This work was funded by the National Institute of Health, the National Heart Lung and Blood Institute (NIH and NHLBI; HL125572-01A1) and the LSUHSC-S start up fund to F.N.E. Gavins.

在视频报道该协议与实验得到LSUHSC-S机构动物护理和使用委员会批准并符合美国国立卫生研究院的指导方针。 注:雄性C57BL / 6小鼠重25 - 47克该研究中使用。小鼠保持与自由饮水标准州城颗粒饲料，在单独通风笼12小时光/暗周期下。该过程将使用无菌技术（ 例如 ，无菌手套，无菌仪器）在无菌条件下进行。 1.术前准备工作<ol><li>诱导用氯胺酮（150毫克/千克）和赛拉嗪（10毫克/千克）的组合麻醉腹膜内注射（IP）。一旦麻醉达到5分钟，每10分钟 - 步行掐最初每3个监测麻醉深度。虽然动物麻醉下，管理无菌眼软膏，以防止干燥。氯胺酮的初始剂量/甲苯噻嗪通常持续AB出30-40分钟。如果需要的话，这是由动物的响应于脚夹送验证额外剂量可以施用。 </li><li>放置小鼠上的温度调节的热垫仰卧位置并维持体温在36.5±0.1℃，这是使用直肠探针验证。 </li><li>剃颈部和消毒用70％的乙醇的皮肤。 </li></ol> 2. MCA闭塞（图1） <ol><li>使一个中线颈切口使用虹膜直剪刀和缩回（使用拉钩）软组织以暴露容器。 </li><li>使用杜蒙钳不破坏迷走神经周围组织解剖CCA和ECA。 </li><li>用6-0丝线绑围绕CCA一个松散的结，进行临时的缝合。 </li><li>使周围的ECA一个永久性缝合紧紧结扎血管（远端CCA分叉处），从它延伸较小的船只。 </li><li>做一个缝合AROUND非洲经委会近端CCA分叉。 </li><li>周围放置颈内动脉（ICA）和翼腭动脉（PPA）的一个微血管剪辑。 </li><li>使在ECA的小切口用微解剖弹簧剪刀，并插入一个180微米的硅尖的单丝。晋级尽量靠近永久缝合尽可能为灯丝的操作更加轻松。 </li><li>拧紧与周围插入长丝的ECA临时缝合，取下夹子微血管。 </li><li>切永久，远端缝合，并使用微解剖弹簧剪刀长丝的入口点之间的ECA。 </li><li>通过ICA引导丝，直到感到有阻力（约9 - 10微米超出了CCA分叉）在马华。 注:在事件感到太大的阻力，而多数长丝仍清晰可见，灯丝可能已进入PPA。如果发生这种情况，拉长丝回分叉处，轻轻脓ħ前进到ICA使用杜蒙钳，和推动长丝，直到它可以在ICA可视化。 </li></ol> 3.再灌注<ol><li>经过30分钟的接合期间，轻轻拉回来使用杜蒙钳拆下灯丝和保护周围的ECA的开口端缝合。 </li><li>通过仔细地松开使用杜蒙钳和血流通过CCA恢复了结扎删除周围的CCA临时缝合。 </li><li>用连续外科缝合关闭切口。皮肤的封闭可以通过连续或间断缝合来完成。