Different populations of progesterone receptor-steroid complexes in binding to specific DNA sequences: effects of salts on kinetics and specificity

The endothelium plays an obligatory role in a number of relaxations of isolated arteries. These endothelium-dependent relaxations are due to the release by the endothelial cells of potent vasodilator substances [endothelium-derived relaxing factors (EDRF)]. The best characterized EDRF is nitric oxide (NO). Nitric oxide is formed by the metabolism of L-arginine by the constitutive NO synthase of endothelial cells. In arterial smooth muscle, the relaxations evoked by EDRF are explained best by the stimulation by NO of soluble guanylate cyclase that leads to the accumulation of cyclic GMP. The endothelial cells also release an unidentified substance that causes hyperpolarization of the cell membrane (endothelium-derived hyperpolarizing factor, EDHF). The release of EDRF from the endothelium can be mediated by both pertussis toxin-sensitive (alpha2-adrenergic activation, serotonin, thrombin, aggregating platelets) and insensitive (adenosine diphosphate, bradykinin) G-proteins. In blood vessels from animals with regenerated endothelium, and/or atherosclerosis, there is a selective loss of the pertussis-toxin sensitive mechanism of EDRF-release which favors the occurrence of vasospasm, thrombosis and cellular growth.     

Brain activation, affect, and aerobic exercise: an examination of both state-independent and state-dependent relationships

N-2-hydroxyethylpiperazine-N'-2-ethanesulphonic acid (HEPES) is a commonly-used buffer. This study determined the effect of HEPES on contractility of the dog basilar artery and tested the hypothesis that HEPES inhibits vasoconstriction of isolated arterial segments by generating H2O2. Rings of dog basilar artery with or without endothelium were suspended under isometric tension and contracted with KCl, serotonin, or prostaglandin F2 alpha (PGF2 alpha) in bicarbonate or HEPES buffer. Addition of HEPES, 30 mmol l-1, before or after contraction with KCl, serotonin or PGF2 alpha significantly decreased maximal tension in rings with or without endothelium. Preincubation with HEPES buffer, 10 mmol l-1, significantly decreased maximal contractions to each agonist in rings with endothelium and to KCl and serotonin in rings without endothelium. HEPES, 30 mmol l-1, noncompetitively inhibited concentration-contraction curves to increasing concentrations of each agonist in rings with or without endothelium. Inhibition by HEPES was completely reversible with washing. The inhibitory effects of HEPES on responses to each agonist in rings with endothelium were significantly less in the dark or after coincubation with catalase. Unlike HEPES, effects of H2O2 were endothelium-dependent in that H2O2 caused contractions in rings with endothelium and relaxations in rings without endothelium. 5-(N,N'-dimethyl)-amiloride and 4,4'-diisothiocyanataostilbene-2,2'-disulfonic acid did not affect contractility in this preparation. These results show that HEPES exerts significant inhibitory effects on arterial smooth muscle contractility. The mechanism does not involve endothelium-dependent relaxation, effects on chloride channels or the sodium-hydrogen exchanger or generation of H2O2 by HEPES in the light.     

Contraction and endothelium dependent relaxation via alpha adrenoceptors are variable in various pig arteries

OBJECTIVE: The aim was to examine differences in the vascular response to alpha adrenoceptor stimulation on both smooth muscle and endothelium among large arteries perfusing various organs. METHODS: In ring preparations of coronary, carotid, iliac, mesenteric, and renal arteries from pigs (n = 24), the magnitudes of contraction to noradrenaline (with beta blocker), with or without endothelium, and endothelium dependent relaxation to noradrenaline (with alpha 1 and beta blocker) were compared in vitro. RESULTS: In both endothelium-intact and denuded groups, the contractions to noradrenaline were significantly smaller in coronary and carotid arteries than in iliac, mesenteric, and renal arteries. The presence of endothelium significantly suppressed the contractions in coronary and carotid arteries, but did not affect the contractions in iliac, mesenteric, and renal arteries. The endothelium dependent relaxations to noradrenaline, which were abolished by LNMMA, were more prominent in coronary and carotid arteries than in iliac, mesenteric, and renal arteries. CONCLUSIONS: The contractions via alpha adrenoceptors were small, and endothelium dependent relaxations via alpha 2 adrenoceptors were large, in the coronary and carotid arteries, which perfuse the vital organs (heart and brain), as compared with those in iliac, mesenteric, and renal arteries. These results suggest that in the state of activation of the sympathetic nervous system, arterial tone in response to alpha adrenoceptor stimulation may be regulated not only by alpha adrenoceptors on vascular smooth muscle but also by those on endothelium, through release of endothelium derived relaxing factor (EDRF) via alpha 2 adrenoceptors.     

