Name: modeling stroke in mice: permanent coagulation of the distal middle cerebral artery
Uploaded: 2014-07-31T14:00:00.000+00:00
Duration: 8 min 14 s
Description: Watch this Scientific Journal Video about Modeling Stroke in Mice: Permanent Coagulation of the Distal Middle Cerebral Artery at JoVE.com

这项工作是由德国研究基金会"慕尼黑集群的神经系统（协同）"的精英集群和戴姆勒 - 奔驰的基础，AL

伦理声明报道在这个视频中的实验是在按照批准了德国政府委员会（Regierung冯Oberbayern，慕尼黑，德国）国家指导方针的使用实验动物和协议进行的。 10周岁，雄性C57BL/6J小鼠被用于这项研究。动物饲养控制温度（22±2℃）下，用12小时明暗周期和访问。镇痛和镇静协议被描述为经当地政府委员会沉淀的食物和水随意 ，但可能会有所不同从其他实验室使用的协议。 材料1。制备和仪器<ol><li>连接的热毯，以保持操作区域温暖和维持麻醉期间的小鼠的体温（37℃）。 </li><li>准备蒸压剪刀，镊子和棉花，dexpanth烯醇眼膏和缝合材料。准备用生理盐水溶液（不带针头）注射器保持水润的操作区域。准备麻醉气体（70％N 2 O + 30％的O 2 +异氟醚）。 </li><li>注入镇痛药腹膜内:安乃近200毫克/千克，卡洛芬4毫克/千克和Buprenorphin 0.1毫克/公斤。 </li><li>将鼠标放置到感应室与4％的异氟醚流量麻醉，直到身体的自发运动和触须停止。 </li><li>传输鼠标与它的鼻子卧位到麻醉面罩和维持异氟醚浓度在4％约一分钟，然后降低它在约1.5％，以维持适当的麻醉。 </li><li>在双眼适用泛醇眼膏。 </li></ol> 2，远端MCAO模型<ol><li>让耳朵和眼睛之间的1厘米的皮肤切口用小剪刀操作手术部位加兹登的aseptical准备后克的皮肤消毒剂。 </li><li>分离皮肤和本地化颞肌。 </li><li>在高频发生器选择凝血功能，双极模式，选择12 W和与连接线连接的电凝镊子。 </li><li>加一滴生理盐水，然后使用镊子从它的顶端和背部部分颅骨分离颞肌，从而使肌肉瓣没有完全去除肌肉。 </li><li>识别马华下面的透明头骨，在颞区的延髓部分，背到眼眶后静脉窦（ 图2A）。如果MCA分叉是不可见的（由于解剖的正常变化）确定容器最喙。 </li><li>添加一些盐水的头骨和薄了骨与钻右上方MCA分支，直到它有一个薄的和半透明的质感（ 图2B）。 </li><li>仔细撤回骨动脉上一个非常薄的镊子。 </li><li>选择双极模式in在7 W.混凝高频发生器与电凝动脉钳子近端和远端分叉（ 图2C）。当分叉是不可见的，由于解剖变异体，凝结正确识别MCA分支（见上文）以大约两个站点。 1mm的。这是没有必要掌握动脉与镊子凝固，用在两侧从上面的镊子小心碰触动脉是充分，导致更少的机械损伤。 </li><li>等待30秒，然后轻轻触碰动脉用钝钳来检查任何血流由于自发再通。万一再通的重复电一次。 </li><li>搬迁颞肌它的位置，覆盖了钻孔。 </li><li>缝合伤口，然后将动物在护理盒在32℃下从麻醉中恢复，并回到笼子里。一般来说，需要5-10分钟的动物从anesth恢复ESIA。 </li><li> 24小时后注射术后镇痛（IP），然后每天直到术后第5天:卡洛芬4毫克/千克。 </li></ol> 3，假手术相同地执行所有的程序，以上述的操作 - 包括头骨和其去除变薄 - 除了不凝固暴露动脉。 4，油缸测试13 <ol><li>将动物在透明的亚克力玻璃圆筒（直径:8厘米，高度:25厘米）的两个反射镜和录像带5分钟前。中央调整相机在两面镜子的前面，气缸，以获得最佳的视频（ 图3A）。 </li><li>对于独立的前肢使用评估，气缸壁与一个前肢得分（1）在全后部接触，（2）着陆只有一个前肢地板上满后后。计数至少20个联系人一个前肢使用慢动作或逐FR视频岚客户端（VLC），注释功能的免费软件（ <a href="http://www.videolan.org/vlc">http://www.videolan.org/vlc</a> ）。 </li><li>对于术前基线分析:每只小鼠进行试验两次，试验之间有1小时的休息。前肢的使用表示为左/右片面的，独立的前肢使用比例。 </li><li>马华凝固后:每只小鼠再次进行测试两次，如上文所述试验之间有1小时的休息。 </li></ol> 5，灌注<ol><li> （氯胺酮和甲苯噻嗪一十六分之一百二十零毫克/千克体重，分别为如 ）麻醉动物。 </li><li>修复动物仰卧位置并用中值切割打开腹腔。删除RIP和胸骨。做一个小切口在右心房。插入灌注导管在左心室和用20毫升盐水的缓慢灌注。 </li><li>斩首动物，打开颅骨，并从颅底轻轻取下脑。 </li><li&gt;冻结的大脑在-40°C的低温异戊烷10分钟。随后从异戊烷和存储中删除的大脑，直到进一步的分析在-80°C。 </li></ol> 6。梗死容量法<ol><li> Cryosectioning:在低温恒温器连续切割的大脑到20微米厚的部分每400微米的幻灯片，并储存在-20幻灯片°C。 </li><li>甲酚紫（CV）染色: <ol><li>准备染色液:将将0.5g的简历醋酸500毫升H 2 O搅拌和加热（60℃）直至结晶溶解。 ，溶液冷却并存储在一个黑暗的瓶子。每次使用前重新加热到60°C和过滤器。 </li><li>干燥载玻片在室温下30分钟。然后将它们放置在95％乙醇中15分钟，然后在70％乙醇中1分钟，并随后在50％乙醇中1分钟。 </li><li>将载玻片在蒸馏水中2分钟，刷新蒸馏水并将其放置在1分钟。随后将幻灯片染色溶液（60℃）F或10分钟，并用蒸馏水洗涤两次他们持续1分钟。 </li><li>将玻片在95％乙醇中2分钟。然后将它们放入100％乙醇5分钟，刷新了100％的乙醇，并放置在2分钟。随后盖上安装介质的幻灯片。 </li><li>分析: 扫描幻灯片和由斯旺森方法14分析间接梗死体积，以纠正水肿: （缺血区域）=（皮质对侧的面积） - （同侧的非缺血皮层区域）（ 图4A）。 </li></ol></li></ol>

