Biomechanical and phenotypic changes in the vasospastic canine basilar artery after subarachnoid hemorrhage.

Because it has been argued that active myogenic tone prolongs cerebral vasospasm for >2 wk after subarachnoid hemorrhage (SAH), we attempted to identify the mechanism that plays the main role in sustaining the prolonged cerebral vasospasm. We especially focused on the roles of biomechanical and phenotypic changes in the cerebral arteries in the mechanisms of prolonged vasospasm after SAH. We used the basilar arteries from a "two-hemorrhage" canine model to make serial measurements of maximal contraction capacity and arterial stiffness (papaverine-insensitive tone) until day 28. We also examined hematoxylin-eosin-stained vasospastic canine basilar arteries for histological changes and immunohistochemically examined them for expression of myosin heavy chain isoforms (SMemb, SM1, and SM2), which are markers of smooth muscle phenotypic changes. Changes in collagen concentration in canine basilar arteries were also measured. Angiographic cerebral vasospasm persisted until day 14 and then gradually diminished; artery diameter returned to the control diameters on day 28. Maximal contraction capacity decreased until day 21 and showed some recovery by day 28. Arterial stiffness, on the other hand, progressed until day 28. Histological examination revealed medial thickening and increased connective tissue until day 21 and a return to control findings by day 28. The increased connective tissue was not accompanied by changes in collagen concentration, suggesting a role of some other protein in the increase in connective tissue. Immunohistochemical studies with anti-SMemb, anti-SM1, and anti-SM2 antibodies showed enhanced expression of SMemb from day 7 to day 21 and disappearance of SM1 and SM2 on days 14 and 21. The changes in myosin heavy chain isoform expression returned to normal on day 28. The above results indicate that biomechanical and phenotypic changes may play a pivotal role in sustaining cerebral vasospasm for >2 wk after SAH, with minimal changes in active myogenic arterial tone.     

Reversal of delayed vasospasm by an inhibitor of the synthesis of 20-HETE

This study characterized the time course of changes in cerebral blood flow (CBF) and vascular diameter in a dual-hemorrhage model of subarachnoid hemorrhage (SAH) in rats and examined whether acute blockade of the synthesis of 20-hydroxyeicosatetraenoic acid (20-HETE) with N-(3-chloro-4-morpholin-4-yl)phenyl-N'-hydroxyimido formamide (TS-011) can reverse delayed vasospasm in this model. Rats received an intracisternal injection of blood (0.4 ml) on day 0 and a second injection 2 days later. CBF was sequentially measured using laser-Doppler flowmetry, and the diameters of the cerebral arteries were determined after filling the cerebral vasculature with a casting compound. CBF fell to 67% of control after the first intracisternal injection of blood but returned to a value near control 24 h later. CBF again fell to 63% of control after a second intracisternal injection of blood and remained 30% below control for 5 days. The fall in CBF after the second intracisternal injection of blood was associated with a sustained 30% reduction in the diameters of the middle cerebral, posterior communicating, and basilar arteries. Acute blockade of the synthesis of 20-HETE with TS-011 (0.1 mg/kg i.v.), 5 days after the second SAH, increased the diameters of the cerebral arteries, and CBF returned to control. These results indicate that the rats develop delayed vasospasm after induction of the dual-hemorrhage model of SAH and that blockade of the synthesis of 20-HETE fully reverses cerebral vasospasm in this model. They also implicate 20-HETE in the development and maintenance of delayed cerebral vasospasm.     

Soluble and Catalytically Active Endothelin Converting Enzyme-1 is Present in Cerebrospinal Fluid of Subarachnoid Hemorrhage Patients

