renal plasma flow (RPF) and glomerular filtration rate (GFR) were determined with the PAH and inulin clearance method, respectively. Mean arterial pressure (MAP) and pulse rate were measured noninvasively at baseline and every 15 min after the start of the study period. L-NMMA alone reduced RPF (-22%, P < 0.001) and GFR (-8%, P < 0.009) and increased MAP (+10%, P < 0.001). BQ-123 alone did not affect these parameters. However, coinfusion of BQ-123 blunted the effects of L-NMMA on RPF (P < 0.001), GFR (P < 0.001), and MAP (P = 0.006). Peripheral and renal hemodynamic effects of acute systemic NO synthase inhibition are at least partially reversed by ET(A) receptor blockade with BQ-123. This indicates a functional antagonism between specific ET(A) receptor antagonist and NO synthase inhibitors at the level of the renal vasculature.     

Paradoxical coronary microcirculatory constriction during ischemia: a synergic function for nitric oxide and endothelin

A paradoxical microcirculatory constriction has been observed in hearts of patients with ischemia, secondary to coronary stenosis. Here, using the isolated mouse heart (Langendorff), we examined the mechanism of this response, assuming involvement of nitric oxide (NO) and endothelin-1 (ET-1) systems. Perfusion pressure was maintained at 65 mmHg for 70 min (protocol 1), or it was reduced to 30 mmHg over two intervals, between the 20- and 40-min marks (protocol 2) or from the 20-min mark onward (protocol 3). In protocol 1, coronary resistance (CR) remained steady in untreated heart, whereas it progressively increased during treatment with the NO synthesis inhibitor N(G)-nitro-l-arginine methyl ester (L-NAME) (2.7-fold) or the ET(A) antagonist BQ-610 (2.8 fold). The ET(B) antagonist BQ-788 had instead no effect by itself but curtailed vasoconstriction to BQ-610. In protocol 2, hypotension raised CR by 2.2-fold. This response was blunted by reactive oxygen species (ROS) scavengers (mannitol and superoxide dismutase plus catalase) and was converted into vasodilation by l-NAME, BQ-610, or BQ-788. Restoration of normal pressure was followed by vasodilation and vasoconstriction, respectively, in untreated and treated preparations. In protocol 3, CR progressively increased with hypotension in the absence but not presence of L-NAME or BQ-610. We conclude that the coronary vasculature is normally relaxed by two concerted processes, a direct action of NO and ET-1 curtailing an ET(B2)-mediated tonic vasoconstriction through ET(A) activation. The negative feedback mechanism on ET(B2) subsides during hypotension, and the ensuing vasoconstriction is ascribed to ET-1 activating ET(A) and ET(B2) and reactive nitrogen oxide species originating from ROS-NO interaction.     

The blockade of endothelin A receptor protects astrocytes against hypoxic injury: common effects of BQ-123 anderythropoietin on the rejuvenation of the astrocyte population

In the present study the role of endothelin (ET) and its receptors (ETA-R and ETB-R) in cellular mechanisms underlying the resistance of astroglial cells to low oxygen level and development of hypoxia has been investigated. To define the influences of ET and its receptors on survival and on antigenic as well as morphologic differentiation of rat astroglial cells in normoxic (NC) and hypoxic culture (HC) the selective antagonists of ETA-R (BQ-123) and ETB-R (BQ-788) were used. Treatment of HC with BQ-123 caused an increase in cell number and inhibited the hypoxia-induced apoptosis by 37%. BQ-123 decreased the hypoxia-induced cytotoxicity in HC. These effects of BQ-123 were abolished in cultures simultaneously treated with BQ-123 and BQ-788. Administration of BQ-788 alone decreased the number of living cells in NC, but not in HC. The activity of caspase-3/-7 was not changed by exposure of NC and HC to BQ-788. The protection provided by BQ-123 to astroglial cells against cytotoxicity in NC and HC was similar to that of erythropoietin (EPO), a cytokine with established neuroprotective effects. The functional improvement of astroglial cells and slowing down of their differentiation under exposure to BQ-123, or EPO, or BQ-123 + EPO has been evidenced by an increased number of nestin+/glial fibrillary acidic protein-positive (GFAP+) astrocytes accompanied by decrease of nestin-/GFAP+ cells. The simultaneous treatment with BQ-123 and EPO additionally decreased the activities of caspase-3/-7 (64%) and release of LDH into the medium (94%). The benefits in the functional states of astrocytes obtained by combined treatment of HC with BQ-123 and EPO suggest a new therapeutic strategy in treatment of hypoxic brain injury.     