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作者没有竞争的利益透露。

本协议描述远端，永久性MCAO的实验性中风模型经颅电 - 即所谓的"凝模式"。这种模式已经同时成为在实验中风研究12中最常用的动物模型之一。相比其他局灶性脑缺血模型，如在本视频呈现的凝血模型具有约10分钟时，由受过训练的科学家进行的很短的运行时间的优点。因此，短暂的麻醉时间可以在此模型中，这是一个实验性中风模型中的有利特性，因为麻醉药对神经保护和中风结果的影响是众所周知的15来实现。此外，如先前由迈克尔等人 11所述，我们证实了所得到的梗死体积和本地化远侧永久性MCA凝固后对应于缺血性脑损伤中的大多数人的招比例大脑的大小。人的中风主要是小的与半球的约5-15％的病变大小，根据以前的人口研究和临床成像试验16-18，而相比之下，广泛中风病变抗压性脑水肿发生在小于10％的临床笔画数19。因此，中风病灶的12％左右由该模型实现了半球的马华领土可以被视为一个平移相关每搏输出量。然而，它必须被考虑到不同的小鼠品系或用于麻醉的协议，可能会影响所产生的损伤体积20。 在此模型中诱导中风后的观察期死亡率是几乎不存在。小于5％的总死亡率，主要是因为由anesthesiological并发症或牺牲的操作过程中死亡的，因为达到排除标准。为了保证低V这种模式和其出色的重复性ariability，我们建议以下排除标准:1）在操作过程中的任何蛛网膜下腔出血。 2）操作时间超过15分钟。 3）再通MCA的两次尝试的电只有短暂MCAO后。此外，动物需要被MCAO后（基本的生理行为，毛皮的外观和体重），以控制疼痛，不适或病态行为的日常检查。 可能对于像激光散斑测量，磁共振成像，行为测试或组织学分析中风结果分析实施多项措施。在这个协议中，我们提供了示例性方法的行为分析和脑梗死体积的分析。几个测试后局灶性脑缺血行为分析已经开发并在实验中风研究使用。先前在此行程模型21,22我们用我们的组感觉功能障碍合适的测试再旋转试验23，标签粘性测试24，测试角25和气缸测试13，这足以证明这部影片。气缸测试一贯描绘电机在不对称末端永久性MCAO后的急性期，也检测到连续的恢复运动功能。 尽管有明显的优势，这中风模型的一些限制必须被考虑在内。首先，颅骨的环锯术是必要的，以凝聚动脉，从而产生对大脑的围手术感染的电位访问，虽然手术伤口，颞肌或大脑本身的细菌感染从未由自己检测或记录被别人采用这种模式。此外，制备和凝固过程中的机械损坏，皮质不能被排除，但可以通过仔细钻探和去除颅骨，手术部位的恒定加湿和最小NE被限制<html> cessary电（参见排除标准）。虽然MCA的如在图2中所描绘的过程在大多数C57BL / 6小鼠的发现，我们描述在协议如何继续在容器中的过程中的正常变化，以减少模型的变异性。此外，我们建议使用多个（在分叉3; 2没有的MCA分叉）闭塞部位，以尽量减少在MCA，这在我们的经验是这种模式的变化的一个重要因素部分再通的风险。 在行为测试而言，只有轻微的行为缺陷可以在上述的行为测试来检测和功能性再生可以中风后的第一个星期内被观察到。因此，更先进的测试系统具有更高的灵敏度和定性试验参数，如本领域技术到达测试27可能更适合于检测长期功能预后在该模型中。 ontent"&gt;最后，由于在MCA不能得到再灌注，这是在中风患者由于自发凝块溶解或治疗28在相当大比例观察到的特征的永久凝固，但是，先前描述的血栓栓塞性中风模型26提供与皮质脑缺血再灌注免费中风模型的选项。综合来看，高重复性，可能的长期观察，由于最小的死亡率和可比性相对梗死体积和定位在尊重人的行程区分"凝结模式"作为基础和转化中风研究的宝贵模型。

