Concentration-dependent Increases In Tension Research Articles

L-arginine and Arginase Products Potentiate Dexmedetomidine-induced Contractions in the Rat Aorta.

The α2-adrenergic sedative/anesthetic agent dexmedetomidine exerts biphasic effects on isolated arteries, causing endothelium-dependent relaxations at concentrations at or below 30 nM, followed by contractions at higher concentrations. L-arginine is a common substrate of endothelial nitric oxide synthase and arginases. This study was designed to investigate the role of L-arginine in modulating the overall vascular response to dexmedetomidine. Isometric tension was measured in isolated aortic rings of Sprague Dawley rats. Cumulative concentrations of dexmedetomidine (10 nM to 10 μM) were added to quiescent rings (with and without endothelium) after previous incubation with vehicle, N-nitro-L-arginine methyl ester hydrochloride (L-NAME; nitric oxide synthase inhibitor), prazosin (α1-adrenergic antagonist), rauwolscine (α2-adrenergic antagonist), L-arginine, (S)-(2-boronethyl)-L-cysteine hydrochloride (arginase inhibitor), N-hydroxy-L-arginine (arginase inhibitor), urea and/or ornithine. In some preparations, immunofluorescent staining, immunoblotting, or measurement of urea content were performed. Dexmedetomidine did not contract control rings with endothelium but evoked concentration-dependent increases in tension in such rings treated with L-NAME (Emax 50 ± 4%) or after endothelium-removal (Emax 74 ± 5%; N = 7 to 12). Exogenous L-arginine augmented the dexmedetomidine-induced contractions in the presence of L-NAME (Emax 75 ± 3%). This potentiation was abolished by (S)-(2-boronethyl)-L-cysteine hydrochloride (Emax 16 ± 4%) and N-hydroxy-L-arginine (Emax 18 ± 4%). Either urea or ornithine, the downstream arginase products, had a similar potentiating effect as L-arginine. Immunoassay measurements demonstrated an upregulation of arginase I by L-arginine treatment in the presence of L-NAME (N = 4). These results suggest that when vascular nitric oxide homeostasis is impaired, the potentiation of the vasoconstrictor effect of dexmedetomidine by L-arginine depends on arginase activity and the production of urea and ornithine.

Effects of Levosimendan on Hydrogen Peroxide Induced Contraction in Human Saphenous Vein

Aim: Increased oxidative stress plays important roles in vascular dysfunction in patients undergoing coronary artery bypass graft surgery. Hydrogen peroxide (H2O2) is used as an experimental model for oxidative stress. The present study was designed to assess the effects of levosimendan pretreatment on the contractile effects induced by H2O2 in human saphenous vein (HSV) segments. Methods: We studied H2O2 induced contractions of isolated HSV mounted in standard tissue baths. H2O2 (10-6 – 10-3 M) was added cumulatively to the organ bath. Concentration-response curves to H2O2 were repeated in the presence of levosimendan (10–8 M). In the second series of experiments, strips were contracted with 5-HT (10–5 M). When the contraction reached a stable plateau, H2O2 was administrated cumulatively into the organ bath. The same procedure was conducted in the presence of levosimendan. Results: Pretreatment of the SV strips with levosimendan significantly reduced the contractile response to each concentration of H2O2 . 5-HT produced contractions in SV strips. Further treatment of these strips with H2O2 resulted in statistically significant concentration-dependent increases in tension. Preincubation of the tissues with levosimendan did not significantly influence the maximum amplitude of the 5-HT-induced tone but inhibited the contractile effect of H2O2 on the 5-HT-induced contraction. Conclusion: Pretreatment of HSV with clinical concentrations of levosimendan inhibits the vasoconstriction caused by oxidative stress, indicating its potential preventive effect against oxidative stress induced graft spasm.