Impaired endothelium-dependent relaxation after cardiac global ischemia and reperfusion: role of warm blood cardioplegia

OBJECTIVES: Experiments were designed to determine whether coronary endothelial dysfunction after cardiac global ischemia and reperfusion could be prevented by warm blood cardioplegic solution. BACKGROUND: The coronary endothelium produces endothelium-derived relaxing factor (EDRF) to prevent vasospasm and thrombosis. After ischemia and reperfusion, endothelium-dependent relaxation (EDR) is diminished as a result of G-protein dysfunction. METHODS: Dogs were exposed to extracorporeal circulation in 37 degrees C (group 1) or 28 degrees C (groups 2 and 3). The heart was ischemic for 120 min while continuous warm blood cardioplegic solution (group 1) or intermittent cold (4 degrees C) crystalloid cardioplegic solution was not used in group 3 animals. The heart was then allowed to function for 60 min of reperfusion. RESULTS: Endothelium-derived relaxation in response to acetylcholine, adenosine diphosphate and sodium fluoride of the coronary rings of group 1 was significantly different from that of groups 2 and 3 but was not significantly different from that of group 4. In contrast, EDR in response to the receptor-independent calcium ionophore agonist A23187 was not significantly different between the four groups. Scanning electron microscopic studies showed that platelet adhesion and aggregation, area of microthrombi, disruption of endothelial cells and separation of the intercellular junction could be found in coronary segments of groups 2 and 3 but not in vessels of groups 1 and 4. CONCLUSIONS: These experiments suggest that cardiac global ischemia and reperfusion impair receptor-mediated release of EDRF from the coronary endothelium with G-protein dysfunction. This type of coronary endothelial dysfunction can be prevented by continuous anterograde infusion of warm blood cardioplegic solution during global ischemia.     

Adventitial expression of recombinant eNOS gene restores NO production in arteries without endothelium

The current study was designed to determine the effect of recombinant endothelial nitric oxide synthase (eNOS) gene expression on endothelium-dependent relaxations to bradykinin in isolated canine basilar, coronary, or femoral arteries. Arterial rings were exposed ex vivo (30 minutes at 37 degrees C) to an adenoviral vector encoding either the eNOS gene (AdCMVeNOS) or the beta-galactosidase reporter gene (AdCMVbeta-Gal). Twenty-four hours after transduction, transgene expression was evident mainly in the adventitia. Expression of recombinant proteins was much higher in basilar arteries than in coronary or femoral arteries. Rings of control, AdCMVbeta-Gal, and AdCMVeNOS arteries with and without endothelium were suspended for isometric tension recording. Levels of cGMP were measured by radioimmunoassay. In AdCMVeNOS basilar arteries with endothelium, relaxations to low concentrations of bradykinin (3 x 10(-11) to 10(-9) mol/L) were significantly augmented. In contrast, in coronary and femoral arteries with endothelium, AdCMVeNOS transduction did not affect relaxations to bradykinin. Removal of the endothelium abolished bradykinin-induced relaxations in control and AdCMVbeta-Gal basilar arteries. However, in basilar arteries transduced with AdCMVeNOS even when the endothelium was removed, stimulation with bradykinin (3 x 10(-11) to 10(-9) mol/L) caused relaxations as well as increases in cGMP production. The relaxations to bradykinin were completely blocked by an NOS inhibitor, NG-nitro-L-arginine methyl ester. Electron microscopic analysis revealed that recombinant eNOS protein was expressed in fibroblasts of the basilar artery adventitia. These results suggest that genetically modified adventitial fibroblasts may restore production of NO in cerebral arteries without endothelium. Our findings support a novel concept in vascular biology that fibroblasts in the adventitia may play a role in the regulation of vascular tone after successful transfer and expression of recombinant eNOS gene.     