Endothelin converting Enzyme-1 (ECE-1) is essential for the production of Endothelin-1 (ET-1), which is associated with vasospasm following subarachnoid hemorrhage (SAH). We have previously demonstrated the presence of a catalytically active soluble form of ECE-1 in the media of endothelial cells. We aimed to determine if this form of ECE-1 exists in vivo, in cerebrospinal fluid (CSF) of SAH patients. We examined CSF taken from SAH subjects for the presence of soluble ECE-1 using a bradykinin based quenched fluorescent substrate assay. We obtained further confirmation by characterizing the CSF mediated cleavage products of BigET-1 and BigET_18–34 (6 μg/ml) using mass spectrometry. The specificity of cleavage was confirmed using the ECE-1 inhibitor {"type":"entrez-protein","attrs":{"text":"CGS35066","term_id":"877962710","term_text":"CGS35066"}}CGS35066 5nmol/L. SAH CSF samples had mean ECE-1 activity of 0.127 ± 0.037 μmols of substrate cleaved/μl of CSF/24 h. The C-terminal peptides generated upon the cleavage of BigET-1 and BigET_18–34 were detected 48 h after incubation of these substrates with CSF. Cleavage of these substrates was inhibited by {"type":"entrez-protein","attrs":{"text":"CGS35066","term_id":"877962710","term_text":"CGS35066"}}CGS35066. Results of Western blots also produced strong evidence for the presence of truncated soluble ECE-1 in CSF. These results strongly suggest the presence of a truncated but catalytically active form of ECE-1 in the CSF of SAH subjects. Further studies are necessary to determine the biological significance of soluble ECE-1 in CSF of SAH subjects, including an association with vasospasm after SAH.Endothelin-1 (ET-1)¹ is the most potent vasoconstrictor in the central nervous system. Elevated levels of ET-1 in cerebrospinal fluid (CSF) have been implicated in the pathogenesis of cerebral vasospasm following subarachnoid hemorrhage (SAH) (1). However, it is not known whether the production of ET-1 within the CSF space contributes to the pathogenesis of vasospasm. ET-1 is produced upon the cleavage of its precursor BigET-1 by the highly specific metalloprotease Endothelin Converting Enzyme-1 (ECE-1). The rate-limiting step of ET-1 production is the expression and localization of ECE-1, whose catalytic activity is confined to the extracellular C-terminal domain. Previously, we have demonstrated that the catalytically active C terminus can be shed from the cell surface (2). This results in the release of a truncated but catalytically active form of ECE-1 into the extracellular milieu.Although the presence of a soluble form of ECE-1 has been demonstrated in vitro, it is yet to be shown in any human biological fluid. In this study, we have used a combination of mass spectrometry, Western blotting as well as quenched fluorescent substrate (QFS) based enzyme assays to demonstrate for the first time the presence of catalytically active, soluble form of ECE-1 in CSF of SAH subjects.     

The effects of continuous prostacyclin infusion on regional blood flow and cerebral vasospasm following subarachnoid haemorrhage: study protocol for a randomized controlled trial

ABSTRACT: BACKGROUND: One of the main causes of mortality and morbidity following subarachnoid haemorrhage (SAH) is the development of cerebral vasospasm, a frequent complication arising in the weeks after the initial bleeding. Despite extensive research, to date no effective treatment of vasospasm exists. Prostacyclin is a potent vasodilator and inhibitor of platelet aggregation. In vitro models have shown a relaxing effect of prostacyclin after induced contraction in cerebral arteries and a recent pilot trial showed positive effect on cerebral vasospasm in a clinical setting. No randomised, clinical trials have been conducted, investigating the possible pharmacodynamic effects of prostacyclin on the human brain following SAH. METHODS: This trial is a single-center, randomised, placebo controlled, parallel group, blinded, clinical, pilot trial. A total of 90 patients with SAH will be randomised to one of 3 intervention arms; epoprostenol 1 ng/kg/min, epoprostenol 2 ng/kg/min or placebo in addition to standard treatment. Trial medication will start day 5 after SAH and continue to day 10. Primary outcome measure is changes in regional cerebral blood flow from baseline in the arterial territories of the anterior cerebral artery, medial cerebral artery and the posterior cerebral artery, measured by CT perfusion scan. The secondary outcomes will be vasospasm measured by CT angiography, ischaemic parameters measured by brain microdialysis, flow velocities in the medial cerebral artery, clinical parameters and outcome (Glasgow Outcome Scale) at 3 months. CONCLUSION: The trial is an explorative, pilot trial designed to investigate the feasibility and possible effects of low-dose prostacyclin on a primary outcome of regional blood flow and vasospasm in the human brain following SAH. Trial registration: Clinicaltrials.gov NCT01447095.     