Role of ERK1/2 signaling pathways in 4-aminopyridine-induced rat pulmonary vasoconstriction

The aim of the present study was to investigate the contribution of extracellular signal regulated kinase-1/2 (ERK1/2) to pulmonary artery contraction in response to 4-aminopyridione (4-AP), an inhibitor of a voltage-gated K + channels that regulate pulmonary vascular tone. Pulmonary artery rings 1–1.5 mm in diameter from male adult Wistar rat were isolated and cut into 3-mm in length. ERK1/2 up-stream kinase (MEK) inhibitors 2′-amino-3′-methoxyflavone (PD98059) and 1,4-diamino-2,3-dicyano-1,4-bis (2-aminophenylthio)-butadiene (U0126), which block the activation of ERK1/2, were used to test the role of ERK1/2 in 4-AP induced pulmonary arterial vasoconstriction and the influences of 4-AP on expressions of phosphorylated ERK1/2 (p-ERK1/2) in cultured rat pulmonary arterial smooth muscle cells (PASMCs) and whole tissues. Our results show that 4-AP elicited concentration-dependent increases in tension of rat pulmonary artery rings, effects that were reduced by pretreatment of the rings with ERK inhibitors, PD98059 (20 μM) and U0126 (10 μM). Moreover, 4-AP increased the expressions of p-ERK1/2 in cultured PASMCs s and whole tissues, which were prevented by pretreatment of the cells or tissues with U0126. These results indicate that ERK1/2 signaling pathway contributes to pulmonary vasoconstriction induced by 4-AP.

The Sympathomimetic Actions of l-Ephedrine and d-Pseudoephedrine: Direct Receptor Activation or Norepinephrine Release?

The basic mechanisms by which ephedrine is preferred over other vasopressors in obstetric anesthesia have not been clearly defined. We examined the sympathomimetic effects of l-ephedrine, currently used as a vasopressor, and d-pseudoephedrine, currently used as a decongestant. In anesthetized rats, l-ephedrine and d-pseudoephedrine caused dose-dependent increases in arterial blood pressure and heart rate, and these effects disappeared after destruction of the sympathetic nerve terminals with 6-hydroxydopamine (6-OHDA) pretreatment. The two ephedrine isomers produced concentration-dependent increases in tension of anococcygeal muscle and sinus rate of right atrium from rats. However, the anococcygeal and atrial responses to d-pseudoephedrine were abolished after 6-OHDA pretreatment, whereas approximately 50% of the responses to l-ephedrine were 6-OHDA-resistant. In human umbilical artery and vein, the two isomers failed to generate any contraction when given at the concentration that is capable of producing significant effects on anococcygeal and atrial tissues. Although direct adrenoceptor activation with l-ephedrine was detectable at tissue levels, the pressor response in vivo was entirely attributable to norepinephrine release from sympathetic nerves. This indirect mechanism could partly explain why l-ephedrine is better at increasing maternal arterial blood pressure while preserving the uteroplacental blood flow that is devoid of the involvement of the sympathetic innervation. The indirectly sympathomimetic property of l-ephedrine may be one of the mechanisms to explain why ephedrine is preferred over alpha-adrenergic agonists as a vasopressor for treatment of intraspinal anesthesia-induced hypotension in obstetrics.

Role of Na +/H + exchanger in acetylcholine-mediated pulmonary artery contraction of Spontaneously hypertensive rats

Compared to sympathetic nervous system, the role of parasympathetic innervation on tone development, especially under diseased conditions, of the pulmonary artery is relatively unknown. In this study, the contractile effect of acetylcholine and the type(s) of muscarinic (M) receptor involved in the pulmonary artery (1st intralobar branch; endothelium-denuded, under resting tension) of the normotensive Wistar–Kyoto (WKY) and age-matched (male, 22–26 weeks old) Spontaneously hypertensive rats (SHR) were investigated. Cumulative administration of acetylcholine (≥0.1 μM) caused a concentration-dependent increase in tension (antagonised by p-fluoro-hexahydro-sila-difenidol and 4-diphenylacetoxy- N-methylpiperidine, both are selective muscarinic M 3 receptor antagonists) and the magnitude of maximum contraction (expressed as % of 50 mM [K +] o-induced contraction) was markedly enhanced in the presence of neostigmine (10 μM, an anti-cholinesterase) (acetylcholine 30 μM, SHR: 72% vs. 35%; WKY: 32% vs. 20%). In SHR only, acetylcholine-elicited contraction was suppressed by 1-[β-[3-(4-Methoxyphenyl)-propoxyl]-4-methoxyphenethyl]-1H-imidazole (SK&F 96365, 1 μM), amiloride (500 μM), ethyl-isopropyl-amiloride (EIPA, 10 μM), 2-[2-[4-(4-Nitrobenzyloxy)phenyl]ethyl]isothiourea (KB-R 7943, 5 μM), 2,4-dichlorobenzamil (10 μM), and an equal molar substitution of [Na +] o (≤30 mM) with choline or N-methyl- d-glucamine. In nominally [Ca 2+] o-free, EGTA (0.5 mM)-containing Krebs' solution, acetylcholine (≥3 μM) only elicited a small contraction. In conclusion, muscarinic M 3 receptor activation is responsible for the pulmonary artery contraction induced by acetylcholine, with a greater magnitude observed in SHR. The exaggerated contraction in SHR is probably due to an influx of [Na +] o through the Na +/H + exchanger and the store-operated channels (SOC) into smooth muscle cells. Elevation of cytosolic [Na +] i subsequently leads to an influx of [Ca 2+] o through the reverse mode of the Na +/Ca 2+ exchanger seems to play a permissive role in mediating the exaggerated contractile response of acetylcholine recorded in the SHR.