中风是导致死亡的第三大最常见的原因和后天残疾的成人在发达国家1其中一个主要的原因。约80％的这种急性神经疾病是由脑缺血再由脑血流的阻塞导致引起，而约15％是由一种脑出血2引起的。尽管目前的研究，组织型纤溶酶原激活剂静脉给药是唯一批准的药物治疗缺血性中风，到目前为止，只适用于中风患者少数因卒中发病后3,4 4.5小时的短批准的时间窗口。由于没有体外模型，它可以正确地模拟大脑，血管和中风期间全身的病理生理机制之间复杂的相互作用，动物模型是中风的临床前研究是必不可少的。 因此，一些缺血性中风模型已经开发了一个变量ETY种。一个最常用的中风模型是"长丝模型"，其中缝线丝被瞬时引入到颈内动脉和转发，直到尖端闭塞大脑中动脉（MCA）的原点，从而导致血流的停止和皮层下及在长时间的情况下闭塞也皮层区域5,6的后续脑梗塞。在缺血性中风的光化学模型，照射皮质容器的光化学闭塞喷射导致小的，局部外切病变7光敏剂后得以实现。该lenticulostriatal动脉MCA远端的永久性闭塞可通过动脉，其瞬时压缩的结扎或电凝永久8,9来实现。在这个模型中所产生的梗塞主要影响新皮层10，因为马华在这个模型中是闭塞远端的静水ulostriatal动脉，它提供基底节。 由于多数人的中风病灶位于大脑中动脉供血区，所有常用的中风模型类似于MCA的闭塞或其分支11之一。马华是提供血液供应到大脑的主要动脉之一;它产生于颈内动脉，路线沿外侧沟在那里再分支和项目基底节和额叶，顶叶和颞叶，包括主运动和感觉皮质的侧面。右侧和左侧的MCA连接到大脑前动脉和后交通动脉，它连接到大脑后动脉，创建Willis环（ 图1）。 正如先前报道的卡迈克尔等人 11，梗塞的远端大脑中的Ar为蓝本计算电池闭塞（MCAO）模型小鼠中包括约10-15％的半球，从而模仿大多数人的中风病灶分别位于皮质MCA ​​供血区11,12。 1981年，田村等人在大鼠8所述的永久，经颅MCAO凝固模型。然而，通过田村描述的模型涉及为了规避动脉的更远端分叉MCA的近端闭塞。因而，原来的"田村模型"诱导不仅皮质而且纹状体病变，类似的"灯丝模型"6得到的病变。在这里，我们描述了永久的远端MCAO模型小鼠经颅电。此外，我们报告相关的组织学和功能的方法来分析这个模型的中风结果。所有的方法都是基于开发并在我们的实验室使用的标准作业程序。