NO and NO-independent mechanisms mediate ETB receptor buffering of ET-1-induced renal vasoconstriction in the rat

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.     

Interaction between nitric oxide and renal myogenic autoregulation in normotensive and hypertensive rats

Blood pressure fluctuates continuously throughout life and autoregulation is the primary mechanism that isolates the kidney from this fluctuation. Compared with Wistar rats, Brown Norway (B-N) rats display impaired renal myogenic autoregulation when blood pressure fluctuation is increased. They also are very susceptible to hypertension-induced renal injury. Because blockade of nitric oxide augments myogenic autoregulation in Wistar rats, we compared the response of the myogenic system in B-N rats to nitric oxide blockade with that of other strains [Wistar, Sprague-Dawley, Long-Evans, spontaneously hypertensive (SHR)]. Renal blood flow dynamics were assessed in isoflurane anesthetized rats before and after inhibition of nitric oxide synthase by Lomega-nitro-arginine methyl-ester (L-NAME, 10 mg/kg, iv). Under control conditions, myogenic autoregulation in the B-N rats was weaker than in the other strains. Myogenic autoregulation was not augmented after L-NAME administration in the SHR, but was augmented in all the normotensive rats. The enhancement was significantly greater in B-N rats so that after L-NAME the efficiency of autoregulation did not differ among the strains. The data suggest that nitric oxide is involved in the impaired myogenic autoregulation seen in B-N rats. Furthermore, the similarity of response in Wistar, Long-Evans, and Sprague-Dawley rats suggests that modulation by nitric oxide is a fundamental property of renal myogenic autoregulation.     

Nitric oxide and endothelin in oxygen-dependent regulation of vascular tone of human umbilical vein

We investigated the possible contribution of nitric oxide (NO) and endothelin (ET) to oxygen-dependent regulation of human umbilical vein vascular tone by simultaneous registration of intracellular membrane potential and isometric tension of vessel strips with and without NO synthase inhibition [10-4 M N omega-nitro-L-arginine methyl ester (L-NAME)], ETA receptor blockade (10(-5) M BQ-123), or ETB receptor blockade (10(-7) M BQ-788) at Po2 values in the bath solution between 5 and 104 mmHg. Increasing PO2 above the physiological intrauterine range resulted in depolarization and an increase of isometric tension, whereas lowering PO2 resulted in hyperpolarization and a decrease in isometric tension. Removal of the endothelium reversed these effects. At PO2 values below 39 mmHg, intact preparations treated with either L-NAME, BQ-788, or BQ-123 were more depolarized than controls. In the case of treatment with L-NAME or BQ-123, this was accompanied by an increase in isometric tension. We conclude that it is NO that mediates the hypoxic hyperpolarization and vasodilatation of the human umbilical vein and that ET, via activation of ETB1 receptors on endothelial cells, contributes to this effect.     

Endothelin type A receptor antagonist normalizes blood pressure in rats exposed to eucapnic intermittent hypoxia

We have reported that eucapnic intermittent hypoxia (E-IH) causes systemic hypertension, elevates plasma endothelin 1 (ET-1) levels, and augments vascular reactivity to ET-1 and that a nonspecific ET-1 receptor antagonist acutely lowers blood pressure in E-IH-exposed rats. However, the effect of chronic ET-1 receptor inhibition has not been evaluated, and the ET receptor subtype mediating the vascular effects has not been established. We hypothesized that E-IH causes systemic hypertension through the increased ET-1 activation of vascular ET type A (ET(A)) receptors. We found that mean arterial pressure (MAP) increased after 14 days of 7 h/day E-IH exposure (109 +/- 2 to 137 +/- 4 mmHg; P < 0.005) but did not change in sham-exposed rats. The ET(A) receptor antagonist BQ-123 (10 to 1,000 nmol/kg iv) acutely decreased MAP dose dependently in conscious E-IH but not sham rats, and continuous infusion of BQ-123 (100 nmol.kg(-1).day(-1) sc for 14 days) prevented E-IH-induced increases in MAP. ET-1-induced constriction was augmented in small mesenteric arteries from rats exposed 14 days to E-IH compared with those from sham rats. Constriction was blocked by the ET(A) receptor antagonist BQ-123 (10 microM) but not by the ET type B (ET(B)) receptor antagonist BQ-788 (100 microM). ET(A) receptor mRNA content was greater in renal medulla and coronary arteries from E-IH rats. ET(B) receptor mRNA was not different in any tissues examined, whereas ET-1 mRNA was increased in the heart and in the renal medulla. Thus augmented ET-1-dependent vasoconstriction via vascular ET(A) receptors appears to elevate blood pressure in E-IH-exposed rats.     