Guanylyl cyclase mediates ANP-induced vasoconstriction of murine splenic vessels.

We have previously shown that ANP causes differential constriction of the splenic vasculature of the rat (veins greater than arteries), which may be inhibited by blocking the production of cGMP with A7195. In this paper, we report experiments done on vessels derived from guanylyl cyclase (GC)-A knockout mice. Small splenic arteries ( approximately 150-microm diameter) and veins ( approximately 250-microm diameter) were dissected from male GC-A-deficient 129sv mice or age-matched wild-type controls and mounted in a wire myograph. In the wild-type mice, ANP exhibited higher potency in the veins than in the arteries (EC(50) values wild-type mice: artery, 8 +/- 3 x 10(-9) M, n = 5 vs. vein, 6 +/- 4 x 10(-10) M, n = 5; P < 0.05). The concentration-response curve for ANP-induced vasoconstriction was also shifted leftward in denuded compared with intact arteries (EC(50) values: denuded artery: 5 +/- 3 x 10(-10) M, n = 5 vs. intact artery, 8 +/- 3 x 10(-9) M, n = 5; P < 0.05), i.e., the denuded vessels were more reactive. By contrast, ANP caused no significant change in tension from baseline in intact splenic arteries, intact splenic veins, or denuded splenic arteries derived from the GC-A-deficient mice, although these vessels did show normal concentration-dependent increases in tension to phenylephrine. We conclude that ANP causes vasoconstriction in the splenic vasculature by an endothelium-independent mechanism, mediated via guanylyl cyclase.

The calibre of the foramen of Panizza in Crocodylus porosus is variable and under adrenergic control.

The foramen of Panizza is located within the outflow tract of the crocodilian heart, between the left and right aortas. It has been suggested that the foramen of Panizza has a variable calibre, which could explain the profound changes in the distribution of flows and pressure profiles recorded in the right and left aortas. We investigated this possibility using a modified in-situ perfused heart preparation in combination with isolated strip preparations from the outflow tract. In the perfused heart preparation, bolus injections of adrenaline increased the resistance in the foramen of Panizza, indicating a decrease in its diameter. Isolated strip preparations from the outflow tract showed a concentration-dependent increase in tension in response to adrenaline, while vasoactive intestinal polypeptide caused a relaxation in adrenaline pre-contracted strip preparations. We propose that an increase in the diameter of the foramen of Panizza may be important during pulmonary to systemic shunts to allow blood to flow from the left to right aorta (reverse foramen flow) in order to supply the carotid and coronary arteries. During non-shunting conditions, a constricted foramen may prevent excess flow from the right to left aorta during diastole.

Tachykinins contribute to nerve-mediated contractions in the human esophagus

Tachykinins mediate nonadrenergic, noncholinergic excitation in the gastrointestinal tract, but their role in esophageal peristalsis remains unclear. We used muscle strips from the distal third of human esophagus, obtained from patients undergoing esophagectomy for cancer, to investigate the contribution of tachykinins to nerve-mediated contractions. Isometric tension responses to agonists or electrical field stimulation were recorded in circular and longitudinal muscle strips. Tachykinins produced concentration-dependent increases in tension in circular and longitudinal muscle strips, with the following order of potency: beta-Ala(8)-neurokinin (NK) A (4-10) > NKB > substance P, suggesting NK(2) receptor involvement. The NK(2) receptor antagonist, SR48968 (1 micromol/L), inhibited responses to tachykinins in both muscles. Nerve activation produced on- and off-contractions in circular muscle and a duration-contraction in longitudinal muscle. Atropine (10 micromol/L)-insensitive nerve-evoked contractions were identified for the 3 types of responses. SR48968 produced concentration-dependent inhibition of atropine-insensitive on- and off-contractions but had no effect on the duration-contraction. At low stimulus frequency (1 Hz), on-contractions showed greater sensitivity to SR48968 than off-contractions. Nerve-mediated contractions in the human esophagus have a significant atropine-insensitive component. Tachykinins acting on NK(2) receptors can account for some, but not all, of this response, suggesting that other excitatory mechanisms also contribute.