Increased production of nitric oxide in coronary arteries during congestive heart failure

Experiments were designed to determine whether a heterogeneity of endothelium-dependent relaxations in arteries from different vascular beds exists in experimental congestive heart failure (CHF) and to determine the mediators of those responses. CHF was produced in dogs by rapid ventricular pacing for 15 d. Rings of coronary, femoral, and renal arteries with and without endothelium from control and CHF dogs were suspended in organ chambers for measurement of isometric force. In arteries contracted with prostaglandin F2 alpha, endothelium-dependent relaxations to BHT 920 (an alpha 2-adrenergic agonist) were increased in coronary arteries from dogs with CHF (maximal relaxation: control -15 +/- 9% vs CHF -92 +/- 5%; n = 5-6; P < 0.05), with a modest enhancement in renal arteries. Relaxations to adenosine diphosphate and the calcium ionophore were unchanged. Relaxations to BHT 920 in CHF were reduced by NG monomethyl-L-arginine (L-NMMA) and pertussis toxin but not by indomethacin. These data suggest that endothelium-dependent relaxations are affected heterogeneously in CHF. The enhanced response to alpha 2-adrenergic agonists in the coronary artery is mediated by nitric oxide through a mechanism sensitive to inhibition by pertussis toxin. This selective increase in endothelium-dependent relaxations in the coronary artery may contribute to preserving coronary blood flow during CHF.     

Endothelial dysfunction in spontaneously hypertensive rats: consequences of chronic treatment with losartan or captopril

BACKGROUND: Hypertension is associated with endothelial dysfunction characterized by decreased endothelium-dependent relaxations and increased endothelium-dependent contractions. Angiotensin converting enzyme inhibitors and thromboxane A2 receptor antagonists decreased the endothelium dysfunction in hypertensive animals. OBJECTIVE: To investigate the effects of prolonged treatment with losartan on endothelium-dependent and -independent relaxations and contractions in aortic rings from spontaneously hypertensive rats (SHR). MATERIAL AND METHODS: Male SHR aged 16 weeks were treated for 12 consecutive weeks either with 10 mg/kg losartan per day or with 60 mg/kg captopril per day administered via their drinking water. The systolic blood pressure was evaluated basally and during week 12. At the end of the treatment period, the vascular reactivity in aortic rings was studies. A group of rats treated with captopril was studies as a reference group. RESULTS: Losartan and captopril reduced the blood pressure significantly and comparably. Both drugs enhanced acetylcholine-induced relaxations and reduced the maximal contractile response to acetylcholine in the presence of NG-nitro-L arginine methyl ester (L-NAME). Contractile responses to phenylephrine, endothelin-l and U46619 were not affected by these treatments. Increased relaxing responses to superoxide dismutase were observed only in captopril-treated rats. Losartan reduced the contractile response to angiotensin II. By contrast this contractile response was elevated in rats treated with captopril. CONCLUSIONS: Prolonged antihypertensive treatments with losartan and captopril decreased the endothelial dysfunction in aortic rings from SHR not only by enhancing NO-dependent relaxations but also by reducing the contractions in response to an endothelium-derived contracting factor. The results further confirm that an endothelium-derived contracting factor plays a role in vascular dysfunction in SHR and the relationships between this factor and angiotensin II.     

The endothelium-dependent response of human hepatic artery after preservation with the UW solution

The University of Wisconsin's (UW) solution has been used commonly for current liver transplantation. However, its effect on the vascular endothelium remains unclear. Experiments were designed to study the effects. Human hepatic arteries harvested from patients with hepatocellular carcinoma undergoing liver resection were preserved in 4 degree C physiological solution (group 1, the content showed on the text) and UW solution (group 2) for 1 hr. Segments of preserved and control (group 3) hepatic arteries were suspended in organ chamber to measure the isometric force. The relaxations to acetylcholine (ACH) and adenosine diphosphate in segments of hepatic artery with endothelium were significantly greater than those segments without endothelium. The maximal relaxation to ACH in arterial segments with endothelium of group 2 was significantly different from those of group 1 and 3 (group 1 to group 3, 82 +/- 2%, 57 +/- 6%, and 83 +/- 4% of the initial tension contracted by neoepinephrine (3 X 10-7 mole/l, P < 0.05). The maximal relaxation to adenosine diphosphate was similar to the response to ACH. Perfusate hypoxia (oxygen tension 30 +/- 5 mmHG) caused endothelium-dependent contraction of the arterial segments (group 1 to group 3, 233 +/- 32%, 276 +/- 35%, and 251 +/- 40% of the initial tension, P < 0.05). Endothelium-independent relaxation and contraction of human hepatic artery to sodium nitroprusside and norepinephrine were not altered by UW solution. In summary, the impaired endothelium-dependent relaxation by UW solution and prominent endothelium-dependent contraction to hypoxia of human hepatic artery would favor vasospasm and thrombus formation after liver transplantation.     