Activation of Rho-associated kinase during augmented contraction of the basilar artery to serotonin after subarachnoid hemorrhage

Delayed cerebral vasospasm after subarachnoid hemorrhage (SAH) may be due, in part, to altered regulation of arterial smooth muscle contraction. Contraction of cerebral arteries to serotonin is augmented after experimental SAH. We hypothesized that activation of Rho-associated kinase (Rho kinase) contributes to augmented contraction of cerebral arteries to serotonin after SAH. Autologous arterial blood (SAH) or artificial cerebrospinal fluid (control) was injected into the cisterna magna of anesthetized rabbits. At 2 days after injection, the basilar artery was excised and isometric contraction of arterial rings was recorded. Maximum contraction of the basilar artery to serotonin was augmented about fourfold in SAH compared with control rabbits (P < 0.01). Contraction to histamine was similar in the two groups. Fasudil hydrochloride (3 mumol/l), an inhibitor of Rho kinase, markedly attenuated serotonin-induced contraction. Fasudil had little effect on contractions induced by histamine or phorbol 12,13-dibutyrate. In addition, phosphorylation of myosin phosphatase, a major target of Rho kinase in regulation of smooth muscle contraction, in the basilar artery was examined by Western blotting. In basilar arteries of SAH, but not control, rabbits, serotonin increased phosphorylation of myosin phosphatase about twofold at Thr(853) of the myosin-targeting subunit. These results suggest that enhanced activation of Rho kinase contributes to augmented contraction of the basilar artery to serotonin after SAH.     

Efficacy of vision restoration therapy after optic neuritis (VISION study): study protocol for a randomized controlled trial

Background

One of the main causes of mortality and morbidity following subarachnoid haemorrhage (SAH) is the development of cerebral vasospasm, a frequent complication arising in the weeks after the initial bleeding. Despite extensive research, to date no effective treatment of vasospasm exists. Prostacyclin is a potent vasodilator and inhibitor of platelet aggregation. In vitro models have shown a relaxing effect of prostacyclin after induced contraction in cerebral arteries, and a recent pilot trial showed a positive effect on cerebral vasospasm in a clinical setting. No randomised, clinical trials have been conducted, investigating the possible pharmacodynamic effects of prostacyclin on the human brain following SAH.

Methods

This trial is a single-centre, randomised, placebo-controlled, parallel group, blinded, clinical, pilot trial. A total of 90 patients with SAH will be randomised to one of three intervention arms: epoprostenol 1?ng/kg/min, epoprostenol 2?ng/kg/min or placebo in addition to standard treatment. Trial medication will start day 5 after SAH and continue to day 10. The primary outcome measure is changes in regional cerebral blood flow from baseline in the arterial territories of the anterior cerebral artery, medial cerebral artery and the posterior cerebral artery, measured by CT perfusion scan. The secondary outcomes will be vasospasm measured by CT angiography, ischaemic parameters measured by brain microdialysis, flow velocities in the medial cerebral artery, clinical parameters and outcome (Glasgow Outcome Scale) at 3?months.

Trial registration

Clinicaltrials.gov NCT01447095.     

Time course of production of hydroxyl free radical after subarachnoid hemorrhage in dogs

Vasospasm after subarachnoid hemorrhage (SAH) is associated with lipid peroxidation. However, lipid peroxides increase in a delayed fashion after SAH and may be a byproduct of but not a cause of vasospasm. This study correlated vasospasm with hydroxyl free radical and lipid peroxide levels. 24 dogs had baseline cerebral angiography and induction of SAH by 2 injections of blood into the cisterna magna at baseline and 2 days later. Angiography was repeated 4, 7, 10, 14 or 21 days after the first injection (n = 4 per group) and a microdialysis catheter was inserted into the premedullary cistern. Control dogs (n = 4) underwent angiography and microdialysis but not SAH. Salicylic acid, 100 mg/kg, was administered intravenously, and microdialysis fluid was collected and analyzed by high pressure liquid chromatography for 2,3- and 2,5-dihydroxybenzoic acids (DHBA). Malondialdehyde was measured in subarachnoid clot removed from the prepontine cistern and in the basilar artery itself at the time of euthanasia. Significant vasospasm developed 4 to 14 days after SAH. Malondialdehyde levels were significantly elevated in the basilar artery and subarachnoid clot 4 days after SAH (p < 0.0001, ANOVA) but not at other times. 2,5-DHBA levels were significantly greater than control at 4 to 14 days and they peaked at 4 days (p < 0.05, ANOVA). 2,3-DHBA was significantly increased at 4 days after SAH (p < 0.05, ANOVA). There were significant correlations between basilar artery malondialdehyde levels and vasospasm and cerebrospinal fluid 2,5-DHBA levels and vasospasm. These results suggest the presence of hydroxyl free radical after SAH and demonstrate a correlation between such production, as measured by trapping with salicylate, and the early phase of vasospasm. The correlation with vasospasm implicates free radicals and lipid peroxidation in this phase of vasospasm.     