Endothelin plays a role in contractions of isolated pig pulmonary vessels induced by diaspirin cross-linked hemoglobin

The current studies were undertaken to investigate the role of endothelin-1 (ET-1) and its receptors in contractions of isolated pulmonary vessels of the pig induced by diaspirin cross-linked hemoglobin (DCLHb). Second-order pulmonary arteries (PAs) and veins (PVs) were isolated from pigs, cut into rings (4 to 5 mm), and mounted at optimal passive tension in 37°C Krebs-filled tissue baths bubbled with 95% O25% CO2. Isometric tension was recorded continuously. In paired rings, concentration responses to ET-1 (10−10 to 10−7 mol/L), DCLHb (10−9 to 3 × 10−6 mol/L), and N-nitro-L-arginine (LNA) (10−6 to 5 × 10−5 mol/L) in the presence and absence of the ETA receptor antagonist BQ123 (3 × 10−5 mol/L) were determined. PVs and PAs with intact endothelium and rings from which the endothelium was removed (denuded) were pretreated with the ETB receptor antagonist BQ788 to determine the contribution of ETB receptors to ET-1, DCLHb, and LNA responses. ET-1, DCLHb, and LNA caused concentration-dependent increases in tension in all vessels. In the presence of BQ123, the 50% effective concentration (EC50) of ET-1 was significantly increased (by 5-fold to 10-fold) in all vessels. DCLHb concentration responses were significantly attenuated—in the PVs by 45% and in the PAs by 79%—during treatment with BQ123. BQ123 attenuated LNA responses in PVs by 35% and in PAs by 87%. Treatment with BQ788 had no effect on endothelium-intact PVs or PAs but significantly increased ET-1 EC50 (log of the molar concentration) from −9.0 ± 0.22 to −7.8 ± 0.05 in denuded PAs. The ET-1 EC50 was significantly decreased in denuded PAs (−9.0 ± 0.22) as compared with responses in endothelium-intact PAs (−8.1 ± 0.18). DCLHb concentration responses were attenuated by 71% and LNA responses by 80% during antagonism with BQ788 in the intact PAs only. These data demonstrate that ET-1 plays a role in DCLHb-induced contractions in the PA and PV. The contributions of ET are mediated by both ETA and ETB receptors in the PA but only by ETA receptors in the PV. These results suggest that the vasoconstrictor actions of DCLHb, which have previously been shown to depend on its interference with endothelium-generated NO, may also involve ET. This may reflect the importance of the interaction of these two endothelium-generated physiologic antagonists in the pulmonary circulation.

Functional evidence for a novel suramin-insensitive pyrimidine receptor in rat small pulmonary arteries.

Uridine nucleotides are known to cause constriction of pulmonary arterial smooth muscle. However, the P2 receptor subtypes underlying the contractile effects of these nucleotides in the pulmonary circulation have not been determined. We have used myography and the patch-clamp recording technique to compare the effects of UTP and UDP in isolated small pulmonary arteries (diameter 100 to 400 microm) and their constituent smooth muscle cells. In endothelium-denuded arteries, both UTP and UDP (0.01 to 3 mmol/L) induced concentration-dependent increases in tension that were independent of P2X receptor stimulation. The UDP-mediated increase in tension was significantly less sensitive to the nonselective P2 receptor blocker suramin than the UTP-mediated increase in tension. In single isolated arterial myocytes, voltage-clamped at -50 mV (close to the resting membrane potential of these cells), application of both UTP and UDP evoked periodic oscillations of inward current primarily because of a Ca2+-activated Cl- current (ICl,Ca). Oscillations of ICl,Ca evoked by UTP were reversibly inhibited by suramin, although those evoked by UDP were insensitive to the antagonist. In addition to confirming the presence of classical P2Y2 receptors, these results also provide functional evidence for the existence of a novel UDP receptor in pulmonary arterial myocytes, which may contribute to pyrimidine-evoked vasoconstriction. This notion is supported by molecular evidence that demonstrates the presence of P2Y6 receptor transcripts in rat pulmonary arterial smooth muscle.