<table><tbody><tr><td>Camera Canon Eos 500D</td><td>Canon</td><td></td><td>Optics: 18-55 mm; 30 fps; 640 x 480 video function</td></tr><tr><td>Mirror</td><td>Kristallform</td><td>2677089</td><td>30 x 30 cm&amp;nbsp;</td></tr><tr><td>Transparent acrylic glass cylinder</td><td>H&amp;#38;S Kunststofftechnik</td><td></td><td>Diameter: 8 cm,&amp;nbsp; height: 25 cm</td></tr><tr><td>Heating blanket</td><td>FHC DC Temperature Controller</td><td></td><td></td></tr><tr><td>Fine Scissors</td><td>FST</td><td>15000-00</td><td></td></tr><tr><td>Mayo Scissors</td><td>FST</td><td>1410-15</td><td></td></tr><tr><td>Forceps</td><td>FST</td><td>11616-15</td><td></td></tr><tr><td>Cottons</td><td>NOBA Verbondmitel Danz</td><td>974116</td><td></td></tr><tr><td>Saline solution</td><td>Braun</td><td>131321</td><td></td></tr><tr><td>Bepanthen pommade</td><td>Bayer</td><td></td><td></td></tr><tr><td>Isoflurane</td><td>Abbot</td><td>B506</td><td></td></tr><tr><td>Anesthesia system for isoflurane</td><td>Drager</td><td></td><td></td></tr><tr><td>Stereomikroskop&amp;nbsp;</td><td>Zeiss</td><td>Stemi DV4</td><td></td></tr><tr><td>Electrosurgical device</td><td>ERBETOM ICC 80/50 HF-Chirurgieger&amp;auml;t</td><td></td><td></td></tr><tr><td>Drill</td><td>Proxxon</td><td>D-34343</td><td></td></tr><tr><td>Ketamine</td><td>Inresa Arzneimittel GmbH</td><td></td><td></td></tr><tr><td>Xylacine</td><td>Albrecht</td><td></td><td></td></tr><tr><td>5ml Syringe&amp;nbsp;</td><td>Braun</td><td></td><td></td></tr><tr><td>Phosphate Buffered Saline PH: 7.4</td><td>Apotheke Innestadt Uni Munchen</td><td>P32799</td><td></td></tr><tr><td>Isopentane</td><td>Fluka</td><td>59070</td><td></td></tr><tr><td>Cryostat</td><td>Thermo Scientific CryoStarNX70</td><td></td><td></td></tr><tr><td>Superfrost Plus Slides</td><td>Thermo Scientific</td><td>J1800AMNZ</td><td></td></tr><tr><td>Cresyl violet</td><td>Sigma Life Science</td><td>C5042-10G</td><td></td></tr><tr><td>Acetic acid</td><td>Sigma Life Science</td><td>695092</td><td></td></tr><tr><td>Ethanol 70%</td><td>CLN Chemikalien Laborbedorf</td><td>521005</td><td></td></tr><tr><td>Ethanol 96%</td><td>CLN Chemikalien Laborbedorf</td><td>522078</td><td></td></tr><tr><td>Ethanol 99%</td><td>CLN Chemikalien Laborbedorf</td><td>ETO-5000-99-1</td><td></td></tr><tr><td>Roti-Histokit mounting medium</td><td>Roth</td><td>6638.1</td><td></td></tr><tr><td>C57Bl/6J mice</td><td>Charles River</td><td>000664</td><td></td></tr></tbody></table>

modeling stroke in mice: permanent coagulation of the distal middle cerebral artery

中风是导致死亡的第三大最常见的原因，在发达国家收购成人残疾的主要原因。只有非常有限的治疗选项可用于急性期的一小部分中风患者。目前的研究集中寻找新的治疗策略，并越来越专注于卒中后亚急性和慢性期，因为更多的患者可能有资格获得治疗干预在长期的时间窗口。这些延迟的机制包括重要的病理生理途径，如中风后的炎症，血管生成，神经可塑性和再生。为了分析这些机制并随后评估新的药物靶标，具有临床相关性，低死亡率和高重现性试验行程车型受到追捧。此外，老鼠是在其中一个焦点中风损伤可诱导和其转基因模型中的广谱是最小的哺乳动物可用。因此，我们在这里描述颅的小鼠模型中，通过lenticulostriatal动脉，即所谓的"凝模式"的电远端大脑中动脉永久凝固。在这个模型中所产生的梗塞位于主要在皮质;有关大脑体积的相对梗死体积对应于大多数人的中风。此外，该模型满足了上述的再现性和低死亡率的准则。在这段视频中，我们展示中风诱导中的"凝固模式"的手术方法和报告组织学和功能分析的工具。