Effect of BQ-123 and nitric oxide inhibition on liver in rats after renal ischemia-reperfusion injury

Ischemia-reperfusion (I/R) injury induces an inflammatory response and production of oxygen-derived reactive species which affect many organs including heart, brain, kidney and gastrointestinal tract. The aim of this study was to assess the hepatic changes after renal I/R injury. Male Sprague Dawley rats were subjected to either sham operation or treatment with L-NAME, L-arginine and BQ-123 during 30 min renal ischemia and 2 h reperfusion injury. Hepatic superoxide dismutase (SOD), catalase, glutathione peroxidase (GSH-Px) activities, and thiobarbituric acid-reactive substances (TBARS) and nitric oxide (NO) levels were evaluated to show hepatic response to renal I/R injury. Catalase and SOD activities showed significant differences between the control and the other groups after I/R. On the other hand, GSH-Px activity did not show any significant changes between the control and the other experimental groups mentioned under above conditions. Meanwhile, levels of TBARS were not different between the control and the other experimental groups, whereas NO level showed changes between the control and experimental groups except the one to which endothelin receptor antagonist agent (BQ-123) subjected. Experimental period may not be enough to determine the changes in GSH-Px activity and level of TBARS. However, catalase and SOD activities decreased in experimental groups treated by chemical agents. NO level decreased in chemicalagent-applied experimental groups but not in the group to which endothelin receptor antagonist BQ-123 was applied alone.     

ET-1-induced pulmonary vasoconstriction shifts from ET(A)- to ET(B)-receptor-mediated reaction after preconstriction.

Endothelin-1 (ET-1) has been reported to induce pulmonary vasoconstriction via either ET(A) or ET(B) receptors, and vasorelaxation after ET-1 injection has been observed. Our study investigated the effects of ET-1 in isolated rabbit lungs, which were studied at basal tone (part I) and after preconstriction (U-46619; part II). Pulmonary arterial pressure (PAP) and lung weight gain were monitored continuously. In part I, ET-1 (10(-8) M; n = 6; control) was injected after pretreatment with the ET(A)-receptor antagonist BQ-123 (10(-6) M; n = 6) or the ET(B)-receptor antagonist BQ-788 (10(-6) M; n = 6). The same protocol was carried out in part II after elevation of pulmonary vascular tone. ET-1 induced an immediate PAP increase (DeltaPAP 4.3 +/- 0.4 mmHg at 10 min) that was attenuated by pretreatment with BQ-123 (P < 0.05 at 10 min and P < 0.01 thereafter) and that was more pronounced after BQ-788 (P < 0.01 at 10 min and P < 0.001 thereafter). In part II, ET-1 induced an immediate rise in PAP with a maximum after 5 min (DeltaPAP 6.3 +/- 1.4 mmHg), leveling off at DeltaPAP 3.2 +/- 0.2 mmHg after 15 min. Pretreatment with BQ-123 failed to attenuate the increase. BQ-788 significantly reduced the peak pressure at 5 min (0.75 +/- 0.4 mmHg; P < 0.001) as well as the plateau pressure thereafter (P < 0.01). We conclude that ET-1 administration causes pulmonary vasoconstriction independent of basal vascular tone, and, at normal vascular tone, the vasoconstriction seems to be mediated via ET(A) receptors. BQ-788 treatment resulted in even more pronounced vasoconstriction. After pulmonary preconstriction, ET(A) antagonism exerted no effects on PAP, whereas ET(B) antagonism blocked the PAP increase. Therefore, ET-1-induced pulmonary vasoconstriction is shifted from an ET(A)-related to an ET(B)-mediated mechanism after pulmonary vascular preconstriction.     