Genetic construction and characterization of an anti-monkey CD3 single-chain immunotoxin with a truncated diphtheria toxin

In arteries, adrenomedullin (ADM) causes relaxations of rings with and without endothelium by stimulating accumulation of cyclic nucleotides resulting from activation of the ADM and calcitonin gene-related peptide (CGRP) receptors. Experiments were designed to determine the mechanism(s) of relaxation to ADM in veins. Rings of canine femoral vein with and without endothelium were suspended in organ chambers for measurement of isometric force. Rings were contracted with prostaglandin F2alpha (2 x 10(-6) M), and cumulative dose-responses to ADM (10(-11) to 10(-7) M) were obtained in the absence or presence of indomethacin (10(-5) M), indomethacin + N(G)-monomethyl-L-arginine (10(-4) M), methylene blue (10(-5) M), particulate guanylate cyclase inhibitor HS-142-1 (10(-5) M), tetraethylammonium (TEA, 10(-2) M), CGRP-receptor antagonist (CGRP 8-37, 10(-6) M), ADM-receptor antagonist (ADM 26-52, 10(-6) M), diphenhydramine (10(-6) M), 8-phenyltheophylline (3 x 10(-6) M), or superoxide dismutase (150 U/ml) plus catalase (1,200 U/ml). ADM produced concentration-dependent relaxations only in veins with endothelium. Relaxations to ADM in rings with endothelium were significantly inhibited only by methylene blue and HS-142-1. In separate experiments, incubation of rings with ADM (10(-8) M) and 3-isobutyl-1-methyl-xanthine (10(-4) M) for 3 min did not significantly affect the accumulation of cyclic guanosine monophosphate (cGMP) and cyclic adenosine monophosphate (cAMP). These data suggest that ADM-mediated relaxation in veins is endothelium dependent and is not associated with activation of CGRP receptors or currently defined ADM receptors. Further, relaxations are not mediated by nitric oxide, indomethacin-sensitive prostanoids, TEA-sensitive hyperpolarizing factors, oxygen free radicals, or accumulation of cyclic nucleotides.     

Effect of low density lipoprotein on adenosine receptor-mediated coronary vasorelaxation in vitro

We investigated the effect of low density lipoprotein (LDL) on vasorelaxations and nitric oxide generation induced by the adenosine analogs, 5'-(N-ethylcarboxamide)adenosine, 2-p-(2-carboxyethyl)phenylethyl-amino-5'N-ethylcarboxamidoadenosine and/or 2-chloroadenosine in porcine coronary artery rings in vitro. Preincubation of tissues with native LDL (100 and 200 microg/ml) for 4 hr in the absence or presence of copper sulfate (5 microM) selectively attenuated the endothelium-dependent relaxations elicited by 5'-(N-ethylcarboxamide)adenosine and 2-p-(2-carboxyethyl)phenylethyl-amino-5'N-ethylcarboxamideoadenosine+ ++ without altering the response to 2-chloroadenosine which produced endothelium-independent relaxation. The 4-hr exposure of tissues to native LDL (100 microg/ml) also inhibited the production of nitrite induced by 5'-(N-ethylcarboxamide)adenosine in endothelium-intact rings. These effects were associated with enhanced oxidation of the lipoprotein. The inhibitory action of LDL on tissue relaxations and nitrite generation as well as the oxidation of the lipoprotein were all prevented by high density lipoprotein (100 microg/ml). In contrast, a relatively short period (20 min) of tissue incubation with native LDL produced no alterations of the relaxations and nitrite production evoked by 5'-(N-ethylcarboxamide)adenosine and 2-p-(2-carboxyethyl)phenylethyl-amino-5'N-ethylcarboxamidoadenosine. Under this condition, the oxidation of LDL was not also significantly altered. In conclusion, the results indicate that in coronary artery LDL, with oxidative modification, causes attenuation of nitric oxide-mediated endothelial responses induced by adenosine receptors activation, and this effect is prevented by high density lipoprotein. Such modulation may be of importance in hypercholesterolemia and in the development of atherosclerosis.     

