Role of endothelium in the responses of human intracranial arteries to a slight reduction of extracellular magnesium.

The effect of a slight reduction of extracellular magnesium on the tone of precontracted small human pial arteries and the dependence of the response on the functional integrity of the vascular endothelium was investigated in vitro. A decrease in extracellular magnesium from 1.2 to 0.8 mM resulted in sustained relaxation when the endothelium was intact. When the endothelium was disrupted, the same reduction resulted in an elevation of tone. These results suggest that small changes in extracellular magnesium modulate human cerebro-arterial tone through an endothelium-derived relaxing factor rather than by altering smooth muscle tone directly.     

Effects of nitric oxide donors and inhibitors of nitric oxide signalling on endothelin‐ and serotonin‐induced contractions in human placental arteries

In order to explore the role of nitric oxide (NO) in the control of fetoplacental vascular tone in normal pregnancy we have examined the effects of NO donors on uteroplacental arteries pre‐contracted with the vasoconstrictor endothelin‐1 (ET‐1) or serotonin (5‐HT). We have furthermore examined the effects of guanylate cyclase inhibitors on the NO‐induced relaxation. Segments of placental arteries (n=102) obtained from 39 placentas immediately after delivery were mounted in organ baths and superfused with Krebs–Ringer solution at 37 °C. The vessel segments were exposed to drugs for various intervals and the tension was recorded isometrically and registered on a polygraph. Cyclic guanosine monophosphate (cGMP) analysis was performed after extraction of vessel segments using a specific radioimmunoassay. The placental artery segments responded to ET‐1 and 5‐HT with a dose‐dependent vasoconstriction. After pre‐contraction with ET‐1 (10^–7 M ) or 5‐HT (10^–6 M ), the vessels relaxed in response to the NO donors glyceryltrinitrate (GTN) (10^–6 M ) and S‐nitroso‐N‐acetyl‐penicillamine (SNAP) (10^–5 M ). In the presence of the non‐specific guanylate cyclase inhibitor LY 83583 (10^–6 M ), the vessels responded with a small contraction. In the presence of the soluble guanylate cyclase (sGC) inhibitor 1H[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one (ODQ) the non‐treated vessels responded with a relaxation. 1H[1,2,4]oxadiazolo[4,3‐a]quinoxalin‐1‐one gave no obvious relaxation in pre‐contracted vessels. Addition of 8‐Br‐cGMP, the cell‐permeant analogue of cGMP, with or without pre‐contraction had no effect on the vessels. Cyclic guanosine monophosphate analysis showed that GTN treatment caused an increase in cGMP after 12 min. Our results indicate that NO acts as a vasodilator in placental vessels. The cGMP‐dependent mechanisms may be involved in NO‐induced relaxation but cGMP‐independent mechanisms appear also to be involved.     

Effects of nitric oxide donors and inhibitors of nitric oxide signalling on endothelin- and serotonin-induced contractions in human placental arteries

In order to explore the role of nitric oxide (NO) in the control of fetoplacental vascular tone in normal pregnancy we have examined the effects of NO donors on uteroplacental arteries pre-contracted with the vasoconstrictor endothelin-1 (ET-1) or serotonin (5-HT). We have furthermore examined the effects of guanylate cyclase inhibitors on the NO-induced relaxation. Segments of placental arteries (n=102) obtained from 39 placentas immediately after delivery were mounted in organ baths and superfused with Krebs-Ringer solution at 37 degrees C. The vessel segments were exposed to drugs for various intervals and the tension was recorded isometrically and registered on a polygraph. Cyclic guanosine monophosphate (cGMP) analysis was performed after extraction of vessel segments using a specific radioimmunoassay. The placental artery segments responded to ET-1 and 5-HT with a dose-dependent vasoconstriction. After pre-contraction with ET-1 (10(-7) M) or 5-HT (10(-6) M), the vessels relaxed in response to the NO donors glyceryltrinitrate (GTN) (10(-6) M) and S-nitroso-N-acetyl-penicillamine (SNAP) (10(-5) M). In the presence of the non-specific guanylate cyclase inhibitor LY 83583 (10(-6) M), the vessels responded with a small contraction. In the presence of the soluble guanylate cyclase (sGC) inhibitor 1H[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (ODQ) the non-treated vessels responded with a relaxation. 1H[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one gave no obvious relaxation in pre-contracted vessels. Addition of 8-Br-cGMP, the cell-permeant analogue of cGMP, with or without pre-contraction had no effect on the vessels. Cyclic guanosine monophosphate analysis showed that GTN treatment caused an increase in cGMP after 12 min. Our results indicate that NO acts as a vasodilator in placental vessels. The cGMP-dependent mechanisms may be involved in NO-induced relaxation but cGMP-independent mechanisms appear also to be involved.     