Expression of Nemo-Like Kinase (NLK) in the Brain in a Rat Experimental Subarachnoid Hemorrhage Model

This study aimed to investigate the expression of the Nemo-like kinase (NLK) in the brain after experimental subarachnoid hemorrhage (SAH) in rats. A total of 90 rats were randomly divided into six groups: control group, day 1, day 3, day 5, day 7, and day 14. Day 1, day 3, day 5, day 7, and day 14 groups were all SAH groups in which the rats were killed on days 1, 3, 5, 7, and 14, respectively. In SAH groups, autologous arterial blood was injected into cisterna magna once on day 0. Cross-sectional area of basilar artery was measured by H&E staining. Immunostaining and immunoblotting experiments were performed to detect the expression of NLK protein. Real-time polymerase chain reaction was used to analyze the presence and quantity of NLK mRNA. The level of oxidative stress in the artery was also measured. The basilar arteries exhibited vasospasm after SAH and became the most severe on day 3. The expressions of NLK protein and mRNA were decreased remarkably in SAH groups compared with the control group. The down-regulated expression of NLK was detected after SAH and the low ebb was on day 3, which was oppositely the peak time of oxidative stress. The expression of NLK was present mainly in the neurons in the brain and smooth muscle cells in the basilar artery. NLK is decreasingly expressed in an opposite time-course to the development of cerebral vasospasm (CVS) and SAH-induced brain injury in this rat experimental model of SAH and these findings might have important implications during the administration of specific NLK agonist to prevent or reduce CVS or neuronal apoptosis caused by SAH.     

Potential role of CD34 in cerebral vasospasm after experimental subarachnoid hemorrhage in rats

Inflammatory responses have been implicated in the elaboration of several forms of central nervous system injury, including cerebral vasospasm after subarachnoid hemorrhage (SAH). A critical event participating in such responses is the recruitment of circulating leukocytes into the inflammatory site. CD34 is a key adhesion molecule responsible for recruitment of monocytes/macrophages and the attachment of leukocytes to endothelial cells. However, it has not been investigated whether, and to what degree, CD34 is induced by SAH and also the role of CD34 in the pathogenesis of cerebral vasospasm following SAH remains unknown. Experiment 1 aimed to investigate the timecourse of the CD34 expression in the basilar artery after SAH. In experiment 2, we chose the maximum time point of vasospasm (day 3) and assessed the effect of monoclonal antibody against CD34 on regulation of cerebral vasospasm. As a result, the elevated expression of CD34 was detected in the basilar artery after SAH and peaked on day 3. After intracisternal administration of CD34 monoclonal antibody, the vasospasm was markedly attenuated after blood injection on day 3. Our results suggest that CD34 is increasingly expressed in a parallel time course to the development of cerebral vasospasm in a rat experimental model of SAH and administration of the specific CD34 antibody could prevent or reduce cerebral vasospasm caused by SAH.     

Phosphoramidon inhibits the conversion of intracisternally administered big endothelin-1 to endothelin-1

It is suggested that endothelin-1 (ET-1), a potent vasoconstrictor peptide, is involved in the pathogenesis of cerebral vasospasm following subarachnoid hemorrhage (SAH). We examined the effects of intracisternal administration of big ET-1 on the cerebral arteries in the absence or presence of pretreatment with phosphoramidon, an inhibitor of ET converting enzyme, in anesthetized dogs. After intracisternal administration of big ET-1 (10 micrograms/dog), the caliber of the basilar artery on the angiogram was decreased to about 59% of the control. This was accompanied by a marked increase in immunoreactive ET in the cerebrospinal fluid. Systemic arterial pressure was markedly elevated following big ET-1 injection. All changes induced by big ET-1 were effectively prevented with phosphoramidon. These data suggest that intracisternally administered big ET-1 is converted to ET-1 and that the generated ET-1 produces cerebral vasospasm and hypertension. A phosphoramidon-sensitive metalloproteinase appears to contribute to this conversion.     