由于短麻醉时间和中度脑损伤，转移到自己的笼子后约10分钟所有的动物都醒了，自由移动在笼子里，与同窝交互。在MCAO手术过程中的死亡率低于5％，主要是由于偶然的蛛网膜下腔出血或麻醉不正确的结果。在中风后，诱导7天的观察时间死亡只发生很少的动物约1-2％。在操作系列10动物对本报告所有的动物经受了这次手术和7天的观察期，他们都没有因为排除标准排除。 马华凝固后行为缺陷是由气瓶检验分析13前爪使用不对称性评估。在该试验中，独立的左和右前爪使用的比率的测量是在指定的时间点中风诱导后和相比得到24基线值MCAO（ 图3B）前小时。动物呈现在肢体不对称使用一个显著变化组合墙勘探缸测试24小时（1.72±0.326，P &lt;0.05），3天（1.36±0.17，P &lt;0.05），MCAO后。虽然在1周的观察时间的比例提高，电机的不对称仍然显著MCAO后第7天（1.35±0.29，P &lt;0.05）较基础值。 我们用甲酚紫染色串行冠状脑切片行程感应（ 图4B）后7天内进行梗死体积测量。平均梗死体积为15.4 立方毫米，从而代表一个脑半球（ 图4C）12％。此行程模式的变化是非常低约10％的标准偏差。病变区包括躯体感觉和运动皮质与皮质下结构只有轻微的感情。此外，梗死的定位区域是只有很少的变化高度可预测的，如图中示意分布图 （ 图4D）。 <img alt="图1" fo:content-width="5in" src="/files/ftp_upload/51729/51729fig1highres.jpg" width="500" /> 图1。Willis环的示意图。威利斯动脉圈是由大脑中动脉（MCA）和大脑前动脉（ACA），该分支从颈内动脉（ICA），以及通过形成大脑后动脉（PCA）和后交通动脉（PComA）。马华跑进外侧沟的地方转移到大脑皮层。占主导地位的马华支提供运动皮质和躯体感觉皮层的一部分的主要部分是永久的展示模型（ACOMA =前交通动脉闭塞; BA =基底动脉; SUCA =高级cerebel拉尔动脉）。 <img alt="图2" fo:content-width="3in" src="/files/ftp_upload/51729/51729fig2highres.jpg" width="400" /> 图2。切除颞肌与大脑中动脉闭塞的示意图后颅看法。 （一）去除颞肌后皮质动脉可以通过部分半透明鼠标头骨（8-12周龄小鼠）进行查看。占主导地位的马华支可以在时间视图的延髓部分，以及进一步皮质动脉从MCA和PCA在尾椎部分分支进行标识。（B）在其主要变化有分叉的主导马华支示意图在横向颞叶皮层（C）后钻出一个钻孔及移除头骨。黑色方块表示在分叉的近端和远端侧的MCA凝血位点。 图3。分析行为缺陷的。 （一）气缸测试设置:将鼠标放置到垂直缸和反射镜，以注册所有使用摄像机运动的背后放置（B）前肢使用不对称是使用气瓶检验分析。独立前肢使用左/右比率计算24小时MCAO前和中风诱导后指定的时间点。 N = 10，* P &lt;0.05，指示的时间点和基准线（控制）值之间。 <img alt="图4" fo:content-width="5in" src="/files/ftp_upload/51729/51729fig4highres.jpg" width="500" /> 图4。体积梗塞分析和梗塞结果远侧MCAO后。一个甲酚紫染色冠状胸罩（一）代表图像在第MCAO后7天。黄色的线是表示选择对侧（右）皮质和红线是选择同侧脑的非梗死（染色）皮质。面色苍白的区域在左侧半球描绘梗死组织区（B）梗死体积的10大脑分析（每个点代表一个人的大脑）24小时，3天，远端马华凝固后的7天内。水平红线代表平均值，误差线表示标准偏差（C）代表甲酚紫染色冠状脑切片马华凝固后，每400μm的第7天。（D）的梗死脑组织MCAO后第7天示意图分布。每张幻灯片在相对 ​​于前囟门给定的部分（:，由牛津大学出版社许可Liesz A 等人，Brain，2011修改后的图像从）描绘梗死分布（色码表示）累积的信息。 &lt;A HREF ="htt​​ps://jove.unicartagenaproxy.elogim.com/files/ftp_upload/51729/51729fig4highres.jpg"目标="_blank"&gt;请点击这里查看该图的放大版本。

Watch this Scientific Journal Video about Modeling Stroke in Mice: Permanent Coagulation of the Distal Middle Cerebral Artery at JoVE.com

Modeling Stroke in Mice: Permanent Coagulation of the Distal Middle Cerebral Artery

大脑中动脉闭塞（MCAO）的各种小鼠模型被广泛应用于实验性脑研究。这里，我们表明经颅永久远侧MCAO模型，产生对应于由多数人的缺血性中风的损害施加一个大小一致的皮层梗死。

造型中风小鼠:在远端大脑中动脉永久性混凝

Stroke is the third most common cause of death and a main cause of acquired adult disability in developed countries. Only very limited therapeutical ...

modeling-stroke-mice-permanent-coagulation-distal-middle-cerebral

Research

JoVE Journal

Medicine

36.9K 次观看.  University Hospital Munich. 大脑中动脉闭塞（MCAO）的各种小鼠模型被广泛应用于实验性脑研究。这里，我们表明经颅永久远侧MCAO模型，产生对应于由多数人的缺血性中风的损害施加一个大小一致的皮层梗死。 

视频: Modeling Stroke in Mice: Permanent Coagulation of the Distal Middle Cerebral Artery

Modeling Stroke in Mice - Middle Cerebral Artery Occlusion with the Filament Model

