Selective hemoglobin inhibition of endothelium-dependent vasodilation of rabbit basilar artery

The effect of hemoglobin on endothelium-dependent vasodilation of the isolated rabbit basilar artery was examined using an isometric tension recording method. Acetylcholine (ACh) (10(-7) - (10(-4) M) evoked a dose-dependent vasodilation of isolated rabbit basilar artery previously contracted by 10(-6) M serotonin. This vasodilating action disappeared after removal of the endothelium. The ACh-induced vasodilation of rabbit basilar artery is thought to be strictly endothelium-dependent. Hemoglobin (10(-7) - 10(-5) M) inhibited this ACh-induced endothelium-dependent vasodilation conditional upon the dose. Adenosine triphosphate (ATP, 10(-7) - 10(-4) M) also relaxed isolated rabbit basilar artery already contracted by 10(-6) M serotonin. This vasodilating action was slightly inhibited by adenosine antagonist, 8-phenyltheophylline (8-PT), and markedly attenuated by removal of the endothelium. This ATP-induced vasodilation is thought to be composed of ATP itself (endothelium-dependent) and ATP degradation products (endothelium-independent) such as adenosine monophosphate or adenosine. Hemoglobin markedly inhibited ATP-induced vasodilation, but there still remained a small vasodilation, which was blocked by 8-PT. Papaverine-induced vasodilation was not affected by removal of the endothelium, and hemoglobin did not inhibit the papaverine-induced vasodilation. These results suggest that rabbit basilar artery has endothelium-dependent vasodilating mechanisms induced by ACh and ATP, and that hemoglobin selectively blocks the endothelium-dependent vasodilation. This finding may relate to the pathogenesis of cerebral vasospasm after subarachnoid hemorrhage: there is a possibility that the presence of hemoglobin released from lysed erythrocytes inhibits the endothelium-dependent vasodilation of cerebral arteries; furthermore, the endothelial degeneration following subarachnoid hemorrhage may impair the vasodilating mechanisms of cerebral artery smooth-muscle cells.     

The effect of nicardipine on vasospasm in rabbit basilar artery after subarachnoid hemorrhage

This study was performed to examine the effect of the dihydropyridine calcium antagonist, nicardipine, on vasospasm after experimental subarachnoid hemorrhage (SAH) in the rabbit. The study was carried out in two parts: 1) effect of intravenous nicardipine (n = 45) and 2) effect of intracisternal nicardipine (n = 21). SAH was induced by injecting 5 ml of autologous arterial blood into the cisterna magna. In the intravenous study, there were five groups: 1) SAH without treatment; 2) SAH with vehicle (saline); 3) SAH and intravenous infusion of low-dose nicardipine (0.01 mg/kg/h); 4) SAH and intravenous infusion of high-dose nicardipine (0.15 mg/kg/h); and 5) controls without SAH. The intravenous infusions were started immediately after SAH and continued for 48 hours until death. In the intracisternal study, there were three groups: 1) SAH without treatment; 2) SAH with intracisternal administration of nicardipine (0.37 mg/h); and 3) controls without SAH. Intracisternal infusions were begun 70 hours after SAH and continued for 2 hours until death. After perfusion-fixation, the basilar artery was removed and processed for morphometric analysis. In the intravenous study, vessels from animals subjected to SAH were significantly narrowed when compared with controls, although after high-dose nicardipine vessel caliber was slightly larger than in the other SAH groups. Animals given intracisternal nicardipine showed a nonsignificant reduction of caliber as compared with controls: only 12% decrease in diameter and 22% decrease in luminal area. In the rabbit SAH model, nicardipine had a very modest effect on vasospasm at the doses tested.     

Hemoglobin penetration in the wall of the rabbit basilar artery after subarachnoid hemorrhage and intracisternal hemoglobin injection

Summary The ability of hemoglobin (Hb) to penetrate the basilar arterial wallin vivo after experimental subarachnoid hemorrhage was examined using immunohistochemistry. The distribution of anti-Hb antibodies in rabbit basilar artery was studied following the injection of autologous blood in the cisterna magna. Vessels removed two or four days after subarachnoid hemorrhage exhibited varying degrees of vasospasm, and exhibited Hb immuno-fluorescence throughout the vessel wall. Hemoglobin immunofluorescence was most conspicuous in the adventitia but was also seen in the smooth muscle and endothelial cell layers in 7 of 10 animals. The degree of vasoconstriction correlated with the total amount of Hb-fluorescence present in the vessel wall. When Hb solution alone was injected into the subarachnoid space, vasoconstriction was evident but penetration into the vascular layers was not as extensive as that observed after injection of autologous blood.These findings demonstrate that Hb is able to penetrate through the arterial wall after subarachnoid hemorrhage. The results provide direct support for the hypothesis that Hb-induced changes in smooth muscle and/or endothelial function can contribute to the pathogenesis of vasospasm.     