皮肤缝合钉也是可以接受的方法。 </li><li>注射小鼠用1ml盐水皮下体积补充和与镇痛卡布洛芬（5毫克/公斤，SC），用于从手术过程，标志指示需要额外的疼痛缓解包括驼背，ungroomed涂层的疼痛和不适的缓解，活性降低，异常装腔作势，和食欲下降。 </li><li>观察整个从麻醉恢复小鼠中使用使用温度控制器调节的加热灯加热的30℃笼并放置捣碎饲料在培养皿上笼的地板上，以鼓励进食。小鼠将在再灌注期间被容纳每笼之一。 </li></ol> 4.假手术<ol><li>受试小鼠的相同步骤，而不单丝插入。 </li></ol> 5.术后神经分数（表1;图2） <ol><li>神经学评价采用18分制评估总适当的再灌注期后，小鼠;发动机;感觉;本体。较高的神经功能评分相当于减少神经功能。 注:如果判断反应迟钝，无法行走将小鼠安乐死。出于人道的安乐死其它的标准将包括的重量损失大于20％，呼吸二应力，和感染周围手术区。 在 CO 2室将用于安乐死和物理方法进行确认死亡将是颈椎脱位或开胸。 </li></ol> 6.脑梗死容积的测量（图3） <ol><li>诱导用氯胺酮（150毫克/千克）和赛拉嗪（10毫克/千克）由脚捏初始每3注入麻醉的IP监视器深度的组合麻醉 - 5分钟，每10分钟一次麻醉实现。 </li><li>放置小鼠仰卧位置并使用虹膜直剪刀从腹部到颈部随后腹膜切开皮肤。 </li><li>抬起使用杜蒙钳胸骨切排骨暴露心脏。 </li><li>打开左和使用两对止血的胸部右侧，露出心脏。 </li><li>插入连接于5mL注射器进入左心室一个＆G针和切割右心房。用室温生理盐水或磷灌注德缓冲盐水（PBS），直到流体变得清晰（通常为3 - 5分钟）。 </li><li>使用虹膜直剪刀和杜蒙钳取出头骨从大脑仔细了。 </li><li>切断嗅球和使用刀片使大脑将适合在基质小脑。使用此矩阵大脑切成偶数段。寒意矩阵使用前要保持组织凉爽。将脑入矩阵，放在冰上。 </li><li>切片的大脑使用两个刀片2毫米冠状段。使用开始从顶部2毫米的刀片进行首次降息，并在地方离开这个刀片。使切头背后另有2毫米。除去附着组织中的第一刀片。直到所有组织被切成重复此过程。 注:应该有4 - 完成后5段。 </li><li>放置分段成24孔板含2％2,3,5- triphenyltetrazalium氯（TTC），然后将其放置在一个浅水浴吨37℃20分钟。将每个段放入一个单独的井有助于保持他们在他们被切断的顺序。确保TTC解决方案完全覆盖组织片段。 10分钟后，把所有的片上。 </li><li>放置10％福尔马林的量小到一个新的24孔板的孔中，并在它们被切成顺序的片段转移到该板。 </li><li>扫描段到计算机并分析梗死大小使用ImageJ分析软件的全脑切片6的百分比（NIH 1.57图片软件）6。 <ol><li>轮廓梗塞面积以产生面积测量。下一个测量整个对侧半球，以确定该地区。通过对侧半球的区域划分梗塞面积再乘以100来确定梗死体积。 </li></ol></li></ol>