Stroke is among the most frequent causes of death and adult disability, especially in highly developed countries. However, treatment options to date are very limited. To meet the need for novel therapeutic approaches, experimental stroke research frequently employs rodent models of focal cerebral ischaemia. Most researchers use permanent or transient occlusion of the middle cerebral artery (MCA) in mice or rats.
Proximal occlusion of the middle cerebral artery (MCA) via the intraluminal suture technique (so called filament or suture model) is probably the most frequently used model in experimental stroke research. The intraluminal MCAO model offers the advantage of inducing reproducible transient or permanent ischaemia of the MCA territory in a relatively non-invasive manner. Intraluminal approaches interrupt the blood flow of the entire territory of this artery. Filament occlusion thus arrests flow proximal to the lenticulo-striate arteries, which supply the basal ganglia. Filament occlusion of the MCA results in reproducible lesions in the cortex and striatum and can be either permanent or transient. In contrast, models inducing distal (to the branching of the lenticulo-striate arteries) MCA occlusion typically spare the striatum and primarily involve the neocortex. In addition these models do require craniectomy. In the model demonstrated in this article, a silicon coated filament is introduced into the common carotid artery and advanced along the internal carotid artery into the Circle of Willis, where it blocks the origin of the middle cerebral artery. In patients, occlusions of the middle cerebral artery are among the most common causes of ischaemic stroke. Since varying ischemic intervals can be chosen freely in this model depending on the time point of reperfusion, ischaemic lesions with varying degrees of severity can be produced. Reperfusion by removal of the occluding filament at least partially models the restoration of blood flow after spontaneous or therapeutic (tPA) lysis of a thromboembolic clot in humans.
In this video we will present the basic technique as well as the major pitfalls and confounders which may limit the predictive value of this model.

Filamentous occlusion of the Middle cerebral artery is a common model for studying ischemic stroke in mice.

建模在小鼠体内的行程 - 中东与灯丝模型大脑中动脉闭塞

Stroke is among the most frequent causes of death and adult disability, especially in highly developed countries. However, treatment options to date are ...

科研

医学

Mouse Model of Middle Cerebral Artery Occlusion

Stroke is the most common fatal neurological disease in the United States 1. The majority of strokes (88%) result from blockage of blood vessels in the brain (ischemic stroke) 2. Since most ischemic strokes (~80%) occur in the territory of middle cerebral artery (MCA) 3, many animal stroke models that have been developed have focused on this artery. The intraluminal monofilament model of middle cerebral artery occlusion (MCAO) involves the insertion of a surgical filament into the external carotid artery and threading it forward into the internal carotid artery (ICA) until the tip occludes the origin of the MCA, resulting in a cessation of blood flow and subsequent brain infarction in the MCA territory 4. The technique can be used to model permanent or transient occlusion 5. If the suture is removed after a certain interval (30 min, 1 h, or 2 h), reperfusion is achieved (transient MCAO); if the filament is left in place (24 h) the procedure is suitable as a model of permanent MCAO. This technique does not require craniectomy, a neurosurgical procedure to remove a portion of skull, which may affect intracranial pressure and temperature 6. It has become the most frequently used method to mimic permanent and transient focal cerebral ischemia in rats and mice 7,8. To evaluate the extent of cerebral infarction, we stain brain slices with 2,3,5-triphenyltetrazolium chloride (TTC) to identify ischemic brain tissue 9. In this video, we demonstrate the MCAO method and the determination of infarct size by TTC staining.

We demonstrate in the video a method for producing a middle cerebral artery occlusion in adult mice using an intraluminal monofilament. We also show how to evaluate the extent of cerebral infarction by 2,3,5-triphenyltetrazolium chloride (TTC) staining.

大脑中动脉闭塞的小鼠模型

Stroke is the most common fatal neurological disease in the United States 1. The majority of strokes (88%) result from blockage of blood vessels in the ...

The Application Of Permanent Middle Cerebral Artery Ligation in the Mouse

Focal cerebral ischemia is among the most common type of stroke seen in patients. Due to the clinical significance there has been a prolonged effort to develop suitable animal models to study the events that unfold during ischemic insult. These techniques include transient or permanent, focal or global ischemia models using many different animal models, with the most common being rodents.
The permanent MCA ligation method which is also referred as pMCAo in the literature is used extensively as a focal ischemia model in rodents 1-6. This method was originally described for rats by Tamura et al. in 1981 7. In this protocol a craniotomy was used to access the MCA and the proximal regions were occluded by electrocoagulation. The infarcts involve mostly cortical and sometimes striatal regions depending on the location of the occlusion. This technique is now well established and used in many laboratories 8-13. Early use of this technique led to the definition and description of &#x201C;infarct core&#x201D; and &#x201C;penumbra&#x201D; 14-16, and it is often used to evaluate potential neuroprotective compounds 10, 12, 13, 17. Although the initial studies were performed in rats, permanent MCA ligation has been used successfully in mice with slight modifications 18-20 .
This model yields reproducible infarcts and increased post-survival rates. Approximately 80% of the ischemic strokes in humans happen in the MCA area 21 and thus this is a very relevant model for stroke studies. Currently, there is a paucity of effective treatments available to stroke patients, and thus there is a need for good models to test potential pharmacological compounds and evaluate physiological outcomes. This method can also be used for studying intracellular hypoxia response mechanisms in vivo. 
Here, we present the MCA ligation surgery in a C57/BL6 mouse. We describe the pre-surgical preparation, MCA ligation surgery and 2,3,5 Triphenyltetrazolium chloride (TTC) staining for quantification of infarct volumes.