Reactivity of rabbit basilar artery to alterations in extracellular potassium and calcium after subarachnoid hemorrhage

Hemolysis of periarterial clots after subarachnoid hemorrhage may liberate large quantities of K+ into the vicinity of cerebral blood vessels and possibly change their sensitivity to endogenous vasoactive agents. The current study examined the influence of subarachnoid hemorrhage on the sensitivity of rabbit basilar arterial segments to K+ and Ca++. An analysis of K+ and Ca++ dose-response curves demonstrated that incubated arterial segments isolated from animals with subarachnoid hemorrhage were substantially more sensitive to these cations than were corresponding controls. We speculate that chronically elevated K+ levels in areas of periarterial clot lysis or brain ischemia might initiate vascular smooth muscle depolarization and vasospasm. Our data provide additional rationale for the use of calcium channel blockers in preventing or treating vasospasm in cases of subarachnoid hemorrhage.     

Inhibition of endothelin-converting enzyme activity in the rabbit basilar artery

OBJECTIVE: Endothelin (ET)-1 may be involved in the regulation of cerebrovascular resistance under pathological conditions, most notably during the development of vasospasm after subarachnoid hemorrhage. Blocking ET-converting enzyme activity may be a promising approach to the prevention of cerebral vasospasm after subarachnoid hemorrhage. METHODS: In this study, the effects of several putative ET-converting enzyme inhibitors were investigated after intracisternal application in rabbits, to inhibit basilar artery contractions induced by big ET-1 (2 x 10(-6) mol/L). RESULTS: In the group pretreated with [D-Val22]big ET-1[16-38] (2 x 10(-5) mol/L) (n = 8), the angiographically measured diameter of the basilar artery changed from 0.63 +/- 0.12 mm to 0.66 +/- 0.12 mm. In the control group (n = 8), the diameter of the basilar artery decreased from 0.71 +/- 0.13 mm to 0.57 +/- 0.15 mm. These results corresponded to an increase in vessel diameter of 5 +/- 10% in the treatment group and a decrease in vessel diameter of 20 +/- 16% in the control group (P = 0.014). In the group pretreated with captopril (2 x 10(-4) mol/L) (n = 8), the angiographically measured diameter of the basilar artery changed from 0.64 +/- 0.11 mm to 0.71 +/- 0.10 mm. These results corresponded to an increase in vessel diameter of 14 +/- 19% in the treatment group, compared with a decrease in vessel diameter of 20 +/- 16% in the control group (P = 0.014). CONCLUSION: These results demonstrate that [D-Val22]big ET-1[16-38] and captopril act as highly potent ET-converting enzyme inhibitors, affecting big ET-1-induced contraction of the rabbit basilar artery.     

A comparison of the vasodilating effects of halothane and isoflurane on the isolated rabbit basilar artery with and without intact endothelium.

Although volatile anesthetics result in cerebral arterial dilation, the precise mechanisms underlying this effect are not known. In vitro tension recordings were used to study the vasodilating potencies of halothane and isoflurane in isolated cerebral vessels and to examine the possible role of the endothelium in modulating any effects observed. Cylindrical segments of the rabbit basilar artery and midline ear artery from the same animal were placed in a flow-through bath of 37 degrees C oxygenated (95% O2/5% CO2) physiologic salt solution and stretched to a resting tension of approximately 2,000 dynes. They were then constricted with 3.0 x 10(-2) M K+, 1.0 x 10(-3) M norepinephrine, or 5.0 x 10(-6) M serotonin and exposed to either halothane or isoflurane at concentrations of approximately 0.5, 1.0, 1.5, and 2.0 MAC in varied order for 15 min at each concentration. A 30-min period of perfusion with anesthetic-free, vasoconstrictor-containing perfusate separated successive exposures to an anesthetic. Vessels prepared in this fashion retained their responsiveness to both vasoconstrictors and volatile anesthetics for as long as 4 h. They also relaxed appropriately to acetylcholine, indicating that the endothelium was intact. Concentrations of volatile anesthetic in the tissue perfusate were directly measured using gas chromatography, and the relationship between bath concentrations (expressed as MAC fractions) and the degree of relaxation were determined. The data were analyzed by parallel line regression. Halothane was found to be a significantly more potent vasodilator of the isolated basilar artery than was isoflurane. For example, in K(+)-constricted vessels, the concentration of halothane needed to produce a 50% reduction in tension was 1.32 MAC, compared with 1.66 MAC for isoflurane. Comparable differences were found in the basilar artery in the presence of other constrictors. However, there was no significant difference between the two agents in their effects upon the ear artery. In a separate series of experiments, the endothelium of basilar artery segments was removed by drying. Removal was confirmed by observing a diminished dilator response to acetylcholine. These vessels were subsequently constricted with K+, and relaxation dose-response curves were obtained for both halothane and isoflurane. There were no differences in the dose-response curves for deendothelialized versus intact vessels, with halothane still the more potent relaxant after endothelial removal. These data demonstrate that halothane and isoflurane cause a dose-dependent relaxation of rabbit cerebral vessels, regardless of the vasoconstrictor used. Halothane was a more potent relaxant of the basilar artery when expressed on a MAC-fraction basis.(ABSTRACT TRUNCATED AT 400 WORDS)     