Nitric oxide-mediated beta 2-adrenoceptor relaxation is impaired in mesenteric veins from portal-hypertensive rats

BACKGROUND & AIMS: beta-Adrenergic relaxation seems to be mediated by nitric oxide. The aim of this study was to evaluate changes induced by portal hypertension in beta 2-adrenergic vasorelaxation. METHODS: Isolated rat mesenteric veins were relaxed by salbutamol, and nerve-mediated vasocontractions were induced by electrical field stimulation. Responses were evaluated in the presence of NG-nitro-L-arginine methyl ester (L-NAME) or tetrodotoxin. Immunocytochemical techniques were used for localization of neuronal NO synthase. RESULTS: Salbutamol-induced relaxations were decreased in rings from portal-hypertensive animals. L-NAME reduced these relaxations, but its effects were more pronounced in sham-operated rats. Tetrodotoxin decreased the effect of salbutamol only in rings from sham-operated animals. Combination of L-NAME and tetrodotoxin did not exert a greater effect than either of these agents alone. Veins from portal-hypertensive animals were more sensitive to S-nitroso-N-acetyl penicillamine. L-NAME increased vasocontractions by electrical stimulation only in rings from sham-operated rats. Veins from portal-hypertensive animals exhibited a specific degeneration of NO-containing nerve endings. CONCLUSIONS: beta 2-Adrenergic relaxation is impaired in mesenteric veins from portal-hypertensive rats, possibly as a result of a defective neuronal release of NO.     

13C-NMR spectroscopy of keratan sulphates--assignments for five sialylated pentasaccharides derived from the non-reducing chain termini of bovine articular cartilage keratan sulphate by keratanase II digestion

The effects of dexfenfluramine on the release of brain dopamine and serotonin into striatal dialysates were measured in freely moving rats. Samples collected every 20 min were assayed for dopamine and serotonin by high-performance liquid chromatography in a single run. The administration of a lower anorectic dose of dexfenfluramine (0.5 or 1.0 mg/kg intraperitoneally) significantly increased dialysate serotonin concentrations without affecting those of dopamine. A higher dexfenfluramine dose (2.5 mg/kg intraperitoneally) increased both serotonin and dopamine. The increase in dopamine could be blocked by administering the mixed-acting serotonin antagonist methiothepin (20 microM), and was reproduced by applying serotonin (3-10 microM) directly to striatal neurons. Tetrodotoxin (1 microM) added to the striatal perfusates decreased the basal release of dopamine and serotonin; it also blocked the effect of dexfenfluramine (2.5 mg/kg intraperitoneally) on dopamine release and decreased the increment in serotonin release (by approximately 70%). Amphetamine (1 mg/kg subcutaneously) or phentermine (2 mg/kg intraperitoneally) increased dialysate dopamine concentrations without affecting those of serotonin, and tetrodotoxin (1 microM) failed to block the response to amphetamine. These findings suggest that (1) lower anorectic doses of dexfenfluramine release serotonin but not dopamine, and (2) higher doses of dexfenfluramine also increase dopamine release by an indirect mechanism mediated via serotonin.     

Effects of cell-permeant calcium chelators on contractility in monkey basilar artery

Vasospasm after traumatic or aneurysmal subarachnoid hemorrhage is associated with smooth muscle contraction, a process that results in part from increased intracellular calcium in smooth muscle cells. These experiments tested the hypothesis that chelation of intracellular calcium with the cell-permeant calcium chelator, 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetracetic acid acetoxymethyl ester (BAPTA-AM), decreases smooth muscle contraction in response to agents that cause contraction by increasing intracellular calcium. Effects of BAPTA-AM on vasoconstriction induced by KCl, prostaglandin F2alpha (PGF2alpha), caffeine, and erythrocyte hemolysate were tested on monkey basilar artery under isometric tension. BAPTA-AM, 30 and 100 micromol/L, caused a significant decrease in resting tension in rings with and without endothelium (30 micromol/L; 8+/-6% [n.s.] and 14+/-5%, 100 micromol/L; 19+/-3% and 32+/-6%,p < 0.05, paired t test). Contractions to caffeine were significantly decreased by 30 micromol/L BAPTA-AM and were abolished at 100 micromol/L in rings with and without endothelium (p < 0.05). BAPTA-AM, 100 micromol/L, competitively inhibited contractions to PGF2alpha. BAPTA-AM, 100 micromol/L, significantly decreased the maximum contractions to KCI in rings with and without endothelium (p < 0.05). There were no significant effects of BAPTA-AM on contractions induced by hemolysate in rings with endothelium but in rings without endothelium, BAPTA-AM, 100 micromol/L, significantly inhibited contractions. In rings with endothelium contractions to hemolysate could be significantly reduced by BAPTA-AM plus indomethacin or indomethacin alone, suggesting that hemolysate releases an eicosanoid from the endothelium by a pathway that is not inhibited by BAPTA. These results suggest that the ability of BAPTA-AM to inhibit smooth muscle contractions will depend on the agonists mediating the contraction. In response to erythrocyte hemolysate, loading of endothelial cells with BAPTA-AM increases the release of a vasoconstricting eicosanoid from these cells that counteracts the decreased contraction caused by loading of smooth muscle cells with BAPTA-AM.     