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The authors have nothing to disclose.

由于它的概念20年前，在MCAO模型对于涉及丝插入人体行程已在研究一个巨大的数字使用。这主要是由于它模拟什么在行程（ 即，缺血性中风）的最常见的形式的临床发生的事实。纹状体是缺血比大脑皮层更敏感，并且同样地，缺血时间长度将翻译成两个纹状体和背外侧皮层是否会受到影响，或者仅仅纹状体。既梗塞和再灌注时间可相应地改变，并且这提供了研究者能够研究短暂性脑缺血发...

中风是死亡的第五大原因遍及世界各地，死于该病一人每隔4分钟。超过80万美国人患有每年中风，这不仅是毁灭性的病人，也为他们的家庭。中风是成人残疾的主要原因，年支出估计为在$ 36.5十亿1的顺序，尽管很少有治疗方法是可用的。 组织纤溶酶原激活物（tPA）是唯一食品和药物管理局（FDA）的许可药物缺血性中风。然而，如果来自中风发作给予患者3-6小时内，并且在这些情况下，有利于仅4％的患者2是唯一有效的。因此，当务之急是中风的重现性好，临床相关动物模型被用于在潜在的治疗策略和治疗这种疾病的发展中提供帮助。要注意， 在体外是很重要</青霉&gt;模型，而在模拟的脑功能障碍的某些方面是有用的，不能够扼要发生在脑和外周中风后的复杂的生理的相互作用。因此， 在体内模型是必不可少的。 中风的最常见的类型是起源于缺血，约占总笔画的87％。其它笔划是脑内出血（9％）和蛛网膜下腔出血（4％），和最经常由栓子到大脑中动脉（MCA）引起的。这是由于在MCA的根部突出曲线，这导致进入大脑层血流成为打乱。在MCA来源于颈内动脉（ICA）和路线沿着横向沟，在那里它的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层的侧表面。 Willis环是由大脑后动脉被创建连接到脑动脉和后交通动脉。 MCAO的腔内长丝或缝合模型是最广泛使用于中风研究之一。但是，有几个不同的变化，以这种模式的，而这些都是根据微丝是否被插入到颈外动脉（ECA，称为龙格法）3，或者它是否被插入的ICA（称为小泉法）4。在小泉方法，在手术侧的总颈动脉（CCA）必须永久如果灯丝被去除以防止在CCA切口出血并列，而在龙格的方法是，必须永久地并列5 ECA 。在这里，因为我们认为这是一个远远优于和缺血性中风更临床相关的手术模式将要使用的方法龙格。此外，使用的硅尖单丝的，特别是与龙格的方法，产生了非常重现的MCAO相对于火焰钝化单丝，这往往产生不完全的闭塞和/或蛛网膜下腔出血6。 腔内长丝方法可作为永久或短暂闭塞4,6的模型。执行瞬态模型，灯丝一段局部缺血之后去除（ 例如，30分钟，60分钟，或2小时），再灌注被允许发生。该模型中，在一定程度上模拟血流后自发或治疗性干预（ 例如 ，tPA的给药）以裂解在人类中的血栓栓塞凝块的恢复。为永久模型，灯丝在适当位置简单地放置一段时间（ 例如 ，24小时），因此，不发生再灌注。腔内长丝方法的另一个优点是，开颅并不需要执行，从而可以原封不动颅骨和避免在颅内压力和温度的任何变化的事实。