Influence of endothelium in the in vitro vasorelaxant effect of magnesium on aortic basal tension in DOCA-salt hypertensive rat.

The objective of this study was to examine the influence of the endothelium on the extracellular magnesium induced relaxation of basal tension in isolated aortas from both mineralocorticoid-salt (DOCA-salt) hypertensive and control normotensive Sprague Dawley male rats. After incubation in magnesium-free physiological salt solution (PSS) (O mM magnesium), the increase of extracellular magnesium (1.2; 4.8 mM magnesium) caused a decrease in aortic tone which was significantly greater when endothelium was disrupted. Magnesium-induced relaxation was also more pronounced when endothelial NO production was blocked by 10(-4) M N omega-nitro-L arginine methyl ester (L-NAME). It is suggested that the vasorelaxation induced by extracellular magnesium is linked to the level of aortic basal tension developed in magnesium-free PSS. The endothelium does not seem to be directly implicated in magnesium-induced vasorelaxation in aortas from normotensive rats. However, in DOCA-salt hypertensive rats, the magnesium-induced relaxation of basal tension was less in the intact aorta (though not when the endothelium was disrupted) when the cyclo-oxygenase pathway was blocked by 10(-6) M indomethacin. These data therefore suggest that extracellular magnesium can promote relaxation by endothelium-dependent and cyclo-oxygenase-dependent mechanisms such as the production of relaxing prostacyclin in isolated aorta from DOCA-salt hypertensive rats.     

Active myogenic tone: a requisite for hemoglobin mediated vascular contraction?

Acellular free hemoglobin (Hb), when intravenously administered to animals and humans, elicits vascular contraction. A primary mechanism for the Hb mediated vasoconstriction is Hb scavenging of nitric oxide (NO), a potent relaxation factor, constitutively secreted by the vascular endothelium. However, in the isolated rat thoracic aorta in basal state, Hb does not elicit contraction. To investigate this apparent paradox, we assessed isolated rat aortic ring isometric contraction responses to Hb under different myogenic tone states: (1) following equilibration at a submaximal tension, (2) following agonist induced contraction, or (3) following a passive mechanical stretch. In vessel rings at basal state, Hb as high as 4 microM did not elicit any measurable contractions. In contrast, in vessel rings tone enhanced with norepinephrine, Hb as low as 0.1 microM Hb elicited a significant additional contraction. In vessel rings with passively induced tone, 4 microM Hb did not elicit a notable contraction. Similarly, in vessel rings in basal state, 0.17-1 mM acetylcholine, a NO dependent vasodilator, did not elicit relaxation. In these vessel rings, exogenous 8-Br-cGMP, a membrane permeable cGMP analog, did not elicit relaxation. In conclusion, in the isolated rat thoracic aorta, Hb mediated contraction may be contingent upon the state of myogenic tone.     

Effects of reduced oxygen tension on endothelium-dependent relaxation induced by acetylcholine differ in rabbit femoral artery and jugular vein

In intact rabbit femoral artery rings pre-contracted with phenylephrine, acetylcholine (ACh; 10(-9)-10(-6) M) produces endothelium-dependent relaxation, abolished after mechanical rubbing to remove the endothelium. The response to ACh was absent at low oxygen tension (less than or equal to 4 kPa) or in the presence of sodium cyanide (1 mM). Intact rabbit jugular veins relaxed to ACh in lower concentration than did the femoral artery, 10(-10)-10(-8) M; at sufficient ACh concentration the relaxation was complete. In veins with completely removed endothelium no relaxation to ACh occurred, and at concentrations above 3 x 10(-7) M the response was a contraction. The relaxation response to ACh of intact veins persisted during contraction at lower oxygen tension or in the presence of 1 mM sodium cyanide. In rubbed veins, cyanide consistently induced a transient contraction, which was absent in intact veins. The study demonstrates pronounced endothelium-dependent relaxation to ACh in a venous preparation, with a markedly lower sensitivity of the relaxation response to hypoxia than in a muscular artery of the same species.     