Effect of P2-purinoceptor antagonists on hemolysate-induced and adenosine 5'-triphosphate-induced contractions of dog basilar artery in vitro

OBJECTIVE: To test the hypothesis that the vasoactive effects of hemolysate of dog erythrocytes on dog basilar artery in vitro are caused by adenosine 5'-triphosphate (ATP). METHODS: Dog erythrocyte hemolysate was assayed for ATP by high-pressure liquid chromatography. Dog basilar arteries were cut into rings and studied under isometric tension to determine the effects of the P2-purinoceptor antagonists suramin, pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid, and reactive blue 2 on contractions induced by hemolysate, prostaglandin F2 alpha (PGF2 alpha), KCl, uridine 5'-triphosphate, and ATP. RESULTS: Dog erythrocyte hemolysate contained 34 mumol/L of ATP. Hemolysate produced concentration-dependent contractions of dog basilar artery. Suramin (100 mumol/L) significantly inhibited contractions to hemolysate, ATP, and uridine 5'-triphosphate but not to PGF2 alpha and KCl (P < 0.05). Pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid (100 mumol/L) caused a small but significant reduction of the contractions to hemolysate and did not affect contractions to PGF2 alpha and KCl. Reactive blue 2 (30 mumol/L) produced significant inhibition of contractions to hemolysate and PGF2 alpha but did not affect contractions to KCl. CONCLUSION: These findings suggest that ATP mediates a smooth muscle contractile response of hemolysate on dog basilar artery. Because erythrocyte cytosol is known to be important in the pathogenesis of vasospasm, these results suggest that ATP may contribute to the vasoconstriction that occurs in vasospasm.     

Effect of the vascular endothelium on contractions induced by prostaglandin F2 alpha in isolated pregnant guinea pig uterine artery

The purpose of this study was to examine the influence of endothelium on prostaglandin F2 alpha-mediated contractions in pregnant guinea pig uterine artery. Consequently, the effects of prostaglandin F2 alpha on pregnant guinea pig uterine arterial rings with both intact and denuded endothelium were studied. In vessels with denuded endothelium prostaglandin F2 alpha (0.1-10 microM) induced contraction (pD2 = 6.17) with greater potency than in vessels with intact endothelium (pD2 = 5.68). NG-Monomethyl-L-arginine (10 microM) did not affect the concentration-response curve for prostaglandin F2 alpha, regardless of endothelial condition. In contrast, in both types of preparation, indomethacin (10 microM) increased the maximal response value obtained with prostaglandin F2 alpha, but this effect was significantly greater in preparations with intact than in those with denuded endothelium (128.3 versus 206.5%). Moreover, indomethacin shifted the concentration-response curve for prostaglandin F2 alpha to the left only in preparations with intact endothelium. The pKA values for prostaglandin F2 alpha itself did not differ between preparations: 5.41 and 5.52 for pregnant guinea pig uterine artery with and without endothelium, respectively. The receptor reserve expressed as KA/EC50 was significantly greater in rings with denuded (4.44) compared to those in rings with intact endothelium (1.86). We conclude that prostaglandin-F2 alpha-induced contraction in pregnant guinea pig uterine artery is modulated by the vascular endothelium. It is probable that cyclooxygenase products relating to vasodilatation and derived from endothelium mediate this effect, acting as a functional endogenous antagonist and thereby reducing the apparent efficacy and potency of prostaglandin F2 alpha.     