ET-1 modulates KCa-channel activity and arterial tension in normoxic and hypoxic human pulmonary vasculature.

The molecular mechanisms by which endothelin (ET)-1 induces pulmonary hypertension are poorly understood. We investigated the effects of ET-1 on outward K+ currents of normoxic and chronically hypoxic human pulmonary arterial (PA) smooth muscle cells (HPSMCs). In normoxic HPSMCs, ET-1 has dual effects. In intact cells, 5 nM ET-1 activates the large-conductance and Ca2+-activated K+ (KCa)-channel current [IK(Ca)] by increasing intracellular Ca2+ concentration, whereas it directly inhibits IK(Ca) in isolated membrane patches. At a higher concentration (10 nM), ET-1-induced IK(Ca) inhibition predominates. In hypoxic HPSMCs, ET-1 at 5 nM significantly reduces IK(Ca). The ETA-receptor antagonist BQ-123 reverses the ET-1-induced decrease in IK(Ca). Chronic BQ-123 treatment also prevents the hypoxia-induced decrease in IK(Ca). In PA rings obtained from human organ donors, ET-1 causes a concentration-dependent increase in tension. The ET-1-mediated increase in tension is reversed by a KCa-channel agonist. The increase in tension at the highest concentration studied (9 nM) was more pronounced in PA rings obtained from patients with chronic obstructive pulmonary disease. These results imply that an ET-1-induced decrease in IK(Ca) contributes to chronic hypoxia-induced pulmonary hypertension.

Characterization of endothelin receptors in human varicose veins

Experiments were designed to characterize endothelin receptors in human varicose veins. Three groups of veins were studied: (1) varicose vein (VV) tributaries of the greater saphenous vein from patients who were undergoing vein stripping for primary varicosity; (2) greater saphenous veins (SVs) from the same patients; and (3) greater saphenous veins from patients without varicosity who were undergoing arterial reconstruction (control). Veins were either cut into rings and suspended in organ chambers for measurement of isometric force, prepared for receptor binding of membrane proteins, or were prepared for measurement of preproendothelin mRNA by reverse transcriptase-polymerase chain reaction (RT-PCR). Endothelin-1 (10(-11) to 10(-7) mol/L) produced similar concentration-dependent contractions in rings with or without endothelium. Maximal tensions were significantly greater in control veins compared with either SVs or VVs. Sarafotoxin S6c (10(-11) to 3 x 10(-7) mol/L), which is selective for the endothelin-B receptor, also produced concentration-dependent increases in tension in all veins. Sarafotoxin S6c responses in VVs were shifted significantly rightward compared with either SVs or control. Maximal tensions to sarafotoxin S6c also were significantly greater in control veins compared with either SVs or VVs. In receptor binding studies, the number of binding sites as defined by competitive inhibition of 125I-endothelin-1 by endothelin-1 was less in VVs than control veins. Competitive inhibition of 125I-endothelin-1 with endothelin-3 (both A and B receptors) or sarafotoxin S6c (B receptors only) suggests that the difference in receptor number between varicose and nonvaricose veins is attributable to differences in the endothelin-B receptor subtype. Binding affinities were not significantly different for either of the receptor subtypes in all veins studied. Preproendothelin mRNA as quantitated by RT-PCR tended to be higher in VVs compared with either SVs or control veins. Decreased contractions to endothelin-1 in both varicose and saphenous veins of patients with primary varicosity may be associated with a decrease in the number of receptors. These receptors may be downregulated in response to increased production of endothelin-1, which is regulated at the transcriptional level.