17beta-estradiol inhibits endothelin-1 production and attenuates cerebral vasospasm after experimental subarachnoid hemorrhage

Though cerebral vasospasm after aneurysmal subarachnoid hemorrhage (SAH) has been recognized for over half a century, it remains a major complication in patients with SAH. Clinical studies have shown that elevated levels of endothelin-1 (ET-1) are present in the cerebrospinal fluid of patients with SAH, suggesting that ET-1-mediated vasoconstriction contributes to vascular constriction after SAH. Administration of estrogen promotes vasodilation in humans and in experimental animals, in part by decreasing the production of ET-1. This study evaluated the influence of 17beta-estradiol (E2) on the production of ET-1 and cerebrovasospasm in an experimental SAH 2-hemorrhage model in rat. A 30-mm Silastic tube filled with E2 in corn oil (0.3 mg/ml) was subcutaneously implanted in male rats just before SAH induction. The degree of vasospasm was determined by averaging the cross-sectional areas of basilar artery 7 days after first SAH. Plasma samples collected before death were assayed for ET-1. The protective effect of E2 in attenuating vasospasm achieved statistical significance when compared with the SAH only or SAH plus vehicle groups (P < 0.01). Concentrations of ET-1 were higher in the SAH only and SAH plus vehicle groups than in controls (P < 0.001). Serum levels of ET-1 in the SAH plus E2 and E2 only groups were significantly lower than those in the SAH only and SAH plus vehicle groups (P < 0.001). There was no significant difference between ET-1 levels in the healthy control and SAH plus E2 groups. A significant correlation was found between the cross-sectional areas of basilar artery and ET-1 levels (P < 0.001). The beneficial effect of E2 in attenuating SAH-induced vasospasm may be due in part to decreasing ET-1 production after SAH. The role of E2 in the treatment of cerebral vasospasm after SAH is promising and is worthy of further investigation.     

A Low Mortality Rat Model to Assess Delayed Cerebral Vasospasm After Experimental Subarachnoid Hemorrhage

Objective: To characterize and establish a reproducible model that demonstrates delayed cerebral vasospasm after aneurysmal subarachnoid hemorrhage (SAH) in rats, in order to identify the initiating events, pathophysiological changes and potential targets for treatment.Methods: Twenty-eight male Sprague-Dawley rats (250 - 300 g) were arbitrarily assigned to one of two groups - SAH or saline control. Rat subarachnoid hemorrhage in the SAH group (n=15) was induced by double injection of autologous blood, 48 hr apart, into the cisterna magna. Similarly, normal saline (n=13) was injected into the cisterna magna of the saline control group. Rats were sacrificed on day five after the second blood injection and the brains were preserved for histological analysis. The degree of vasospasm was measured using sections of the basilar artery, by measuring the internal luminal cross sectional area using NIH Image-J software. The significance was tested using Tukey/Kramer''s statistical analysis.Results: After analysis of histological sections, basilar artery luminal cross sectional area were smaller in the SAH than in the saline group, consistent with cerebral vasospasm in the former group. In the SAH group, basilar artery internal area (.056 μm ± 3) were significantly smaller from vasospasm five days after the second blood injection (seven days after the initial blood injection), compared to the saline control group with internal area (.069 ± 3; p=0.004). There were no mortalities from cerebral vasospasm.Conclusion: The rat double SAH model induces a mild, survivable, basilar artery vasospasm that can be used to study the pathophysiological mechanisms of cerebral vasospasm in a small animal model. A low and acceptable mortality rate is a significant criterion to be satisfied for an ideal SAH animal model so that the mechanisms of vasospasm can be elucidated ^{7, 8}. Further modifications of the model can be made to adjust for increased severity of vasospasm and neurological exams.     

Melatonin Ameliorates Cerebral Vasospasm After Experimental Subarachnoidal Haemorrhage Correcting Imbalance of Nitric Oxide Levels in Rats

In the present study, we investigated the in vivo effects of melatonin on SAH-induced cerebral vasospasm and oxidative stress, resulting from SAH in an experimental rat model. Twenty-eight rats (225–250 g) were divided into four groups equally: group 1; control, group 2; SAH, group 3; SAH plus placebo, and group 4; SAH plus melatonin. We used double haemorrhage method for SAH groups. Beginning 6 h after SAH, 20 mg/kg melatonin or equal volume of 0.9% saline was administered intraperitoneally twice daily for 5 days to groups 3 and 4, respectively. Melatonin or 0.9% saline injections were continued up to fifth day after SAH and rats were sacrificed at the end of this period. Brain sections at the level of the pons were examined by light microscopy. The lumen diameter and the vessel wall thickness of basilar artery were measured using a micrometer. The serum levels of cerebral vasodilator nitric oxide (NO), the brain levels of an intrinsic antioxidant superoxide dismutase (SOD) and a NO regulator arginase activities were measured. The brain levels of inducible nitric oxide (iNOS) and nitrotyrosine, a nitrosative stress parameter immunohistochemiacally determined. In conclusion, melatonin administration ameliorated cerebral vasospasm by increasing serum NO level and decreasing the brain the levels of arginase and oxidative stress. It is therefore possible that increased brain arginase activity after SAH may also have a significant role in the pathogenesis of vasospasm by limiting the availability of arginine for NO production.     