The role of the endothelium in the control of venous tone: studies on isolated human veins

The role of the endothelium in the vasomotor control of veins was investigated in 14 isolated ring preparations of presumably normal saphenous veins obtained from vein grafts in connection with vascular surgery. The investigations were performed with the specimens mounted for recording isometric tension in organ baths. Paired rings were used, one normal and the other de-endothelialized by gentle rubbing. The responses to noradrenaline (10(-8)-2 X 10(-5) M), acetylcholine (10(-6) M) and 120 mM KCl solution were tested. After precontraction with 10(-7) M noradrenaline, acetylcholine at 10(-6) M did not induce cholinergic relaxation, but in the majority of experiments induced a further increase in tone. Maximal contraction with noradrenaline was significantly higher in normal compared to de-endothelialized vessels. Therefore, endothelium-derived relaxation as in arteries did not occur in human saphenous veins and the existence of an endothelial-derived contracting factor in response to acetylcholine and noradrenaline is a possibility.     

Arterial size determines the enhancement of contractile responses after suppression of endothelium-derived relaxing factor formation

We studied the effect of endothelium-derived relaxing factor (EDRF) on norepinephrine-induced contractile responses and on the tissue guanosine-3,5-phosphate (cGMP) concentration of isolated rabbit arteries with an increasing endothelium to smooth muscle cell ratio (aorta, femoral and mesenteric arteries). After suppression of EDRF formation (either by N ^G-nitro-l-arginine or, in mesenteric arteries, by saponin), contractions elicited by cumulative doses of norepinephrine were unaltered in aorta but were enhanced by 22.5% in femoral arteries and by 44.3% in mesenteric arteries (at the highest norepinephrine concentration). The cGMP concentration (pmol/mg protein) of unstimulated, endotheliumintact vessels decreased after suppression of EDRF formation from 1.09±0.24 to 0.74±0.28 in aortic, from 2.86±0.4 to 0.61±0.19 in femoral and from 6.3±0.9 to 0.7±0.15 in mesenteric arterial segments. The basal cGMP concentration did not differ in endothelium-denuded segments of these arteries, suggesting a similar basal activity of soluble guanylate cyclase (sGC). A higher sensitivity of sGC may have contributed to the higher cGMP concentration observed in the smaller arteries, since in the presence of sodium nitroprusside the cGMP concentration of endothelium-denuded segments increased 1.8-fold in aortic, 2.9-fold in femoral and 2.4 fold in mesenteric arterial segments. However, these differences in sGC activation cannot be solely responsible for the high basal cGMP concentration in endotheliumintact mesenteric arteries. The greater ratio of endothelium to smooth muscle cell layers in the smaller arteries might result in a higher EDRF concentration in the vascular wall and subsequently in a higher cGMP concentration. In conclusion, these data support the view of a greater importance of EDRF-mediated vascular control in small arteries than in large conduit arteries.     

Hypercapnic vasodilatation in isolated rat basilar arteries is exerted via low pH and does not involve nitric oxide synthase stimulation or cyclic GMP production