Middle cerebral artery (MCA) ligation is a technique to study focal cerebral ischemia in animal models. In this method, the middle cerebral artery is exposed by craniotomy and ligated by cauterization. This method gives highly reproducible infarct volumes and increased post-operative survival rates compared to other methods available.

在小鼠永久性大脑中动脉结扎术中的应用

Focal cerebral ischemia is among the most common type of stroke seen in patients. Due to the clinical significance there has been a prolonged effort to ...

Induction of Ischemic Stroke and Ischemia-reperfusion in Mice Using the Middle Artery Occlusion Technique and Visualization of Infarct Area

Cerebrovascular disease is highly prevalent in the global population and encompasses several types of conditions, including stroke. To study the impact of stroke on tissue injury and to evaluate the effectiveness of therapeutic interventions, several experimental models in a variety of species were developed. They include complete global cerebral ischemia, incomplete global ischemia, focal cerebral ischemia, and multifocal cerebral ischemia. The model described in this protocol is based on the middle cerebral artery occlusion (MCAO) and is related to the focal ischemia category. This technique produces consistent focal ischemia in a strictly defined region of the hemisphere and is less invasive than other methods. The procedure described is performed on mice, given the availability of several genetic variants and the high number of tests standardized for mice to aid in the behavioral and neurodeficit evaluation.

We describe a mouse model of stroke induced by the occlusion of the middle cerebral artery using a silicone coated suture. The protocol can be applied to induce permanent occlusion or a temporary ischemia, followed by reperfusion.

缺血性中风和缺血再灌注小鼠的诱导使用梗塞区的中心动脉闭塞技术和可视化

Cerebrovascular disease is highly prevalent in the global population and encompasses several types of conditions, including stroke. To study the impact of ...

Middle Cerebral Artery Occlusion Allowing Reperfusion via Common Carotid Artery Repair in Mice

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue plasminogen activator (tPA) as an approved drug treatment to target ischemic strokes. Current research in the field of ischemic stroke focuses on better understanding the pathophysiology of stroke, to develop and investigate novel pharmaceutical targets. Reliable experimental stroke models are crucial for the progression of potential treatments. The middle cerebral artery occlusion (MCAO) model is clinically relevant and the most frequently used surgical model of ischemic stroke in rodents. However, the outcomes of this model, such as lesion volume, are associated with high levels of variability, particularly in mice. The alternative MCAO model described here allows the reperfusion of the common carotid artery (CCA) and the increased perfusion of the middle cerebral artery (MCA) territory, using a tissue pad with fibrinogen-based sealant to repair the vessel, and the improved welfare of the mice by avoiding external carotid artery (ECA) ligation. This reduces the reliance on the Circle of Willis, which is known to be highly anatomically variable in mice. Representative data show that using this alternative surgical approach decreases the variability in lesion volumes between the traditional MCAO approach and the alternative approach described here.

Intraluminal filament occlusion of the middle cerebral artery is the most frequently used in vivo model of experimental stroke in rodents. An alternative surgical approach to allow common carotid artery repair is performed here, which allows the reperfusion of the common carotid artery and a full reperfusion to the middle cerebral artery territory.

颅内动脉闭塞允许经普通颈动脉修复再灌注的研究

The ischemic stroke is a major cause of adult long-term disability and death worldwide. The current treatments available are limited, with only tissue ...

Modeling Stroke in Mice: Focal Cortical Lesions by Photothrombosis

Stroke is a leading cause of death and acquired adult disability in developed countries. Despite extensive investigation for novel therapeutic strategies, there remain limited therapeutic options for stroke patients. Therefore, more research is needed for pathophysiological pathways such as post-stroke inflammation, angiogenesis, neuronal plasticity, and regeneration. Given the inability of in vitro models to reproduce the complexity of the brain, experimental stroke models are essential for the analysis and subsequent evaluation of novel drug targets for these mechanisms. In addition, detailed standardized models for all procedures are urgently needed to overcome the so-called replication crisis. As an effort within the ImmunoStroke research consortium, a standardized photothrombotic mouse model using an intraperitoneal injection of Rose Bengal and the illumination of the intact skull with a 561 nm laser is described. This model allows the performance of stroke in mice with allocation to any cortical region of the brain without invasive surgery; thus, enabling the study of stroke in various areas of the brain. In this video, the surgical methods of stroke induction in the photothrombotic model along with&#160;histological analysis are demonstrated.