Acute and delayed vasoconstriction after subarachnoid hemorrhage: local cerebral blood flow, histopathology, and morphology in the rat basilar artery.

The decreased local cerebral blood flow (LCBF) and cerebral ischemia that occur after subarachnoid hemorrhage (SAH) may be caused by acute and/or delayed vasospasm. In 36 Sprague-Dawley (350-450 g) rats SAH was induced by transclival puncture of the basilar artery. Mean arterial blood pressure (MABP), LCBF, intracranial pressure (ICP), and cerebral perfusion pressure (CPP) were measured in all rats for 30 min before and 60 min after SAH was induced. One set of control (n : 7) and experimental animals (n : 7) was sacrificed after the 60 min of initial post-hemorrhage measurements were recorded. Four days after SAH induction, LCBF and MABP were measured again for 60 min in subgroups of surviving experimental rats (n : 7) and control rats (n : 7). Histopathologic and morphologic examinations of the basilar artery were performed in each subgroup. There was a sharp drop in LCBF just after SAH was induced (55.50 +/- 11.46 mlLD/min/100 g and 16.1 +/- 3.6 mlLD/min/100 g for baseline and post-SAH, respectively; p < 0.001). The flow then gradually increased but had not returned to pre-SAH values by 60 min (p < 0.05). At 4 days after SAH induction, although LCBF was lower than that observed in the control group and pre-SAH values, it was not significantly different from either of these flow rates (p > 0.05). ICP (baseline 7.05 +/- 0.4 mmHg) increased acutely to 75.2 +/- 7.1 mmHg, but returned to normal levels by 60 min after SAH. CPP (baseline 84.5 +/- 6.3 mmHg) dropped accordingly (to 18.6 +/- 3.1 mmHg), and then increased, reaching 72.2 +/- 4.9 mmHg at 60 min after SAH (p > 0.05). Examinations of the arteries revealed decreased inner luminal diameter and distortion of the elastica layer in the early stage. LCBF in nonsurviver rats (n : 8) was lower than that in the animals that survived (p < 0.01). At 4 days post-hemorrhage, the rats' basilar arteries showed marked vasculopathy. The findings showed that acute SAH alters LCBF, ICP, and CPP, and that decreased LCBF affects mortality rate. Subsequent vasculopathy occurs in delayed fashion, and this was observed at 4 days after the hemorrhage event.     

Phosphoramidon prevents cerebral vasospasm following subarachnoid hemorrhage in dogs: the relationship to endothelin-1 levels in the cerebrospinal fluid

There is increasing evidence that the conversion of big endothelin-1 (big ET-1) to endothelin-1 (ET-1) is specifically inhibited by the metalloproteinase inhibitor phosphoramidon. We investigated the effect of phosphoramidon on delayed cerebral vasospasm from subarachnoid hemorrhage (SAH) using a two-hemorrhage canine model. The magnitude of the vasospasm and the drug effect were determined angiographically. On SAH Day 7, diameter of the basilar artery decreased to about 55% of the control value obtained before SAH (on Day 0). Immunoreactive ET (IR-ET) in the cerebrospinal fluid (CSF) significantly increased after SAH (on Day 7). The intracisternal pretreatment of phosphoramidon potently suppressed the decrease in diameter of the basilar artery after SAH, i.e., observed decrease was only about 20%, compared with the value before SAH. In the phosphoramidon group, IR-ET in CSF markedly increased (on SAH Day 2), but the increased levels of IR-ET significantly declined on SAH Day 7. These results clearly indicate that phosphoramidon effectively prevents delayed cerebral vasospasm. Whether the prevention is due to the inhibition of conversion of big ET-1 to ET-1 is now under study.     

Effect of endothelin-1 receptor antagonist BQ-123 on basilar artery diameter after subarachnoid hemorrhage (SAH) in rats.