The relaxant effect of hypercapnia (15% CO₂) was studied in isolated circular segments of rat basilar arteries with intact endothelium. The nitric oxide synthase inhibitor nitro-l -arginine (l -NOARG) and the cytosolic guanylate cyclase inhibitor methylene blue (MB), significantly reduced this relaxation by 54% and 70%, respectively. The effect of l -NOARG was completely reversed by l -arginine. Blockade of nerve excitation with tetrodotoxin (TTX) had no affect on the 15% CO₂ elicited vasodilatation. Measurements of cGMP in vessel segments showed no significant increase in cGMP content in response to hypercapnia. l -NOARG and MB, but not TTX, significantly reduced the basal cGMP content in cerebral vessels. Adding 1.5% halothane to the incubation medium did not result in a significant increase in cGMP content. Lowering the pH by cumulative application of 0.12 m HCl resulted in relaxation identical to that obtained by lowering the pH with 15% CO₂. In vessel segments in which the endothelium had been removed beforehand 15% CO₂ induced relaxation that was not different from that seen in vessels with intact endothelium. l -NOARG had no affect in endothelium denuded vessels. The results suggest that high CO₂ elicits vasodilatation of isolated rat basilar arteries by a mechanism independent of nitric oxide synthase (NOS) activity. The markedly reduced basal cGMP levels in cerebral vessels by l -NOARG and MB suggest that there exists a basal NO formation in the cerebral vessel wall.     

Smooth muscle dilatation in the human uterine artery induced by substance P, vasoactive intestinal polypeptide, calcitonin gene-related peptide and atrial natriuretic peptide: relation to endothelium-derived relaxing substances.

The relaxatory influences of substance P (SP), vasoactive intestinal polypeptide (VIP), calcitonin gene-related peptide (CGRP) and atrial natriuretic peptide (ANP) were investigated in human uterine arteries precontracted by noradrenaline in vitro. SP, VIP, CGRP and ANP all relaxed isolated uterine arteries with intact endothelium. When tested on vessels devoid of their endothelium VIP and SP had no effect on smooth muscular tone, while ANP and CGRP still induced unchanged vasodilatation. These results suggest an involvement of an endothelium-derived relaxing substance in the mechanisms by which VIP and SP induce relaxation of the isolated human uterine artery. On the other hand, ANP and CGRP seem to act on the same vessel preparation in vitro independently of the vascular endothelium. Both addition of noradrenaline and exchange of sodium against potassium in the organ chambers resulted in smooth muscle contraction irrespective of the integrity of the endothelium.     

Relaxation induced by KCl,NaCl and sucrose in rabbit coronary arteries

Contractions induced in isolated ring segments of rabbit coronary arteries by 2-(2-aminoethyl)-pyridine (AEP) were transiently relaxed when KCl, NaCl or sucrose were added to the bathing solution without osmotic correction. When these solutes were added with osmolarity changes minimized by reducing the concentration of another constituent of the medium the relaxations were reduced or abolished. AEP contraction was associated with depolarization and solute-induced relaxation with a lessening of the depolarization. Ouabain depolarized the vessel and diminished both the relaxation and repolarization induced by hyperosmolar addition of NaCl and sucrose to AEP-contracted segments. Sucrose and NaCl also relaxed potassium-induced contractions and their effect was greater at [K]_o 25 mM than at [K]_o 60 mM. Hyperosmolar relaxation still occurred but was significantly smaller in vessels subjected to prolonged sodium pump inhibition produced by cold storage followed by re-warming in K-free or ouabaincontaining solution. It is concluded that hyperosmolar relaxation is mainly due to hyperpolarization and is influenced by the level of membrane potential. The inhibition of hyperosmolar relaxation by ouabain may be due to its depolarizing action.Supported by DHHS grant HL 26238. The study was performed during the tenure by K. D. Keef of a Fellowship of the American Heart Association-Greater Los Angeles Affiliate     

Clonidine induced endothelium-dependent tonic contraction in circular muscle of the rat hepatic portal vein.