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			<td height="22" style="height:22px;width:436px;">Male C57BL/6 mice</td>
			<td style="width:291px;">Jackson Laboratory, Bar Harbor, ME</td>
			<td style="width:397px;">#000664</td>
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			<td height="22" style="height:22px;width:436px;">Ketamine Hydrochloride</td>
			<td style="width:291px;">Morris &#38; Dickson, Shreveport, LA</td>
			<td>67457-108-10</td>
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			<td height="22" style="height:22px;width:436px;">Xylazine</td>
			<td style="width:291px;">Akorn, Inc, Lake Forest, IL</td>
			<td>NADA# 139-236</td>
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			<td height="22" style="height:22px;width:436px;">DC temperature control system</td>
			<td style="width:291px;">FHC, Bowdoin, ME</td>
			<td>40-90-8D</td>
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			<td height="22" style="height:22px;width:436px;">Mini rectal thermistor probe</td>
			<td style="width:291px;">FHC, Bowdoin, ME</td>
			<td>40-80-5D-02</td>
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			<td height="22" style="height:22px;width:436px;">Heating pad</td>
			<td style="width:291px;">FHC, Bowdoin, ME</td>
			<td>40-90-2-06</td>
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			<td style="width:291px;">Amazon, Bellevue, WA</td>
			<td>#64800</td>
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			<td height="22" style="height:22px;width:436px;">70% ethanol</td>
			<td style="width:291px;">Worldwide Medical Products, Bristol, PA</td>
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			<td height="22" style="height:22px;width:436px;">Dissecting microscope</td>
			<td style="width:291px;">Olympus, Center Valley, PA</td>
			<td>SZ40</td>
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			<td height="22" style="height:22px;width:436px;">Iris scissors (straight)</td>
			<td style="width:291px;">Fine Science Tools, Foster City, CA</td>
			<td>11251-20</td>
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			<td height="22" style="height:22px;width:436px;">Dumont forceps (45&#176; bent tip)</td>
			<td style="width:291px;">Fine Science Tools, Foster City, CA</td>
			<td>11297-00</td>
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			<td height="22" style="height:22px;width:436px;">Micro vessel clip</td>
			<td style="width:291px;">Fine Science Tools, Foster City, CA</td>
			<td>18055-05</td>
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			<td height="22" style="height:22px;width:436px;">Micro dissecting spring scissors (straight)</td>
			<td style="width:291px;">Fine Science Tools, Foster City, CA</td>
			<td>14088-10</td>
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			<td style="width:291px;">Fine Science Tools, Foster City, CA</td>
			<td>18200-11 (Helen used 17022-13)</td>
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			<td height="22" style="height:22px;width:436px;">Cotton tipped applicators</td>
			<td style="width:291px;">Fisher Scientific, Waltham, MA</td>
			<td>23-400-100</td>
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			<td height="22" style="height:22px;width:436px;">Gauze sponges</td>
			<td style="width:291px;">Covidien, Mansfield, MA</td>
			<td>#9023</td>
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			<td height="22" style="height:22px;width:436px;">6-0 silk braided surgical suture</td>
			<td style="width:291px;">Roboz, Gaithersburg, MD</td>
			<td>SUT-1073-11</td>
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			<td height="22" style="height:22px;width:436px;">0.9% sodium chloride</td>
			<td style="width:291px;">Morris &#38; Dickson, Lake Forest, IL</td>
			<td>0409-4888-20</td>
			<td></td>
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			<td height="22" style="height:22px;width:436px;">6-0 medium MCAO suture (silicon rubber coated monofilament)</td>
			<td style="width:291px;">Doccol Corporation, Sharon, MA</td>
			<td>6023PKRe</td>
			<td></td>
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			<td height="22" style="height:22px;width:436px;">Sofsilk 6-0 silicone coated braided silk</td>
			<td style="width:291px;">Covidien, Mansfield, MA</td>
			<td>SUT-14-1</td>
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			<td height="22" style="height:22px;width:436px;">Carprofen</td>
			<td style="width:291px;">Pfizer, New York, NY</td>
			<td>NADA# 141-199</td>
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			<td height="22" style="height:22px;width:436px;">Puralube</td>
			<td style="width:291px;">Dechra, Norwich, UK</td>
			<td>NDC 17033-211-38</td>
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			<td height="22" style="height:22px;width:436px;">Physitemp temperature controller</td>
			<td style="width:291px;">Harvard Apparatus, Holliston, MA</td>
			<td>TCAT-2AC</td>
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			<td height="22" style="height:22px;width:436px;">Heat lamp</td>
			<td style="width:291px;">Harvard Apparatus, Holliston, MA</td>
			<td>HL-1</td>
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			<td height="22" style="height:22px;width:436px;">Laser doppler probe</td>
			<td style="width:291px;">AD Instruments, Colorado Springs, CO</td>
			<td>MSP100XP</td>
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			<td style="width:291px;">Fisher Scientific, Waltham, MA</td>
			<td>#353226</td>
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			<td height="22" style="height:22px;width:436px;">Phosphate buffered saline (PBS)</td>
			<td style="width:291px;">Life Technologies, Carlsbad, CA</td>
			<td>20012-050</td>
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			<td height="22" style="height:22px;width:436px;">Single edge razor blades</td>
			<td style="width:291px;">Fisher Scientific, Waltham, MA</td>
			<td>12-640</td>
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			<td height="22" style="height:22px;width:436px;">2,3,5-triphenyltetrazalium chloride (TTC)</td>
			<td style="width:291px;">Sigma Aldrich, St. Louis, MO</td>
			<td>T8877-50G</td>
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			<td height="22" style="height:22px;width:436px;">Mouse brain matrix slicer</td>
			<td style="width:291px;">Braintree Scientific, Braintree, MA</td>
			<td>BS-A 5000C</td>
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			<td height="22" style="height:22px;width:436px;">Water bath</td>
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			<td>#182</td>
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			<td height="22" style="height:22px;width:436px;">10% formalin</td>
			<td style="width:291px;">Sigma Aldrich, St. Louis, MO</td>
			<td>HT501128-4L</td>
			<td></td>
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			<td height="22" style="height:22px;width:436px;">Image J analysis software</td>
			<td style="width:291px;">NIH, Bethesda, MD</td>
			<td>free download</td>
			<td></td>
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			<td height="22" style="height:22px;width:436px;">Retractor</td>
			<td style="width:291px;">Medical Device Purchase, Newcastle, CA</td>
			<td>MP-740</td>
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surgical approach for middle cerebral artery occlusion and reperfusion induced stroke in mice