Hemoglobin-induced contraction of pig pulmonary veins

The effects of hemoglobin A o (HbA o), αα cross-linked hemoglobin (ααHb), cyanomet αα cross-linked hemoglobin (cyanometααHb), and human serum albumin (HSA) were compared under basal conditions and during relaxation with acetylcholine (ACh), sodium nitroprusside (SNP), and papaverine (PAP) in porcine pulmonary veins. Isometric tension changes were recorded in isolated rings (3 to 4 mm) that were suspended in Krebs solution bubbled with 95% O 2/5% CO 2. Increasing concentrations of HbA o and ααHb (10 −9−3 × 10 −6 mol/L) caused concentration-dependent increases in tension that reached a maximum of 4.20 ± 0.3 gm and 3.78 ± 0.6 gm, respectively. CyanometααHb and HSA (10 −9− 3 × 10 −6mol/L) did not cause significant increases in tension. The maximum responses to HbA o and ααHb were significantly increased during relaxation with ACh and SNP but not with PAP. In contrast, SNP (10 −4 mol/L) and PAP (10 −5 mol/L), but not ACh, reversed contractions induced by HbA o and ααHb. These studies support the concept that hemoglobin-induced vascular contraction is primarily mediated by inactivation of the vasodilator nitric oxide in vitro. We suggest that this mechanism is common to acellular hemoglobins in which the ligand binding site is unimpaired and in which the heme iron is in the ferrous ( +2) state.

Chronic increases in blood flow upregulate endothelin-B receptors in arterial smooth muscle.

Experiments were designed to characterize endothelin receptors in arteries after chronic increases in blood flow. A fistula was created between the femoral artery and vein in one hindlimb of dogs; contralateral blood vessels were sham operated. Sham- and fistula-operated arteries were removed 6 wk postoperatively. Some arteries were prepared for measurement of isometric force or for isolation of membrane proteins. Other arteries were used for histological staining with an endothelin-B (ETB) receptor antibody. In arteries suspended for the measurement of isometric force, endothelin-1 produced concentration-dependent increases in tension that were significantly greater in fistula- than in sham-operated arteries without endothelium. The ETB-receptor-selective peptide sarafotoxin S6c produced concentration-dependent increases in tension only in fistula-operated arteries. In receptor-binding studies of membrane proteins, Scatchard analysis of saturation binding with 125I-labeled endothelin-1 (125I-endothelin-1) indicated that the total number of receptors was greater in fistula-operated arteries; affinity was threefold less in fistula- than in sham-operated arteries. Competitive displacement of 125I-endothelin-1 by endothelin-3 was significant for a two-site model in membranes prepared from sham-and fistula-operated arteries. Competitive inhibition of 125I-endothelin-1 binding by sarafotoxin S6c was significant for a one-site binding model in all arteries. Sarafotoxin S6c binding sites were elevated significantly in fistula-operated arteries. Immunohistochemical staining for the ETB receptor was significantly greater in both the endothelium and smooth muscle of fistula- than in sham-operated arteries. These results suggest that chronic increases in blood flow upregulate endothelin receptors, including ETB receptors in arterial smooth muscle.

Effect of angiotensin receptor blockade in the rabbit aorta: influence of the endothelium

Contractile responsiveness of the rabbit aorta (endothelium intact and denuded) to angiotensin I, II, and III was compared. The effects of converting-enzyme inhibition with enalapril, the selective AT1-receptor antagonist (losartan), and the AT2-receptor antagonist (PD 123319) on these contractile profiles were examined. In all preparations, it was found that the angiotensins produced concentration-dependent increases in tension. Differences in sensitivity were encountered; in endothelium-intact preparations, the mean EC50 values (nM with 95% confidence interval in parentheses) for angiotensin I, II, and III were 9 (95% CI 7-11), 40 (20-60), and 30 (10-40), respectively, and for denuded preparations they were 20 (11-29), 0.8 (0.7-0.9), and 30 (20-40), respectively. Enalapril decreased the maximal tension developed to angiotensin I and II, which was greater in endothelium-intact preparations. Losartan was a competitive antagonist against angiotensin I and angiotensin II in both intact and denuded preparations, with pA2 values as follows: against angiotensin I, 9.0 and 9.3 for intact and denuded, respectively; against angiotensin II, 8.3 and 8.9 for intact and denuded, respectively. Losartan antagonized angiotensin III, but the slopes of the Schild analysis were significantly less than unity. In endothelium-intact preparations, PD 123319 failed to significantly antagonize responsiveness to angiotensin I. Against angiotensin II, PD 123319 was a competitive antagonist with a pA2 of 8.3. The antagonism for PD 123319 against angiotensin III was insurmountable. In endothelium-denuded preparations, PD 123319 failed to antagonize angiotensin I and angiotensin III. Although PD 123319 appeared to inhibit the responsivenss of the rabbit aorta by angiotensin II, the slope of the Schild plot was significantly less than unity. These experiments provide evidence that angiotensin I possesses different actions from angiotensin II and III and that a functional endothelium modulates the underlying vascular response to angiotensin. In addition, the endothelium modulates the antagonism by losartan and PD 123319, supporting the notion that the endothelium possesses distinct angiotensin receptors.