Clonidine, an alpha2-agonist, has been shown to be useful in the treatment of hepatic portal hypertension in cirrhosis. The mechanism has been attributed to a clonidine-induced decrease in sympathetic activity. While clonidine has been shown to stimulate the alpha2-adrenoceptors of blood vessels, there is limited knowledge of the effects of clonidine on the circular muscle of the hepatic portal vein which regulates its blood flow. To investigate clonidine-induced contraction of the circular muscle of the hepatic portal vein and to clarify the possible role of the endothelium in the contraction, we examined the effects of clonidine on the isometric contraction of endothelium-intact and -removed ring preparations of the rat hepatic portal vein. In endothelium-intact preparations, clonidine caused a concentration-dependent increase in the amplitude of contractions. Inhibition of NO synthesis with Nomega-nitro-L-arginine (L-NNA) elevated the resting tone, and increased the amplitude of the clonidine-induced contractions. Inhibition of cyclooxygenase by diclofenac did not change the amplitude of the clonidine-induced contractions observed both in the presence and absence of L-NNA. Application of a single concentration of clonidine induced a clear increase in amplitude of both twitch and tonic contractions. Twitch and tonic contractions induced by clonidine were inhibited by yohimbine. When the endothelium was damaged by sodium deoxycholate, tonic contractions induced by clonidine were completely suppressed, whereas the increase in twitch contractions was not influenced by chemical damage of the endothelium. Neither SKF-96365, a nonselective cation channel blocker, nor superoxide dismutase, a free radical scavenger, in the presence of catalase, changed the tonic contraction induced by clonidine. These results indicate that stimulation of alpha2-adrenoceptors enhanced twitch contractions and induced tonic contractions in the circular muscle of the rat hepatic portal vein, especially in the absence of NO. The latter, but not the former, occurs through an endothelium-dependent pathway.     

Reversal of alpha 1-receptor mediated relaxation in intestinal smooth muscle

The alpha 1-receptor agonist phenylephrine relaxed longitudinal rabbit jejunal muscle contracted in vitro by low concentrations of barium ions (1 mM). When the Ba2+ concentration was increased to 10-15 mM the response to phenylephrine was a contraction, and at Ba2+ concentrations in between the high and low range this response was biphasic--a relaxation followed by a contractile phase. The alpha 2-receptor agonist clonidine did not affect the tone of the Ba2+ contracted preparation. When the muscle preparation was contracted by Sr2+ (1-20 mM) in the presence of Ca2+ (2.5 mM), phenylephrine relaxed it, and no contractile response to phenylephrine was observed. In the absence of extracellular Ca2+, 5 mM Ba2+ caused a contraction. Under these conditions phenylephrine had no effect on the tissue tone. When Ca2+ was added in a low concentration (0.2-2 mM), phenylephrine elicited a gradually increasing contractile response. At 5 mM Ca2+ the contractile response was replaced by the normal relaxation. The contractile response to phenylephrine in the presence of 5 mM Ba2+ and 2.5 mM Ca2+ was partially blocked by low concentrations of verapamil. In higher concentrations verapamil abolished the tissue tonus completely. The contractile response to phenylephrine in the presence of 5 mM Ba2+ and 2.5 mM Ca2+ could be reverted to the normal relaxation by the addition of 20 mM Mg2+. Increasing the K+ concentration from the normal 5.9 to 62.9 mM blocked the phenylephrine-induced relaxation. No contractile response to phenylephrine occurred. It is concluded that Ba2+ could reverse the response of alpha 1 receptor stimulation in rabbit jejunum from a relaxation to a contraction and that this contractile response was dependent on the presence of Ca2+.     

尸体防腐固定灌注对血管口径的影响

用测微目镜测量了31侧大鼠股动、静脉的外径,比较其生前,死后及灌注防腐液后的变化、并对53侧胎儿的大隐静脉及股动、静脉灌注前后的血管外径进行了测量和比较。死后未经灌注的血管与活体的血管外径近似;而灌注后上述血管的口径均有明显增大。根据回归方程提供了胎儿灌注前微小动脉外径的估算数据。     