Mitogen-activated protein kinase mediation of hemolysate-induced contraction in rabbit basilar artery.

OBJECT: Mitogen-activated protein kinase (MAPK) is an important signaling factor in vascular proliferation and contraction, which are the two features of cerebral vasospasm that follow subarachnoid hemorrhage. The authors studied the possible involvement of MAPK in hemolysate-induced signal transduction and contraction in rabbit basilar artery (BA). METHODS: Isometric tension was used to record the contractile response of rabbit BA to hemolysate, and Western blots were obtained using antibodies for MAPK. The following results are reported. 1) Hemolysate produced a concentration-dependent contraction of rabbit BA; however, preincubation of arteries with the MAPK kinase (MEK) inhibitor PD-98059 markedly reduced this contraction. The administration of PD-98059 also relaxed, in a concentration-dependent fashion, the sustained contraction induced by 10% hemolysate. 2) The Janus tyrosine kinase 2 inhibitor AG-490, preincubated with arterial rings, reduced the contractile response to hemolysate but failed to relax the sustained contraction induced by this agent. The Src-tyrosine kinase inhibitor damnacanthal and the phosphatidylinositol 3-kinase inhibitor wortmannin failed to reduce hemolysate-induced contraction. 3) Hemolysate produced a time-dependent elevation of MAPK immunoreactivity as seen on Western blots of rabbit BA. The MAPK was enhanced 1 minute after hemolysate exposure and the effect reached maximum levels at 5 minutes. The immunoreactivity of MAPK decayed slowly over time, but the level of this kinase was still higher than the basal level, even at 2 hours after exposure to hemolysate. Preincubation of arteries with the MEK inhibitor PD-98059 abolished the effect of hemolysate on MAPK immunoreactivity. CONCLUSIONS: Hemolysate produced contraction of rabbit BA, possibly by activation of MAPK, and therefore MAPK inhibitors may be useful in the treatment of cerebral vasospasm.     

Immediate ultrastructural effects of endothelin-1 on rabbit basilar artery

Summary In this study intra-arterial Endothelin-1 was applied to rabbit basilar arteries and morphological findings were compared between two groups who were either perfusion fixed or immersion fixed. We planned to establish the quantitative dimension of the drug-induced morphological alterations, independent of the fixation technique's effect.There was an abundance of collagenous fibres deposited among the smooth muscle cells which was not observed in control arteries and after immersion fixation. These degenerative changes are similar to the finding following subarachnoid haemorrhage. The only fixation-related difference was the fact that lamina elastica interna was not corrugated in the perfusion fixation group.It is concluded that, the observed changes in the connective tissue of the arterial wall alter the passive elastic properties and so affect the degree of the response to the vasoactive messengers.     

Effects of subarachnoid hemorrhage on platelet-derived vasoconstriction of rabbit basilar artery

The effects of subarachnoid hemorrhage on platelet-derived vasoconstriction of the isolated rabbit basilar artery were examined using an isometric tension recording method. The subarachnoid hemorrhage was induced by injecting arterial blood in the cisterna magna. The following points were confirmed: (1) the maximal contraction produced by the platelets (10⁷/mL) treated with indomethacin or dazoxiben (thromboxane synthetase inhibitor) were suppressed (65% or 70% of the control); (2) the contraction of the arteries treated with ONO-3708 (thromboxane A₂ antagonist) or ketanserin was inhibited (73% or 8.4%), as was contraction after subarachnoid hemorrhage (67% or 14%); (3) platelet-induced contraction was potentiated after subarachnoid hemorrhage; and (4) serotonin-induced contraction was potentiated after subarachnoid hemorrhage. However, synthetic thromboxane A₂-induced contraction was not potentiated.