Possible role of Na(+)-K(+)-ATPase in the regulation of human corpus cavernosum smooth muscle contractility by nitric oxide.

1. This study was designed to determine the role of sodium-potassium adenosine triphosphatase (Na(+)-K(+)-ATPase) in the regulation of human corpus cavernosum smooth muscle contractility by nitric oxide (NO). In addition, we determined if the modulation of Na(+)-K(+)-ATPase activity by NO is dependent on the increase in intracellular cyclic GMP concentration. 2. The effect of NO donors, sodium-nitroprusside (SNP) and S-nitroso-glutathione (S-NO-Glu), and a permeable cyclic GMP analogue, 8-bromo-cyclic GMP, on Na(+)-K(+)-ATPase activity (measured as ouabain-sensitive 86Rb-uptake) was studied in human cultured corpus cavernosum smooth muscle cells (HCCSMC). In addition, the effect of the cyclic GMP lowering agent, methylene blue, on NO-induced increase in Na(+)-K(+)-ATPase activity was studied. 3. SNP (1 microM) caused time-dependent increases in ouabain-sensitive Rb-uptake (33-72%) over 2-20 min in HCCSMC. The stimulation of ouabain-sensitive Rb-uptake by SNP was concentration-dependent (30 and 102% with 0.1 and 1 microM SNP, respectively). Similarly, significant increases in ouabain-sensitive Rb-uptake were obtained with 1 and 10 microM S-NO-Glu. In contrast, incubation of HCCSMC with 8-bromo-cyclic GMP (100 microM) did not increase ouabain-sensitive Rb-uptake. 4. S-NO-Glu induced-increase in intracellular cyclic GMP synthesis, but not the increase in ouabain-sensitive Rb-uptake, was completely inhibited by methylene blue in HCCSMC. 5. The Na(+)-K(+)-ATPase inhibitor, ouabain, caused a concentration-dependent increase in tension (0.5 to 2 fold) in tissues contracted with 15 mM KCl. SNP and S-NO-Glu caused a concentration-dependent relaxation (concentration required to cause half maximal relaxation (ED50) = 0.04 and 0.2 microM, respectively) of HCC strips contracted with 15 mM K+. Ouabain (0.1 to 10 microM) inhibited the response to SNP and S-NO-Glu by shifting the concentration-response curves to the right and preventing full smooth muscle relaxation.6. These results indicate that the activity of Na+-K+-ATPase modulates the contractility of HCC smooth muscle, and that NO stimulates Na+-K+-ATPase activity in HCCSMC independently of its ability to increase the intracellular cyclic GMP concentration. They also suggest that stimulation of Na+-K+-ATPase activity plays an important role in NO-induced relaxation of HCC smooth muscle

Influence of the vascular endothelium on angiotensin II-induced contractions in rabbit renal artery.

The influence of vascular endothelium on angiotensin II-induced contraction and the underlying mechanisms in the rabbit renal artery were investigated. In endothelium-intact preparations, angiotensin II (3-100 nM) caused a concentration-dependent increase in tension by maximally (Emax) 0.74 +/- 0.05 g. Removal of the endothelium significantly enhanced the angiotensin II-induced contractions (Emax: 3.91 +/- 0.19 g). Indomethacin (10 microM) did not influence the angiotensin II-induced contractions. Methylene blue (10 microM) and NG-methyl-l-arginine (L-NMMA, 5 microM) significantly enhanced angiotensin II-induced contractions by 418 +/- 29% and 200 +/- 14%, respectively, in endothelium intact preparations, but not in those devoid of endothelium. L-arginine (1 mM), but not D-arginine, reversed the L-NMMA-induced enhancement of the angiotensin II-induced contraction. The present results suggest that angiotensin II-induced contractions in rabbit renal artery are largely subject to the influence of the endothelium. The endothelium-derived relaxant factor (EDRF), rather than cyclo-oxygenase products, appears to be involved in mediating the inhibitory effects of the endothelium. Nitric oxide (NO) derived from endothelium may play a major role in inhibiting angiotensin II-induced contractions in this preparation.

Modulation of contractions to and receptors for endothelins in canine veins.

