Influence of Pregnancy on Blood-Brain Barrier Integrity During Seizures in Rats
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Evans Blue
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The blood-brain barrier (BBB) is an active and selective barrier that shields the brain from endogenous and exogenous insults. Different stimuli may lead to the disruption of this barrier, including inflammation and trauma. Several methods are used to evaluate BBB disruption. The most widely used method is Evans blue (EB) dye extravasation. EB cannot normally pass through the BBB and thus its presence in brain tissue indicates alterations in permeability. This protocol details the steps of EB extravasation in rodents. Important aspects regarding critical steps and advantages are also provided. © 2019 by John Wiley & Sons, Inc.
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AIM: To discuss the influence of resveratrol on the damage of blood brain barrier(BBB) in permanent focal cerebral ischemic injury in mice.METHODS: Ischemia was induced by intraluminal permanent middle cerebral artery occlusion(MCAO)and Evans blue was injected.The extent and quantity of Evans blue extravasation were measured and the expression of MMP-9 was shown by RT-PCR.RESULTS: Resveratrol significantly(diminished) the extent and quantity of Evans blue extravasation [(101±15)mm~3 vs(149±17) mm~3,(8.6±2.9)% vs((18.6±)3.8)%,P0.01] and inhibited the expression of MMP-9 [(0.693±0.008) vs(1.537±0.037),P0.01].(CONCLUSION:) Resveratrol,due to its neuroprotective effect,can inhibit the activity of MMP-9 and alleviate the disruption of BBB.
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Vascular permeability
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Objective:The purpose of this study was to correlate permeability parameters measured with dynamic contrast-enhanced magnetic resonance imaging (DCE-MRI) using a clinical 3-tesla scanner with extravasation of Evans blue in a rat model with transient cerebral ischemia.Materials and Methods: Sprague-Dawley rats (n = 13) with transient middle cerebral artery occlusion were imaged using a 3-tesla MRI with an 8-channel wrist coil.DCE-MRI was performed 12 hours, 18 hours, and 36 hours after reperfusion.Permeability parameters (K trans , ve, and vp) from DCE-MRI were calculated.Evans blue was injected after DCE-MRI and extravasation of Evans blue was correlated as a reference with the integrity of the blood-brain barrier.Correlation analysis was performed between permeability parameters and the extravasation of Evans blue.Results: All permeability parameters (K trans , ve, and vp) showed a linear correlation with extravasation of Evans blue.Among them, K trans showed highest values of both the correlation coefficient and the coefficient of determination (0.687 and 0.473 respectively, p < 0.001).Conclusion: Permeability parameters obtained by DCE-MRI at 3-T are well-correlated with Evans blue extravasation, and K trans shows the strongest correlation among the tested parameters.
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The influence of chronic arterial hypertension upon the permeability to albumin of the cerebral capillaries, i.e. the blood-brain barrier, was studied in normotensive Wistar and spontaneously hypertensive Wistar rats with experimental subarachnoid hemorrhage. The blood-brain barrier permeability to albumin was assessed quantitatively by spectrophotometric determination of Evans blue extravasation. Subarachnoid hemorrhage was produced by injecting autologous blood into the cortical subarachnoid space. A significant increase of Evans blue albumin extravasation was found in the spontaneously hypertensive rats with subarachnoid hemorrhage as compared with normotensive animals suffering from subarachnoid hemorrhage. Subarachnoid hemorrhage in this model alone caused a significant Evans blue extravasation, whereas sham-operation did not. These findings emphasize the necessity for effective attempts to reduce the leakage of the capillary system in the early stage of subarachnoid hemorrhage.
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Immersion of the hind paws of anaesthetized rats in hot water for 5 min induced massive plasma protein leakage as indicated by extravasation of Evans blue dye in the skin. The threshold temperature which caused noticeable plasma extravasation was 45°C, a maximal response was obtained between 55°C and 60°C. Pretreatment of rats 2 days after birth with 50 mg kg −1 capsaicin significantly reduced the Evans blue extravasation induced by hot water at 50°C, 55°C and 60°C, whereas guanethidine pretreatment 24 h before the experiment caused a significantly increased response at 40°C, 45°C and 50°C. When Evans blue was injected between 10 and 120 min after immersion of the paw in hot water, a significant extravasation of the dye was no longer detectable. However, the weight of the paw as well as the weight of the piece of skin taken for Evans blue quantification increased during this period indicating the progressive development of oedema in the skin and underlying tissues. In rats treated with capsaicin as neonates, the increase in paw weight after immersion in water of 50°C for 5 min was significantly delayed during the first hour, but there was no difference after two hours. In rats pretreated with D ‐Arg 1 , D ‐Pro 2 , D ‐Trp 7,9 , Leu 11 ‐substance P, a substance P (SP) antagonist, the Evans blue extravasation was significantly reduced. However, the response, which remained in rats treated with capsaicin as neonates was not blocked by the SP‐antagonist. It is concluded that activation of peripheral branches of sensory SP neurones contributes to the initial massive protein extravasation and to the subsequent rate of development of oedema following heat injury. Release of histamine did not significantly contribute to this response at the lower temperatures, although the response was reduced by histamine receptor blocking drugs at 55°C and 60°C. Decreasing the sympathetic vasoconstrictor tone by guanethidine resulted in an increased response.
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The contribution of neutrophils to the action of endotoxin on plasma exudation in the airways of anaesthetized guinea-pigs was quantified by measuring the extravasation of Evans blue dye. Endotoxin (Salmonella enteritidis) caused a dose-dependent increase in microvascular leakage to Evans blue dye which was maximal after 25 min (p less than 0.05). The minimum dose tested that induced a significant rise in leakage was 1.5 mg.kg-1 for "central" intrapulmonary airways (ipa); 4.5 mg.kg-1 for trachea and main bronchi and 7.5 mg.kg-1 for nasal mucosa, larynx and "peripheral" ipa. Depletion of circulating neutrophil numbers by 97% using an antibody to guinea-pig neutrophils caused no significant diminution of the effects of endotoxin on leakage in any part of the airway. There was no significant influx of neutrophils into the airway interstitium at the time of maximum extravasation of Evans blue. We conclude that endotoxin-induced airway microvascular permeability is dependent upon mechanisms other than circulating neutrophils.
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The influence of chronic arterial hypertension upon the permeability to albumin of the cerebral capillaries, i.e. the blood-brain barrier, was studied in normotensive Wistar and spontaneously hypertensive Wistar rats with experimental subarachnoid hemorrhage. The blood-brain barrier permeability to albumin was assessed quantitatively by spectrophotometric determination of Evans blue extravasation. Subarachnoid hemorrhage was produced by injecting autologous blood into the cortical subarachnoid space. A significant increase of Evans blue albumin extravasation was found in the spontaneously hypertensive rats with subarachnoid hemorrhage as compared with normotensive animals suffering from subarachnoid hemorrhage. Subarachnoid hemorrhage in this model alone caused a significant Evans blue extravasation, whereas sham-operation did not. These findings emphasize the necessity for effective attempts to reduce the leakage of the capillary system in the early stage of subarachnoid hemorrhage.
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Extravasation
Vascular permeability
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Autologous blood
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