The present experiments suggest that both serotonin and thromboxane A₂ contribute to vasoconstrictions induced by the platelets, before and after subarachnoid hemorrhage. The platelet-derived contraction response is potentiated after subarachnoid hemorrhage and serotonin is responsible for the increased reactivity.     

Contraction and relaxation of rabbit basilar artery by thiopental

Tension measurements were made on rabbit basilar artery segments to test the direct effect of thiopental on serotonin (5HT)- and norepinephrine (NE)-induced contraction. At concentrations of 3 X 10(-5) M or greater, thiopental caused relaxation of NE-induced tension. Thiopental doses greater than 3 X 10(-4) M had a similar relaxing effect on 5HT-induced contraction. However, thiopental doses below 3 X 10(-4) M demonstrated an unexpected dose-dependent potentiation of the 5HT-induced contraction.     

Effect of subarachnoid hemorrhage on serotonin uptake and release in the rabbit basilar artery

This study analyzes the changes induced by subarachnoid hemorrhage (SAH) on the serotonin (5-hydroxytryptamine, 5-HT) uptake and release evoked in rabbit basilar arteries by tyramine. Rabbits were injected with 5 ml of autologous arterial blood into the cisterna magna to produce SAH. Tritium accumulation in basilar arteries was measured after 30 minutes of incubation with 10(-7) M [3H]5-HT and a subsequent 120-minute superfusion (1 ml/min) period. The uptake of 5-HT by arteries 1, 2, 3, and 7 days after SAH was found to be 109%, 69%, 57% (P less than 0.05), and 67% (P less than 0.05) (n = 4, 4, 9, and 6; P less than 0.05) of control (n = 13; 16.8 +/- 1.2 X 10(2) dpm/mg tissue), respectively. The neuronal (cocaine-sensitive) uptake of 5-HT in the arteries 3 days after SAH decreased to approximately 38% of control, whereas the extraneuronal (cocaine-insensitive) uptake of both groups had almost the same absolute value (n = 6 and 6; 4.4 +/- 0.4 and 4.8 +/- 0.4 X 10(2) dpm/mg). Autoradiographic study disclosed that dense clusters of silver grains in the adventitia were not observed after treatment with cocaine (3 X 10(-5) M), although a diffuse distribution of grains was present throughout the vascular wall. The labeled arteries were stimulated by superfusion of tyramine, which is known to replace amines in the sympathetic nerve ending. Tyramine (10(-6) and 10(-4) M)-induced 3H efflux was significantly potentiated by SAH (n = 6) and was suppressed by treatment with cocaine.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effect of intravenous lidoflazine on whole blood-induced basilar artery contraction. An in vivo study

Lidoflazine, a piperazine derivative of known selectivity for vascular smooth muscle, was evaluated as a possible agent for prophylaxis of cerebral vascular contraction induced by subarachnoid perfusion with whole blood. Previous studies from this laboratory have indicated its efficacy in preventing basilar artery contraction induced by serotonin. The animals treated with a subarachnoid perfusion of whole blood had a mean 30% reduction in vessel diameter over the control value. Groups that were treated with 0.5 mg/kg of lidoflazine and 1.0 mg/kg of lidoflazine and then perfused with whole blood in the subarachnoid space had reductions in control diameter of 2.8% and 6.8%, respectively. One group treated with 2.0 mg/kg of lidoflazine and then perfused with whole blood actually had an increase in diameter of 6.8% over the control value. Lidoflazine, when administered intravenously at a slow rate, will not adversely lower systemic blood pressure and can prevent the contraction of cerebral vessels when the stimulus for contraction is whole blood within the subarachnoid space.     

Subarachnoid hemorrhage inhibition of endothelium-derived relaxing factor in rabbit basilar artery

Vascular contractions in response to KCl and serotonin (5-hydroxytryptamine, 5-HT) in rabbit basilar artery were studied in vitro using an isometric tension-measurement technique. Hemoglobin (10(-5)M) markedly augmented contractions induced by 5-HT (10(-9) to 10(-6)M) and slightly augmented those induced by KCl (20 to 80 mM) in arteries with intact endothelium. On the other hand, the augmentation induced by hemoglobin was almost abolished in arteries that were chemically denuded of endothelial cells by pretreatment with saponin. Since hemoglobin is known to be a selective inhibitor of endothelium-derived relaxing factor (EDRF), it is possible that the augmentation of contraction by hemoglobin in endothelium-intact arteries was mediated via an inhibition of spontaneously released EDRF. The effect of subarachnoid hemorrhage (SAH) on spontaneously released EDRF was investigated by injecting 5 ml of blood into the cisterna magna and sacrificing the rabbits 2 days later. Arteries after SAH showed a significant reduction in hemoglobin-induced augmentation compared to that seen in control arteries with intact endothelium. This result suggests that spontaneously released EDRF is significantly reduced after SAH. It is concluded that EDRF is released spontaneously in the rabbit basilar artery and that inhibition of its release might be involved in pathogenesis of cerebral vasospasm.     