The endothelium of the rat renal artery plays an obligatory role in A2 adenosine receptor-mediated relaxation induced by 5'-N-ethylcarboxamidoadenosine and N6-cyclopentyladenosine

Studies were undertaken in the rat isolated renal artery in order to determine if adenosine receptor agonists were capable of inducing the release of nitric oxide from the renovascular endothelium. N6-cyclopentyladenosine (CPA) and 5'-N-ethylcarboxamidoadenosine (NECA) produced concentration-dependent relaxations in endothelium intact renal artery rings. The NECA curve was biphasic with a first phase pA50 of 6.05. The CPA curve was monophasic with a pA50 of 4.35. In the absence of endothelium the curves to both NECA and CPA were monophasic with pA50 values of 3.37 and 3.50, respectively. The A2a adenosine receptor-selective agonist CGS21680 (2-[p-(2-carboxyethyl)-phenethylamino]-5'-N-ethylcarboxamidoadenos ine) was inactive in endothelium intact tissues. Relaxant responses to CPA and NECA in the presence of endothelium were antagonized by 8-p-sulfophenyltheophylline and by 1,3-dipropyl-8-cyclopentylxanthine only at a nonselective concentration (3 x 10(-6) M) suggesting activation of A2 adenosine receptors. The responses to CPA and NECA in the absence of endothelium are not due to activation of A1 or A2 adenosine receptor subtypes because they are resistant to blockade by these xanthines. CPA and NECA responses in the presence of endothelium were inhibited by NG-nitro-L-arginine methylester (L-NAME), a nitric oxide synthase inhibitor, but not by the cyclooxygenase inhibitor indomethacin or the K+ATP channel antagonist glibenclamide. These results suggest that the rat renal artery contains A2b adenosine receptors that are located exclusively on the endothelium and cause the release of nitric oxide.     

Antihypertensive and vasodilator actions of antioxidants in spontaneously hypertensive rats

This study was designed to determine whether the antioxidants ascorbic acid, aminotriazole, and glutathione acutely reduce blood pressure (BP) by endothelium-independent or -dependent vasorelaxation in spontaneously hypertensive rats. Blood pressure of male Wistar-Kyoto (WKY) and spontaneously hypertensive rats (SHR) was measured before and 4 h after administration of antioxidants. Thoracic aortic rings with and without endothelium were suspended in organ chambers for isometric tension recordings. Each of the antioxidants, administered in vivo, significantly decreased blood pressure in SHR but had no significant effect on BP in WKY rats. The endothelium-dependent impaired relaxation of SHR aortic rings to acetylcholine (ACh) was improved by prior in vivo administration of each antioxidant. ACh-induced relaxations of aortic rings from WKY was not affected by prior antioxidant treatment. Addition of each antioxidant directly to the organ chamber containing SHR or WKY aortas produced dose- and endothelium-dependent relaxations. Moreover, antioxidant pretreatment of SHR aortic rings significantly potentiated ACh-induced relaxations in these aortas, suggesting that this effect was endothelium dependent. Relaxations induced by the antioxidants alone or by ACh in the presence of antioxidants were inhibited by addition of either xanthine plus xanthine oxidase or nitro-L-arginine. These findings suggest that either excess production of oxidants or a deficiency of antioxidant systems may contribute to the high blood pressure and the endothelium-dependent impairment of vascular relaxation in SHR.     

Effects of en bloc esophagectomy on nutritional and immune status in patients with esophageal carcinoma

We tested the hypothesis that extravascular adenosine induces the release of vasodilatory products from endothelial cells lining skeletal muscle vessels. Endothelium-intact (n = 35) and -denuded (n = 5) dog semitendinosus intramuscular arteries were isolated, cannulated, and placed in 100-mL baths containing Krebs-Henseleit bicarbonate buffer (Krebs) at 37 degrees C and gassed with 95% O2--5% CO2. Each vessel, as well as a parallel tubing segment (avascular control), was perfused at 3.5 +/- 0.2 mL/min (inflow pressure 94 +/- 2 mmHg; 1 mmHg = 133.3 Pa) with Krebs containing 100 microM phenylephrine, 6% dextran, and 15 units/mL superoxide dismutase. Perfusate from all segments dripped onto endothelium-denuded dog femoral artery rings. The addition of 10 microM acetylcholine to the perfusate to test the functional integrity of endothelium-intact donor segments did not alter resistance in vessel segments or change force in rings. The addition of 100 microM adenosine to the extravascular bath decreased resistance 1.5 +/- 0.4 mmHg.mL-1.min-1 in vessel segments but was without effect on downstream rings. When acetylcholine was retested in the presence of extravascular adenosine, a relaxation (16 +/- 6%) occurred in rings receiving perfusate from endothelium-intact segments but not endothelium-denuded or tubing segments. This relaxation was eliminated by N omega-nitro-L-arginine (10 microM), a nitric oxide synthase inhibitor, and was attenuated to 4 +/- 1% by 8-phenyltheophylline (10 microM), an adenosine receptor antagonist. Thus adenosine, in conjunction with acetylcholine, acting through a receptor-mediated event, resulted in the release of nitric oxide from the endothelium of perfused intramuscular arteries, indicating the potential for extravascular conditions to influence the release of endothelium-derived products.     