Experiments were designed to characterize endothelin receptors in canine femoral veins and to determine whether their distribution or sensitivity could be altered by chronic changes in blood flow and oxygen tension in veins proximal to an arteriovenous fistula. Endothelium was removed from unoperated or fistula-operated femoral veins of anesthetized dogs. Veins were cut into rings and suspended in organ chambers for the measurement of isometric force or frozen for isolation of membrane proteins. Endothelin-1, endothelin-3, and sarafotoxin S6c caused concentration-dependent increases in tension in all rings. In rings of unoperated veins, maximal tensions were significantly less to endothelin-3 and sarafotoxin S6c than to endothelin-1. In rings of fistula-operated veins, maximal tensions to endothelin-1 and endothelin-3 were the same. Contractions to endothelin-1 or endothelin-3 in unoperated veins were not inhibited by an antagonist of endothelin-A receptors, BQ-123. Binding of 125I-labeled endothelin-1 (125I-endothelin-1) to membranes from veins without endothelium increased as a function of membrane protein. Affinity of receptors, as determined by competitive inhibition of 125I-endothelin-1 was as follows: endothelin-1 > endothelin-3 > sarafotoxin S6c. Competitive inhibition of 125I-endothelin-1 by endothelin-3 and sarafotoxin S6c was significant for a two-site binding model in all veins. The total number of binding sites was reduced significantly in fistula-operated veins; the relative proportions of high- and low-affinity binding sites did not change. Affinity of high- and low-affinity receptors increased in fistula-operated veins.(ABSTRACT TRUNCATED AT 250 WORDS)

Acute intraoperative arterial elongation: histologic, morphologic, and vascular reactivity studies.

This study focuses on the histomorphologic damage produced by an acute elongation process, as well as on quantifying the alterations in arterial contractility following the application of this technique. Light microscopy and scanning electron microscopy studies were prepared from expanded and non-expanded pig superficial femoral arteries (SFA) harvested immediately following expansion, and again at 24- and 72-hr intervals. Histologically, the expanded vessels showed minor, patchy, endothelial slough, but not fragmentation of the internal or external elastic lamina. At 24 hr, the endothelium showed reactive changes, but no evidence of smooth-muscle necrosis of the tunica media was observed. At 72 hr, healing of the endothelium was evident by SEM. Similar specimens, also from the SFA, were harvested and placed in organ chambers immediately following expansion and 24 hr later, to measure contractility when exposed to alpha-adrenergic agonists. The vessels were exposed to the contractile agonists, phenylephrine and 5-hydroxytryptamine, which evoked similar concentration-dependent increases in tension in both the expanded group and the controls. From these observations, the authors conclude that acute intraoperative elongation of arteries results in only minor endothelial damage, without affecting the inherent contractility of the vessel wall.

Autonomic modulation of contractions to endothelin-1 in canine coronary arteries.

Endothelin-1 contracts vascular smooth muscle and inhibits release of neurotransmitter from adrenergic and cholinergic neurons. Experiments were designed to investigate the interaction of these mechanisms in a blood vessel that receives both adrenergic and cholinergic innervation. Rings cut from canine left anterior descending coronary arteries were suspended in organ chambers for the measurement of isometric force. In some rings, the endothelium was removed. Endothelin-1 caused concentration-dependent increases in tension in all rings. During electrical stimulation (1 Hz, 9 V, 2 msec), the contractions to endothelin-1 were reduced significantly. In rings without endothelium, this decrease was greater in the presence of atropine (10(-6) M) and was eliminated by a combination of phentolamine (10(-5) M) and propranolol (5 x 10(-6) M). Contractions to endothelin-1 during electrical stimulation in rings with endothelium were significantly less than those without endothelium. This difference was eliminated by atropine and NG-monomethyl L-arginine (10(-4) M). The presynaptic effects of endothelin-1 were studied by measurement of tritium-labeled norepinephrine. Phasic electrical stimulation induced release of norepinephrine; this was inhibited by endothelin-1 at high concentrations (4 x 10(-7) M) in the presence of atropine. These results suggest that the major effect of endothelin-1 is postsynaptic in canine coronary arteries. However, contractions to endothelin-1 may be modulated by the level of sympathetic and parasympathetic tone. In situations in which innervation to the coronary arteries is altered, for example, in hearts used for transplantation, the contractile effects of endothelin-1 would prevail.