中风是全世界死亡的首要原因，仍然是长期的残疾成年人的主要原因之一。约87％的中风是起源于缺血发生在大脑中动脉（MCA）的领土。目前唯一食品和药物管理局（FDA）批准的药物对这种破坏性疾病的治疗是组织纤溶酶原激活物（tPA）。然而，tPA的有管理一个小的治疗窗（3 - 6小时），且仅在究竟是谁收到患者的4％有效。目前的研究重点是了解中风的病理生理机制，以便找到潜在的治疗靶点。因此，可靠的模型是至关重要的，而MCA闭塞（MCAO）模型（也称为腔内丝或缝线模型）被认为是缺血性中风的最临床相关的手术模式，是相当非侵入性和容易复制。典型地，MCAO模型一起使用啮齿动物，尤其是由于小鼠所有可用的该物种的遗传变异。这里，我们描述（和存在于该视频）如何成功地执行小鼠MCAO模型（再灌注），以产生可靠和可重复的数据。

小鼠进行30分钟的MCAO引起的脑缺血（ 图1），随后一个周期再灌注的（24小时和1周在这里提出，但再灌注的长度可以变化）。缺血期间的死亡率是最小的（约2％）。交的缺血，死亡率（第24小时之内）为26％左右。 激光多普勒血流仪被用来确认在之前和MCAO /再灌注后MCA区血流灌注。 图4清楚地表明，当?...

Watch this Scientific Journal Video about Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice at JoVE.com

Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

为了了解中风的病理生理学，它使用可靠的模型是很重要的。本文将描述在小鼠中的最常用的中风模型中的一个，称为大脑中动脉闭塞（MCAO）模型（也称为腔内灯丝或缝合模型）再灌注。

对大脑中动脉闭塞再灌注损伤诱发中风小鼠的手术方法

Stroke is a leading cause of death worldwide and continues to be one of the major causes of long-term adult disabilities. About 87% of strokes are ...

surgical-approach-for-middle-cerebral-artery-occlusion-reperfusion

Research

JoVE Journal

Medicine

21.5K 次观看.  Louisiana State University. 为了了解中风的病理生理学，它使用可靠的模型是很重要的。本文将描述在小鼠中的最常用的中风模型中的一个，称为大脑中动脉闭塞（MCAO）模型（也称为腔内灯丝或缝合模型）再灌注。 

视频: Surgical Approach for Middle Cerebral Artery Occlusion and Reperfusion Induced Stroke in Mice

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 ...

Intraluminal Middle Cerebral Artery Occlusion (MCAO) Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Mice

Stroke is the third leading cause of death and the leading cause of disability in the world, with an estimated cost of near $70 billion 
in the United States in 20091,2. The intraluminal middle cerebral artery occlusion (MCAO) model was developed by Koizumi4 in 1986 to simulate this 
impactful human pathology in the rat. A modification of the MCAO method was later presented by Longa3. Both techniques have been widely used to identify 
molecular mechanisms of brain injury resulting from ischemic stroke and potential therapeutic modalities5. This relatively noninvasive method in rats has been 
extended to use in mice to take advantage of transgenic and knockout strains6,7. To model focal cerebral ischemia, an intraluminal suture is advanced via 
the internal carotid artery to occlude the base of the MCA. Retracting the suture after a specified period of time mimics spontaneous reperfusion, but the 
suture can also be permanently retained. This video will be demonstrating the two major approaches for performing intraluminal MCAO procedure in mice in a 
stepwise fashion, as well as providing insights for potential drawbacks and pitfalls. The ischemic brain tissue will subsequently be stained by 
2,3,5-triphenyltetrazolium chloride (TTC) to evaluate the extent of cerebral infarction8.

Intraluminal Middle Cerebral Artery Occlusion MCAO Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Mice

The intraluminal middle cerebral artery occlusion (MCAO) model is the most frequent used model among experimental ischemic stroke models. Here we will demonstrate the entire model in detail with the guide of Laser Doppler flowmetry, and its representative results.

腔内中东大脑中动脉闭塞（MCAO）缺血性中风模型在小鼠体内的激光多普勒血流指导

Stroke is the third leading cause of death and the leading cause of disability in the world, with an estimated cost of near $70 billion 
in the United ...