Described here is the photothrombotic stroke model, where a stroke is produced through the intact skull by inducing permanent microvascular occlusion using laser illumination after administration of a photosensitive dye.

小鼠模型中风:光血栓形成的焦皮质病变

Stroke is a leading cause of death and acquired adult disability in developed countries. Despite extensive investigation for novel therapeutic strategies, ...

神经科学

Modeling Stroke in Mice: Transient Middle Cerebral Artery Occlusion via the External Carotid Artery

Stroke is the third most common cause of mortality and the leading cause of acquired adult disability in developed countries. To date, therapeutic options are limited to a small proportion of stroke patients within the first hours after stroke. Novel therapeutic strategies are being extensively investigated, especially to prolong the therapeutic time window. These current investigations include the study of important pathophysiological pathways after stroke, such as post-stroke inflammation, angiogenesis, neuronal plasticity, and regeneration. Over the last decade, there has been increasing concern about the poor reproducibility of experimental results and scientific findings among independent research groups. To overcome the so-called &#8220;replication crisis&#8221;, detailed standardized models for all procedures are urgently needed. As an effort within the &#8220;ImmunoStroke&#8221; research consortium (https://immunostroke.de/), a standardized mouse model of transient middle cerebral artery occlusion (MCAo) is proposed. This model allows the complete restoration of the blood flow upon removal of the filament, simulating the therapeutic or spontaneous clot lysis that occurs in a large proportion of human strokes. The surgical procedure of this &#8220;filament&#8221; stroke model and tools for its functional analysis are demonstrated in the accompanying video.

Different models of middle cerebral artery occlusion (MCAo) are used in experimental stroke research. Here, an experimental stroke model of transient MCAo via the external carotid artery (ECA) is described, which aims to mimic human stroke, in which the cerebrovascular thrombus is removed due to spontaneous clot lysis or therapy.

模拟小鼠中风:通过颈外动脉的短暂性脑中动脉闭塞

Stroke is the third most common cause of mortality and the leading cause of acquired adult disability in developed countries. To date, therapeutic options ...

建立经颅大脑中动脉闭塞小鼠模型

A Modified Transcranial Middle Cerebral Artery Occlusion Model to Study Stroke Outcomes in Aged Mice

In experimental stroke research, middle cerebral artery occlusion (MCAO) with an intraluminal filament is widely used to model ischemic stroke in mice. The filament MCAO model typically exhibits a massive cerebral infarction in C57Bl/6 mice that sometimes includes brain tissue in the territory supplied by the posterior cerebral artery, which is largely due to a high incidence of posterior communicating artery atresia. This phenomenon is considered a major contributor to the high mortality rate observed in C57Bl/6 mice during long-term stroke recovery after filament MCAO. Thus, many chronic stroke studies exploit distal MCAO models. However, these models usually produce infarction only in the cortex area, and consequently, the assessment of post-stroke neurologic deficits could be a challenge. This study has established a modified transcranial MCAO model in which the MCA at the trunk is partially occluded either permanently or transiently via a small cranial window. Since the occlusion location is relatively proximal to the origin of the MCA, this model generates brain damage in both the cortex and striatum. Extensive characterization of this model has demonstrated an excellent long-term survival rate, even in aged mice, as well as readily detectable neurologic deficits. Therefore, the MCAO mouse model described here represents a valuable tool for experimental stroke research.

作者聚焦:中风研究的创新技术和未来方向 

This protocol demonstrates a unique mouse stroke model with a medium-sized infarct and an excellent survival rate. This model allows preclinical stroke researchers to extend the ischemia duration, use aged mice, and assess long-term functional outcomes.

改进的经颅大脑中动脉闭塞模型研究老年小鼠脑卒中结局

In experimental stroke research, middle cerebral artery occlusion (MCAO) with an intraluminal filament is widely used to model ischemic stroke in mice. ...

造型中风小鼠:在远端大脑中动脉永久性混凝

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此视频中的章节

建模在小鼠体内的行程 - 中东与灯丝模型大脑中动脉闭塞

大脑中动脉闭塞的小鼠模型

在小鼠永久性大脑中动脉结扎术中的应用

缺血性中风和缺血再灌注小鼠的诱导使用梗塞区的中心动脉闭塞技术和可视化

颅内动脉闭塞允许经普通颈动脉修复再灌注的研究

小鼠模型中风:光血栓形成的焦皮质病变

模拟小鼠中风:通过颈外动脉的短暂性脑中动脉闭塞

改进的经颅大脑中动脉闭塞模型研究老年小鼠脑卒中结局

改进的经颅大脑中动脉闭塞模型研究老年小鼠脑卒中结局

改进的经颅大脑中动脉闭塞模型研究老年小鼠脑卒中结局