Human cytokines modulate arterial vascular tone via endothelial receptors

Only a few cytokines have been tested for their possible role in modulating vascular function. Moreover, no direct effect of cytokines on vascular tone has yet been thoroughly studied. We therefore examined whether a wide range of well-defined cytokines could directly affect vascular tone in isolated human arterial and venous segments from various organs. We found that the cytokines stem cell factor (maximal response with 1 mM), granulocyte colony-stimulating factor (0, 1 mM) and erythropoietin (1 mM) relaxed, while tumor necrosis factor alpha (0.1 mM), interleukin (IL) 6 (10 mM) and IL-10 (0.1 mM) induced contraction of arterial but not of venous segments. The cytokines (maximal concentration tested was 1 mM) IL-3, IL-5, IL-13, macrophage colony-stimulating factor and granulocyte-macrophage colony-stimulating factor had no apparent effects on either arterial or venous tone. These vascular effects were endothelium-dependent as denuded arteries did not respond to any cytokine, and inhibition of nitric oxide synthase or endothelin receptor A abrogated the cytokine-induced changes in vascular tone. With immunohistochemistry we found receptors for the active cytokines on the arterial endothelium. In conclusion, several cytokines may modulate arterial vascular tone via endothelium-dependent mechanisms. Therefore cytokines might significantly modify blood supply to inflamed or ischemic tissues with elevated local concentrations of cytokines.     

Modulation of serotonin-induced vasospasm by endothelium and monoamine oxidase

Endothelial modulation of flow induced by intraluminal serotonin (5-HT) in isolated and perfused bovine coronary artery segments was studied. A constant-pressure continuous perfusion apparatus was utilized. Control coronary arteries were perfused with a fixed volume of serotonin-containing solution followed by a serotonin-free solution, and flow-rate changes during onset and relaxation of vasospasm were measured. Both monoamine oxidase inhibition by iproniazide and endothelium disruption by collagenase increased the rate of onset and magnitude of vasospasm. When the endothelium was intact the vasospasm continued to increase, reaching maximum well after the end of the serotonin perfusion, followed by slow relaxation toward baseline. This contrasted with de-endothelialized vessels in which the increase in contractile response terminated abruptly at the end of the serotonin perfusion and returned rapidly to baseline. Coronary arteries stimulated with prostaglandin F2 alpha responded similarly to de-endothelialized vessels stimulated by 5-HT, although further de-endothelialization of F2 alpha-stimulated vessels showed increased rates of onset and relaxation of vasospasm, suggesting a physical barrier role for the endothelium towards unmetabolized agents. These observations are consistent with the hypothesis that endothelial cells are capable of taking up, storing and subsequently releasing serotonin. The results suggest a protective role of the endothelium as a metabolic and physical barrier. This may represent an anatomical substrate favouring the development of localized vasospasm at sites where the endothelium is injured.     

Endothelial mediated enhancement of noradrenaline induced vasoconstriction in normal and varicose veins

The role of the endothelium to influence smooth muscle contraction in normal and varicose veins was investigated in vitro using saphenous vein preparations obtained at vascular surgery. Paired control and endothelium-denuded rings were tested simultaneously and contracted with noradrenaline (10(-9)-10(-4) M). Identical experiments were also performed on sheep femoral veins and arteries. Noradrenaline induced maximal tension was 30% lower in varicose compared to normal veins. In normal veins removal of endothelium significantly reduced the maximal response to noradrenaline by 40% whereas in varicose veins no significant reduction could be seen. In the sheep femoral vein removal of the endothelium also resulted in decrease of noradrenaline-induced contraction. It can be concluded that in the human saphenous vein the endothelium has a contraction facilitating effect in response to stimulation with noradrenaline. In varicose veins the endothelial-mediated enhancement of noradrenaline-induced vasoconstriction is reduced probably because of endothelial damage. This observation may be of importance in the pathogenesis of varicose veins.     

Contraction-initiated NO-dependent lymphatic relaxation: a self-regulatory mechanism in rat thoracic duct

The objectives of this study were to evaluate the physiological importance of the flow and shear generated by phasic contractions of lymphatic vessels and the mechanisms responsible for the influences of such shear on lymphatic pumping. Lymphatic segments of the rat thoracic duct were isolated, cannulated and pressurized. The diastolic diameters were measured in phasically non-active segments. The diastolic and systolic diameters, half-relaxation time (HRT), contraction frequency, ejection fraction and fractional pump flow were determined in phasically active segments. Since imposed flow was excluded, flow and shear occurred only as a result of the intrinsic contractions in phasically active segments whereas in phasically non-active segments contraction-generated flow and shear were absent. The influences of incrementally increased transmural pressure (from 1 to 5 cmH₂O) were examined in control conditions and after NO synthase blockade ( l -NAME 10⁻⁴ m ) or cyclooxygenase blockade (indomethacin 10⁻⁵ m ). The spontaneous phasic contractions produced a flow-dependent diastolic relaxation. This reduction of the lymphatic tone is a regulatory mechanism that maintains pumping in thoracic duct in an energy-saving/efficient mode: it improves diastolic filling (enhanced lusitropy – lowering HRT), makes lymphatic contractions stronger (enhanced inotropy – higher contraction amplitude) and propels more fluid forward during each contraction (elevated ejection fraction) while decreasing contraction frequency (reduced chronotropy). The findings also demonstrated that the NO pathway, not the cyclooxygenase pathway is responsible for this reduction of lymphatic tone and is the prevailing pathway responsible for the self-regulatory adjustment of thoracic duct pumping to changes in lymph flow pattern.     