Bilateral Common Carotid Artery Occlusion as an Adequate Preconditioning Stimulus to Induce Early Ischemic Tolerance to Focal Cerebral Ischemia

There is accumulating evidence, that ischemic preconditioning - a non-damaging ischemic challenge to the brain - confers a transient protection to a subsequent damaging ischemic insult. We have established bilateral common carotid artery occlusion as a preconditioning stimulus to induce early ischemic tolerance to transient focal cerebral ischemia in C57Bl6/J mice. In this video, we will demonstrate the methodology used for this study.

There is accumulating evidence, that ischemic preconditioning (PC) &#x2013; a non-damaging ischemic challenge to the brain - confers a transient protection to a subsequent damaging ischemic insult. We established bilateral common carotid artery occlusion (BCCAO) as a preconditioning stimulus to induce early ischemic tolerance (IT) to transient focal cerebral ischemia (induced by middle cerebral artery occlusion, MCAO) in C57Bl6/J mice.

双侧颈总动脉闭塞足够的预处理刺激诱导局灶性脑缺血早期缺血耐受

There is accumulating evidence, that ischemic preconditioning - a non-damaging ischemic challenge to the brain - confers a transient protection to a ...

Endothelin-1 Induced Middle Cerebral Artery Occlusion Model for Ischemic Stroke with Laser Doppler Flowmetry Guidance in Rat

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in 20091, 2. Several models of cerebral ischemia have been developed to mimic the human condition of stroke. It has been suggested that up to 80% of all strokes result from ischemic damage in the middle cerebral artery (MCA) area3. In the early 1990s, endothelin-1 (ET-1) 4 was used to induce ischemia by applying it directly adjacent to the surface of the MCA after craniotomy. Later, this model was modified 5 by using a stereotaxic injection of ET-1 adjacent to the MCA to produce focal cerebral ischemia. The main advantages of this model include the ability to perform the procedure quickly, the ability to control artery constriction by altering the dose of ET-1 delivered, no need to manipulate the extracranial vessels supplying blood to the brain as well as gradual reperfusion rates that more closely mimics the reperfusion in humans5-7. On the other hand, the ET-1 model has disadvantages that include the need for a craniotomy, as well as higher variability in stroke volume8. This variability can be reduced with the use of laser Doppler flowmetry (LDF) to verify cerebral ischemia during ET-1 infusion. Factors that affect stroke variability include precision of infusion and the batch of the ET-1 used6. Another important consideration is that although reperfusion is a common occurrence in human stroke, the duration of occlusion for ET-1 induced MCAO may not closely mimic that of human stroke where many patients have partial reperfusion over a period of hours to days following occlusion9, 10. This protocol will describe in detail the ET-1 induced MCAO model for ischemic stroke in rats. It will also draw attention to special considerations and potential drawbacks throughout the procedure.

Several animal models of cerebral ischemia have been developed to simulate the human condition of stroke. This protocol describes the endothelin-1 (ET-1) induced middle cerebral artery occlusion (MCAO) model for ischemic stroke in rats. In addition, important considerations, advantages, and shortcomings of this model are discussed.

内皮素-1诱导大脑中动脉闭塞模型缺血性中风激光多普勒血流仪在大鼠指导

Stroke is the number one cause of disability and third leading cause of death in the world, costing an estimated $70 billion in the United States in ...

Utilizing a Cranial Window to Visualize the Middle Cerebral Artery During Endothelin-1 Induced Middle Cerebral Artery Occlusion

Creation of a cranial window is a method that allows direct visualization of structures on the cortical surface of the brain1-3. This technique can be performed in many locations overlying the rat cerebrum, but is most easily carried out by creating a craniectomy over the readily accessible frontal or parietal bones. Most frequently, we have used this technique in combination with the endothelin-1 middle cerebral artery occlusion model of ischemic stroke to quantify the changes in middle cerebral artery vessel diameter that occur with injection of endothelin-1 into the brain parenchyma adjacent to the proximal MCA4, 5. In order to visualize the proximal portion of the MCA during endothelin -1 induced MCAO, we use a technique to create a cranial window through the temporal bone on the lateral aspect of the rat skull (Figure 1). Cerebral arteries can be visualized either with the dura intact or with the dura incised and retracted. Most commonly, we leave the dura intact during visualization since endothelin-1 induced MCAO involves delivery of the vasoconstricting peptide into the brain parenchyma. This bypasses the need to incise the dura directly over the visualized vessels for drug delivery. This protocol will describe how to create a cranial window to visualize cerebral arteries in a step-wise fashion, as well as how to avoid many of the potential pitfalls pertaining to this method.