Endothelin-mediated vasoconstriction in postischemic newborn intestine

We previously suggested that the profound, sustained vasoconstriction noted in 3-day-old swine intestine after a moderate episode of ischemia-reperfusion (I/R) reflects the unmasking of underlying constrictor tone consequent to a loss of endothelium-derived nitric oxide (NO). In this study, we sought to determine whether endothelin-1 (ET-1) was the unmasked constrictor and whether selective loss of endothelial ET(B) receptors, which mediate NO-based vasodilation, participated in the hemodynamic consequences of I/R in newborn intestine. Studies were performed in innervated, autoperfused intestinal loops in 3- and 35-day-old swine. Selective blockade of ET(A) receptors with BQ-610 had no effect on hemodynamics under control conditions; however, when administered before and during I/R, BQ-610 significantly attenuated the post-I/R vasoconstriction and reduction in arteriovenous O(2) difference in the younger group. In 3-day-old intestine, reduction of intestinal O(2) uptake to a level similar to that noted after I/R by lowering tissue temperature had no effect on the response to BQ-610 or ET-1, indicating that the change in response to BQ-610 noted after I/R was not simply consequent to the reduction in tissue O(2) demand. In studies in mesenteric artery rings suspended in myographs, we observed a leftward shift in the dose-response curve for ET-1 after selective blockade of ET(B) receptors with BQ-788 in 3- but not 35-day-old swine. Rings exposed to I/R in vivo behaved in a manner similar to control rings treated with BQ-788 or endothelium-denuded non-I/R rings.     

Contribution of endothelin to pulmonary vascular tone under normoxic and hypoxic conditions

The contribution of endothelin to resting pulmonary vascular tone and hypoxic pulmonary vasoconstriction in humans is unknown. We studied the hemodynamic effects of BQ-123, an endothelin type A receptor antagonist, on healthy volunteers exposed to normoxia and hypoxia. Hemodynamics were measured at room air and after 15 min of exposure to hypoxia (arterial PO(2) 99.8 +/- 1.8 and 49.4 +/- 0.4 mmHg, respectively). Measurements were then repeated in the presence of BQ-123. BQ-123 decreased pulmonary vascular resistance (PVR) 26% and systemic vascular resistance (SVR) 21%, whereas it increased cardiac output (CO) 22% (all P < 0.05). Hypoxia raised CO 28% and PVR 95%, whereas it reduced SVR 23% (all P < 0.01). During BQ-123 infusion, hypoxia increased CO 29% and PVR 97% and decreased SVR 22% (all P < 0.01). The pulmonary vasoconstrictive response to hypoxia was similar in the absence and presence of BQ-123 [P = not significant (NS)]. In vehicle-treated control subjects, hypoxic pulmonary vasoconstriction did not change with repeated exposure to hypoxia (P = NS). Endothelin contributes to basal pulmonary and systemic vascular tone during normoxia, but does not mediate the additional pulmonary vasoconstriction induced by acute hypoxia.     

ET-1 induces cortical spreading depression via activation of the ETA receptor/phospholipase C pathway in vivo

Recently, it has been shown that brain topical superfusion of endothelin (ET)-1 at concentrations around 100 nM induces repetitive cortical spreading depressions (CSDs) in vivo. It has remained unclear whether this effect of ET-1 is related to a primary neuronal/astroglial effect, such as an increase in neuronal excitability or induction of interastroglial calcium waves, or a penumbra-like condition after vasoconstriction. In vitro, ET-1 regulates interastroglial communication via combined activation of ET(A) and ET(B) receptors, whereas it induces vasoconstriction via single activation of ET(A) receptors. We have determined the ET receptor profile and intracellular signaling pathway of ET-1-induced CSDs in vivo. In contrast to the ET(B) receptor antagonist BQ-788 and concentration dependently, the ET(A) receptor antagonist BQ-123 completely blocked the occurrence of ET-1-induced CSDs. The ET(B) receptor antagonist did not increase the efficacy of the ET(A) receptor antagonist. Direct stimulation of ET(B) receptors with the selective ET(B) agonist BQ-3020 did not trigger CSDs. The phospholipase C (PLC) antagonist U-73122 inhibited CSD occurrence in contrast to the protein kinase C inhibitor G?-6983. Our findings indicate that ET-1 induces CSDs through ET(A) receptor and PLC activation. We conclude that the induction of interastroglial calcium waves is unlikely the primary cause of ET-1-induced CSDs. On the basis of the receptor profile, likely primary targets of ET-1 mediating CSD are either neurons or vascular smooth muscle cells.