Alpha adrenergic activation and hemoglobin mediated contraction in the isolated rat thoracic aorta.

A primary mechanism for Hb mediated vascular contraction appears to be Hb scavenging of endothelium derive nitric oxide (NO), a potent vasodilator. In isolated rat thoracic aorta, however, Hb elicits contraction only after precontraction. The present study investigated a possible role of the alpha adrenergic activation in the Hb mediated contraction. Thoracic aortic rings harvested from normal male SD rats were prepared in a tissue bath and isometric tension changes were evaluated. In vessel rings precontracted with 50nM norepinephrine (NE), 1 microM Hb produced an additional 21.8+/-13.2% increase in tension. Pretreatment with 70nM phentolamine, an alpha adrenergic antagonist, prevented the 50nM NE induced contraction. In these vessels, subsequent treatment with 2-4 microM Hb did not elicit contraction. In vessel rings precontracted with 37mM KCl, 2 microM Hb produced an additional 21.8+/-20.1% tension increase (P<0.05). Pretreatment with phentolamine did neither prevent KCl induced contraction nor affect subsequent Hb mediated additional contraction. To test whether there is a threshold level of basal tension for Hb to trigger contraction, a group of vessel rings were passively stretched to match the tension generated by NE before Hb treatment. In these passively stretched vessel rings, Hb did not produce a significant contraction. Pretreatment with 10mM EGTA, a Ca++ chelator, significantly reduced NE induced contraction (9.7+/-5.9 vs 137.7+/-60.0%, P<0.01) but did not prevent it. EGTA also significantly reduced 2 microM Hb induced contraction (27.2+/-29.3% vs 8.9+/-7.7%, P<0.05). In contrast, pretreatment with verapamil, a Ca++ channel blocker, did not completely block NE and Hb induced contractions. In conclusion, alpha adrenergic activation is not a requisite for the Hb mediated contraction in isolated rat aortic rings. The mechanism how prior tone enhancement allows Hb mediated contraction remains unclear but results from this study suggest a factor that controls cytosolic Ca++ levels may be involved.     

Effects of bradykinin and leukotrienes B4 and D4 on the bovine brochial arteryin vitro: Role of the endothelium

The effects of bradykinin, leukotriene (LT) B₄ and LTD₄ on the bovine bronchial artery were studied using isolated, perfused vessel segments. Intrapulmonary bronchial arteries were precontracted to the EC₅₀ of 5-hydroxytryptamine in the organ bath, after which increasing concentrations of bradykinin, LTB₄ or LTD₄ were perfused through the vessel lumen. In arteries with intact endothelium, bradykinin induced a concentration-dependent relaxation beginning at 1 pmol/l (p<0.01), whereas LTD₄ and LTB₄ had no significant effect. Indomethacin (10 mol/l) significantly (p<0.01) inhibited bradykinin-induced endothelium-denuded arteries bradykinin had no observed effect, whereas LTB₄ and LTD₄ significantly increased tone with threshold concentrations of 50 pmol/l (p<0.025) and 5 pmol/l (p<0.05), respectively. The data indicate that in the bronchial artery: 1) bradykinin is a potent vasodilator; 2) bradykinin-induced relaxation is endothelium-dependent and only partially blocked by a cyclooxygenase inhibitor; 3) removal of the endothelium unmasks a potent contractile effect of LTB₄ and LTD₄.