This article describes a method for visualizing rat cerebral arteries through a cranial window using temporal craniectomy in order to view proximal portions of the middle cerebral artery (Figure 1). This versatile method can be combined with various techniques of drug delivery to measure cerebral artery reactivity in vivo.

利用颅窗口，在可视化的大脑中动脉内皮素-1诱导大脑中动脉闭塞

Creation of a cranial window is a method that allows direct visualization of structures on the cortical surface of the brain1-3. This technique can be ...

Optimized System for Cerebral Perfusion Monitoring in the Rat Stroke Model of Intraluminal Middle Cerebral Artery Occlusion

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal models of acute ischemic stroke allows to confirm successful arterial occlusion and exclude subarachnoid hemorrhage. Cerebral perfusion monitoring can also be used to study intracranial collateral circulation, which is emerging as a powerful determinant of stroke outcome and a possible therapeutic target. Despite a recognized role of Laser Doppler perfusion monitoring as part of the current guidelines for experimental cerebral ischemia, a number of technical difficulties exist that limit its widespread use. One of the major issues is obtaining a secure and prolonged attachment of a deep-penetration Laser Doppler probe to the animal skull. In this video, we show our optimized system for cerebral perfusion monitoring during transient middle cerebral artery occlusion by intraluminal filament in the rat. We developed in-house a simple method to obtain a custom made holder for twin-fibre (deep-penetration) Laser Doppler probes, which allow multi-site monitoring if needed. A continuous and prolonged monitoring of cerebral perfusion could easily be obtained over the intact skull.

Cerebral perfusion monitoring has been demonstrated to improve accuracy in ischemic stroke models. Technical difficulties often limit the use of this essential tool for cerebrovascular research. In this video, an optimized system is shown to obtain a single or multi-site hemodynamic monitoring during intraluminal middle cerebral artery occlusion in rats.

优化系统的脑灌注监测在中风模型大鼠的大脑中动脉闭塞腔内

The translational potential of pre-clinical stroke research depends on the accuracy of experimental modeling. Cerebral perfusion monitoring in animal ...

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

Stroke is the third most common cause of death and a main cause of acquired adult disability in developed countries. Only very limited therapeutical options are available for a small proportion of stroke patients in the acute phase. Current research is intensively searching for novel therapeutic strategies and is increasingly focusing on the sub-acute and chronic phase after stroke because more patients might be eligible for therapeutic interventions in a prolonged time window. These delayed mechanisms include important pathophysiological pathways such as post-stroke inflammation, angiogenesis, neuronal plasticity and regeneration. In order to analyze these mechanisms and to subsequently evaluate novel drug targets, experimental stroke models with clinical relevance, low mortality and high reproducibility are sought after. Moreover, mice are the smallest mammals in which a focal stroke lesion can be induced and for which a broad spectrum of transgenic models are available. Therefore, we describe here the mouse model of transcranial, permanent coagulation of the middle cerebral artery via electrocoagulation distal of the lenticulostriatal arteries, the so-called &#8220;coagulation model&#8221;. The resulting infarct in this model is located mainly in the cortex; the relative infarct volume in relation to brain size corresponds to the majority of human strokes. Moreover, the model fulfills the above-mentioned criteria of reproducibility and low mortality. In this video we demonstrate the surgical methods of stroke induction in the &#8220;coagulation model&#8221; and report histological and functional analysis tools.

Various murine models of middle cerebral artery occlusion (MCAO) are widely used in experimental brain research. Here, we demonstrate the model of transcranial permanent distal MCAO which produces consistent cortical infarction of a size corresponding to damage imposed by the majority of human ischemic strokes.

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 ...