Aim of the study was to quantify cerebral vasospasm in rats after subarachnoid hemorrhage (SAH) by morphometric examination of basilar artery and to evaluate the influence of endothelin receptor blocker BQ-123 on basilar artery constriction. The rat cisterna magna (CM) was cannulated and after 7 days SAH was developed by administration of 100 microl autologic, non-heparinized blood to the CM. The sham subarachnoid hemorrhage was developed by intracisternal administration of 100 microl of artificial cerebrospinal fluid. Endothelin receptor blocker BQ-123 was injected into the CM in a dose of 40 nmol diluted in 50 microl of cerebrospinal fluid 20 min. before SAH, and 24h and 48 h after SAH. After perfusion fixation the brains were removed from the skull and histological preparations of basilar artery were done. The internal diameter and wall thickness of basilar arteries was measured by interactive morphometric method. The most severe vasospasm was found in rats after SAH. The presence of numerous infiltrations composed of neutrophils and macrophages correlated with advanced vasospasm (index of constriction 5 times lower than in normal), suggesting the role of other factors participating in the late phase of vasospasms after SAH. Administration of BQ-123 in the late phase after SAH caused the dilatation of basilar artery. Following the administration of BQ-123 in the late phase (48 h after SAH) the basilar artery dilated, its wall became thinner, and the number of leukocyte infiltrations in the subarachnoid space decreased compared to the values after SAH alone.     

内皮舒张因子与氧自由基在脑血管痉挛发病中的作用

实验性兔蛛网膜下腔出血后,基底动脉壁丙二醛（ＭＤＡ）含量及超氧化物峻化酶（ＳＯＤ）、过氧化氢酶（ＣＡＴ）活性发生改变,基底动脉出现痉挛,应用ＳＯＤ后上述变化减轻。离体采用生物检定法发现,基底动脉受自由基损伤后,去甲肾上腺素（ＮＥ）诱导的血管收缩效应增强,而ＡＣｈ诱导的血管舒张效应减弱。用ＳＯＤ防止了ＡＣｈ诱导的血管舒张效应的减弱。结果表明,氧自由基参与了脑血管痉挛的发生,而脑血管受自由基损伤后,其内皮舒张因子释放减少是脑血管痉挛发病的重要因素。     

Effect of experimental subarachnoid hemorrhage on calcium incorporation and adrenergic innervation of the cerebral arteries of the cat

The effect of experimental subarachnoid hemorrhage (SAH) on the adrenergic innervation and on the 45Ca2+ uptake of cat cerebral arteries was analyzed. Intracisternal injections of autologous blood reduced the noradrenaline content of perivascular nerve endings and 3H-noradrenaline uptake. These values returned to normal levels in a period of two weeks after SAH. The activity of dopamine-beta-hydroxylase was also reduced 3 and 7 days after SAH. Superior cervical gangliectomy and intracisternal injection of 6-hydroxydopamine also reduced these three parameters. The uptake of 45Ca2+ by arteries from animals submitted to SAH was greater than if the blood vessels were from untreated cats. Lantanum brought about a less 45Ca2+ displacement in the arterial segments from untreated animals than in those from cats after SAH. These results suggest that SAH induces a transient adrenergic denervation as well as changes in the membrane of smooth muscle cell which increase the quantity of Ca2+ bound to it. All these modifications might be involved in the cause of chronic cerebral vasospasm that appears after SAH.     

大鼠脑血管痉挛时NO和ET—1变化及尼莫地平的影响

目的探讨蛛网膜下腔出血(SAH)后脑血管痉挛(CVS)时脑组织一氧化氮(NO)和内皮素-1(ET-1)含量变化及尼莫地平(ND)对其影响。方法将135只Wistar大鼠随机均分为SAH组、ND处理组和假手术组,观察手术前后基底动脉管径,及24h内局部脑血流量(rCBF)、脑组织NO和ET-1含量动态改变,并行海马病理检查。结果SAH后rCBF明显而持续降低,基底动脉管径显著缩小;海马CAl区锥体细胞严重受损;脑组织NO和ET-1含量均在SAH后1～24h显著增加(P＜0．05～0．01)。ND处理后使上述异常变化均减轻。结论SAH后脑组织NO、ET-1增多可能参与了CVS所致脑损害过程,ND通过减轻CVS和拮抗脑组织NO及ET-1的病理性改变而发挥脑保护作用。