Vascular hyporesponsiveness in aortic rings from cirrhotic rats: role of nitric oxide and endothelium

1. The role of nitric oxide as mediator of the vascular alterations present in different models of experimental liver cirrhosis is controversial. In the present study, we evaluated the role of nitric oxide and that of the endothelium in the response to phenylephrine and acetylcholine of isolated aortic rings from chronic bile duct-ligated (29 days) rats and their corresponding controls. Experiments were performed in rings with or without endothelium, in rings pretreated with N-omega-nitro-L-arginine methyl ester (10(-4) mol/l) to inhibit nitric oxide synthesis and in rings pretreated with aminoguanidine (10(-4) mol/l) to inhibit inducible nitric oxide synthesis. 2. Under basal conditions, the maximum absolute tension developed in response to cumulative addition of phenylephrine was significantly decreased in rings from bile duct-ligated animals (1.62 +/- 0.06 g) compared with the control rings (2.15 +/- 0.099). This hyporesponsiveness to phenylephrine of rings from bile duct-ligated animals was corrected after treatment with N-omega-nitro-L-arginine methyl ester and reduced, but not completely eliminated, in rings without endothelium. In contrast, aminoguanidine did not modify the lower response to phenylephrine rings from bile duct-ligated animals. ED50 values were not different between groups under any experimental conditions. 3. The endothelium-dependent vasodilatation to acetylcholine in phenylephrine-constricted rings was similar in both groups of animals, control and bile duct ligated, under all experimental conditions. N-omega-nitro-L-arginine methyl ester pretreatment and removal of the endothelium completely abolished the response to acetylcholine in cirrhotic and control rings. 4. These results demonstrate that in aortic rings from cirrhotic, bile duct-ligated rats, increased production of nitric oxide, mainly of endothelial origin, is responsible for the lower contractile response to phenylephrine. Our data, however, do not support the involvement of the inducible nitric oxide synthase isoform in this alteration. In contrast, endothelial vasodilatory response to acetylcholine is not altered in this model of cirrhosis, which indicates that not all mechanisms of nitric oxide release are abnormal.     

Endothelin-A receptors mediate vascular smooth-muscle response to moderate acidosis in the canine tibial nutrient artery

Perfusate pH may influence the tone of vascular smooth muscle by affecting the release of endothelium-derived vasoactive factors or by directly modulating function of the smooth muscle. This study was designed to investigate the role of endothelium-derived factors on acidosis-induced responses of isolated canine tibial nutrient artery suspended in an organ chamber for the measurement of isometric contractile force. To investigate the specific role of the endothelium in half the rings, the endothelium was removed mechanically. Concentration-response curves to KCl were obtained in the absence or presence of inhibition of two important endothelium-derived relaxing factors, nitric oxide and prostacyclin, and an inhibitor of receptors for the endothelium-derived contracting factor, endothelin-1. Acidification of the perfusate from pH 7.45 to 7.0 significantly attenuated the contractions to KCl in arterial rings with endothelium (the mean of the effective concentration causing 50% of the maximal response for KCl at pH 7.45 and 7.0 was 12.31 +/- 0.40 nM and 14.60 +/- 0.55 nM, respectively). This difference was abolished by mechanical removal of the endothelium. In rings with endothelium, inhibition of nitric oxide or prostacyclin did not abolish the attenuation of KCl-induced contractions occurring with acidosis (the mean of the effective concentration causing 50% of the maximal response for KCl at pH 7.45 and 7.0 was 11.18 +/- 0.60 nM and 13.60 +/- 0.60 nM, respectively). Inhibition of endothelin-A receptors did not alter contractions to KCl at pH 7.45. However, the acidosis-induced attenuation of contractions with KCl was abolished by the endothelin-A-receptor antagonist BQ-123 (the mean of the effective concentration causing 50% of the maximal response at pH 7.45 and 7.0 was 13.8 +/- 1.34 nM and 13.2 +/- 1.34 nM, respectively). These results suggest that acidosis-induced relaxation of canine tibial nutrient artery is endothelium dependent and that activation of endothelin-A receptors during acidosis is coupled to a release of an endothelium-derived relaxing factor.