Embolic Middle Cerebral Artery Occlusion (MCAO) for Ischemic Stroke with Homologous Blood Clots in Rats

Clinically, thrombolytic therapy with use of recombinant tissue plasminogen activator (tPA) remains the most effective treatment for acute ischemic stroke. However, the use of tPA is limited by its narrow therapeutic window and by increased risk of hemorrhagic transformation. There is an urgent need to develop suitable stroke models to study new thrombolytic agents and strategies for treatment of ischemic stroke. At present, two major types of ischemic stroke models have been developed in rats and mice: intraluminal suture MCAO and embolic MCAO. Although MCAO models via the intraluminal suture technique have been widely used in mechanism-driven stroke research, these suture models do not mimic the clinical situation and are not suitable for thrombolytic studies. Among these models, the embolic MCAO model closely mimics human ischemic stroke and is suitable for preclinical investigation of thrombolytic therapy. This embolic model was first developed in rats by Overgaard et al.1 in 1992 and further characterized by Zhang et al. in 19972. Although embolic MCAO has gained increasing attention, there are technical problems faced by many laboratories. To meet increasing needs for thrombolytic research, we present a highly reproducible model of embolic MCAO in the rat, which can develop a predictable infarct volume within the MCA territory. In brief, a modified PE-50 tube is gently advanced from the external carotid artery (ECA) into the lumen of the internal carotid artery (ICA) until the tip of the catheter reaches the origin of the MCA. Through the catheter, a single homologous blood clot is placed at the origin of the MCA. To identify the success of MCA occlusion, regional cerebral blood flow was monitored, neurological deficits and infarct volumes were measured. The techniques presented in this paper should help investigators to overcome technical problems for establishing this model for stroke research.

Embolic Middle Cerebral Artery Occlusion MCAO for Ischemic Stroke with Homologous Blood Clots in Rats

In this paper, we demonstrate a standard method for producing an embolic middle cerebral artery occlusion with homologous blood clots (fibrin-rich) in adult rat. This model closely mimics human ischemic stroke and is suitable for preclinical study of thrombolytic therapy for ischemic stroke.

栓塞性脑梗塞（缺血）的大鼠缺血性中风与血同源凝块

Clinically, thrombolytic therapy with use of recombinant tissue plasminogen activator (tPA) remains the most effective treatment for acute ischemic ...

Quantification of Neurovascular Protection Following Repetitive Hypoxic Preconditioning and Transient Middle Cerebral Artery Occlusion in Mice

Experimental animal models of stroke are invaluable tools for understanding stroke pathology and developing more effective treatment strategies. A 2 week protocol for repetitive hypoxic preconditioning (RHP) induces long-term protection against central nervous system (CNS) injury in a mouse model of focal ischemic stroke. RHP consists of 9 stochastic exposures to hypoxia that vary in both duration (2 or 4 hr) and intensity (8% and 11% O2). RHP reduces infarct volumes, blood-brain barrier (BBB) disruption, and the post-stroke inflammatory response for weeks following the last exposure to hypoxia, suggesting a long-term induction of an endogenous CNS-protective phenotype. The methodology for the dual quantification of infarct volume and BBB disruption is effective in assessing neurovascular protection in mice with RHP or other putative neuroprotectants. Adult male Swiss Webster mice were preconditioned by RHP or duration-equivalent exposures to 21% O2 (i.e. room air). A 60 min transient middle cerebral artery occlusion (tMCAo) was induced 2 weeks following the last hypoxic exposure. Both the occlusion and reperfusion were confirmed by transcranial laser Doppler flowmetry. Twenty-two hr after reperfusion, Evans Blue (EB) was intravenously administered through a tail vein injection. 2 hr later, animals were sacrificed by isoflurane overdose and brain sections were stained with 2,3,5- triphenyltetrazolium chloride (TTC). Infarcts volumes were then quantified. Next, EB was extracted from the tissue over 48 hr to determine BBB disruption after tMCAo. In summary, RHP is a simple protocol that can be replicated, with minimal cost, to induce long-term endogenous neurovascular protection from stroke injury in mice, with the translational potential for other CNS-based and systemic pro-inflammatory disease states.

This protocol describes repetitive hypoxic preconditioning, or brief exposures to systemic hypoxia that reduce infarct volumes and blood-brain barrier disruption following transient middle cerebral artery occlusion in mice. It also details dual quantification of infarct volume and blood-brain barrier disruption after stroke to assess the efficacy of neurovascular protection.

神经血管保护定量继在小鼠重复缺氧预处理和瞬态大脑中动脉闭塞

Experimental animal models of stroke are invaluable tools for understanding stroke pathology and developing more effective treatment strategies. A 2 week ...