Study the effects of zonisamide on fine structure of rabbit basilar artery and hippocampus in rabbit subarachnoid hemorrhage model

Background

In this study, we investigated the effect of a novel antiepileptic drug, zonisamide (ZNS), on the basilar artery and hippocampus in a rabbit subarachnoid hemorrhage (SAH) model.

Methods

Three groups of New Zealand white rabbits were used: a sham (non-SAH) group, an SAH?+?saline group, and SAH?+?drug treatment group that received ZNS. In the treatment group, the subjects were given ZNS for 3 days after the SAH. Hippocampal sections were evaluated for neural tissue degeneration. Basilar artery lumen areas and arterial wall thickness were also measured in all groups.

Results

The mean luminal area of the SAH?+?ZNS was significantly greater than the SAH?+?saline group. In addition, the arterial wall thickness of SAH?+?ZNS group was significantly thinner than the SAH?+?saline group. The neuronal degeneration scores of the hippocampal CA1 regions in the SAH?+?ZNS group were significantly lower than the SAH?+?saline treatment animals.

Conclusions

These results indicate that ZNS has a vasodilatatory effect on the basilar artery and a neuronal protective effect in the CA1 region of the hippocampus in a rabbit SAH model.     

Effects of bupivacaine and calcium antagonists on the rat uterine artery

Bupivacaine is a commonly used local anesthetic in obstetrical practice, but since this compound also has a constrictor action on vascular smooth muscle it can be hazardous to the fetus. The aim of the present study was to analyze the effect of bupivacaine on the uterine vasculature using the rat uterine artery as a model. Small arterial segments were mounted in tissue chambers for isometric recording of vascular tension using a specially designed teflon-steel gauge. Bupivacaine induced marked vasoconstriction and this vasoconstriction was reduced considerably by two different Ca antagonists, verapamil and nifedipine. Verapamil (10(-5) mol.l-1) reduced bupivacaine-produced arterial contraction by a mean of 78% and nifedipine (2.9 x 10(-7) mol.l-1) reduced arterial contraction by a mean of 57%.     

Effects of ischemia and hemoglobin on vascular function in isolated rabbit aortas

The vascular endothelium is important in the modulation of vascular tone via production of endothelium-derived relaxing and contracting factors. The abdominal aortas of five groups of rabbits were subjected to varying lengths of ischemia (0, 1, 2, 3, or 4 hours), removed, sectioned into transverse rings, and placed in tissue baths containing Krebs' buffer at 37 degrees C and aerated with 95% O2/5% CO2. After equilibration the rings were tested for endothelium-dependent vasodilation with methacholine and nonendothelium-dependent vasodilation with nitroprusside. Endothelium-dependent relaxation as elicited by methacholine was impaired at 3 and 4 hours of ischemia but was not significantly different at 1 and 2 hours as compared to control, whereas endothelium-independent vasodilation remained normal throughout the different periods of ischemia. The addition of 1 x 10(-6) mol/L rabbit hemoglobin reduced the time needed to demonstrate significant impairment of endothelial function to 2 hours. Endothelium-independent vasodilation was not affected by hemoglobin. We conclude that hemoglobin exacerbates ischemia vascular dysfunction in the rabbit aorta.     

Hemorrhage-induced alterations of rabbit basilar artery reactivity and sensitivity to serotonin

Subarachnoid hemorrhage has a profound effect on cerebrovascular reactivity. The present study noted a progressive change in the sensitivity and reactivity of rabbit basilar artery to serotonin after experimentally induced hemorrhage. The basilar artery exhibited an initial diminished response to serotonin for periods up to 6 hours after hemorrhage, whereafter the vessel gradually became hyperresponsive. The hypersensitivity became maximal 36 hours after hemorrhage and then began to return to normal. Such early onset of serotonin hypersensitivity and reactivity after subarachnoid hemorrhage has not been previously reported. The level of tension developed, however, suggests that serotonin alone is unlikely to cause vasospasm. The strict differentiation of spasm into early and delayed components is questioned.