We would like to thank Dr. Lei Chen (Icahn School of Medicine at Mount Sinai, NY) who first established this model in our laboratory. Supported in part by HL126559, DA039576, MH098891, MH63022, MH072567, DA027569, and NSC 2015/17/B/NZ7/02985. Dr. Luc Bertrand is supported in part by a postdoctoral fellowship from the American Heart Association (16POST31170002).

所有的程序是由迈阿密机构动物护理和使用委员会（IACUC）的大学按照卫生研究院（NIH）的指导方针的国家机构的批准。需要无菌设备和无菌技术的使用。 1.准备缝合闭塞<ol><li>使用直径0.21毫米的缝合线20之间的小鼠 - 25克0.23毫米至25只小鼠 - 35克体重的。缝线的MCAO程序的类型的选择取决于动物的体重。 </li><li>使用银笔，标记从涂硅尖开始9毫米缝合。这将作为插入长度的导向。 </li></ol> 2.准备手术<ol><li>麻醉小鼠用异氟烷与氧混合，用实验室麻醉系统。在凌晨2商用机器上设置5和氧气流量使用异氟醚（见材料表 ）。动物转移到surgerŸ表面用鼻锥维持麻醉（异氟烷使用1.5设置 - 在2 2.5和氧气流量）。 <ol><li>确保鼠标的呼吸率约为1 - 每秒2呼吸喘气没有。此外，确保动物不会出现晶须刺激的反应和踏板反射（趾捏）。监控呼吸率和精力的手术过程中，至少每5分钟。 </li></ol></li><li>放眼用润滑剂上每只眼睛一滴使用无菌拭子，以防止它们在手术过程中进行干燥。可替代地，从一个无菌药物管适用眼药膏的眼球。 </li><li>翻转在它的后面的动物和刮胡子切口区域。之后，彻底消毒用70％的乙醇，接着洗必泰，并用70％乙醇的最终拭子的外科手术区域。放置在一个温暖的手术表面的动物立体显微镜下。 </li></ol> 3.颈总动脉夹层和内部/ EXTERNAL分支<ol><li>用手术剪和镊子，在颈部执行浅中线切口，从乳房骨上面的下颚下面（大约3 - 第4厘米）。手术单应放在周围的手术区域，以防止仪器从与非无菌表面接触。外科被单在当前视频省略以有助于成膜。 </li><li>使用镊子，仔细分离脂肪和结缔组织暴露气管。组织应该轻松，自然分开双方。 </li><li>放置在小鼠的颈背部枕头（圆形物体，约0.5厘米，直径），以延伸颈部，进一步暴露手术的区域。 </li><li>打开使用一个组织牵开器或钩切口。 </li><li>上气管的动物的左侧，小心tweeze相距结缔组织以暴露左颈总动脉（CCA）。要小心，不要损坏与CCA捆绑的神经和主脉。 </li><li>继续露出CCA，小心翼翼地从底层组织分离，并露出顶部"Y"分支内部的脑动脉（ICA）和外部脑动脉（ECA）的。要卸下CCA帮助，将CCA下弯钳刺穿结缔组织，然后慢慢让他们打开。 </li></ol> 4.制备CCA为缺血缝线插针<ol><li>插入的长度约4厘米的CCA下尼龙缝线的3段。确保CCA不扭曲，因为这将极大地缝合线的插入变得复杂。 </li><li>在最低点可能，使用永久结关闭底部缝合。 </li><li>领带就在ICA下方的顶缝/ ECA分支使用可移动的滑结。 </li><li>配合使用可移动滑结中间缝合，但保持敞开的。它离开足够的空间是重要的，因为不阻碍缝合插入。 </li><li>采用显微春天科幻ssors，在CCA进行底部和中部之间缝合切口。执行0.2毫米的切口接近底部缝合。 </li></ol> 5.大脑中动脉闭塞<ol><li>使用镊子，将缝线缺血进入CCA切口，引导其到顶部。 4.5步后迅速执行此步骤，以防止CCA开放的凝血和关闭。在缝合插入受阻的情况下，用钳提示重新打开切口。 </li><li>轻轻用滑结，以限制其周围血流阕中间缝合在MCAO缝合硅氧烷部分，但足够宽松以允许其自由移动。 </li><li>仔细撤消顶部缝合，确保了缺血缝合不滑出。 </li><li>通过几毫米的插入中的ICA的MCAO缝合，然后重新关闭的顶部缝合，以相同的方式如在步骤5.2中描述。 </li><li>引导缺血缝合到闭塞区域。成功插入指示从CCA切口少量血液回流的，而且银9毫米明显的标志是位于CCA切口和ICA / ECA分叉之间。可使用的监测方法如激光多普勒血流监测系统15，16来获得成功闭塞进一步确认。 注:在血流90％A下降至在闭塞区域侧中部脑区域表示成功闭塞。 </li><li>成功闭塞后，紧紧牵制中部和顶部缝合。如果执行假的插入，不重合缝合而是立即跳到步骤7.3。 </li></ol> 6.切口关闭和术后护理<ol><li>塔克在缝线到切口区域。 </li><li>删除拉钩或组织挂钩和枕头。 </li><li>使用无菌盐水和棉签清洁缝合区。 </li><li>关闭使用尼龙线/针和镊子的切口。 </li> &lt;利&gt;管理抗炎（ 例如 ，卡洛芬10毫克/千克，SC）和镇痛药（ 例如 ，丁丙诺啡0.1毫克/千克SC，每天两次）的药物，以减轻术后不适。放置在老鼠放在一个加热垫一个笼子里，以防止低温和给水软化食品广告阴唇访问。 </li><li>监测动物的恢复（5 - 10分钟）和中风的迹象。他们可以从轻度麻痹侧各不相同，盘旋对侧侧翼和滚动的剧烈收缩。几个评价观点中列出了讨论。显示呼吸窘迫或严重癫痫发作的迹象动物应实施安乐死。 </li><li>取决于缺血再灌注协议，从30分钟到120分钟或更离开代替MCAO缝合。减少阻塞时间，可以防止死亡。 </li><li>对于永久性闭塞，留在原地的缝合缺血24小时，但显著的死亡率是可以预期的。这段时间过去后，继续STE第7页。 <ol><li>如果使用永久性闭塞模型，养老鼠笼，直到终点。当时，去除缝线（步骤7.1到7.8）。这个过程也可以安乐死之后进行。 </li></ol></li></ol> 7.再灌注<ol><li>再次麻醉动物（按照2.1的说明），并删除创伤闭合缝线。 </li><li>使用镊子和组织分离器，重新打开切口暴露CCA。 </li><li>小心取出顶部缝合，直到涂硅部分位于中间结对缺血缝合轻轻一拉。 </li><li>重做在缝合顶端结防止血液流入通过缝合（它不必是一滑结）。 </li><li>仔细撤消中间结，拉动缝线过去结顶，但保持了CCA内。 </li><li>盖紧结顶阻断动脉血流。 </li><li>拉出完全缺血缝合然后拧紧中间结。 </li><li>在MCAO缝合可以是重复使用几次。在每次使用后，仔细清洁缝合在70％乙醇以除去使用无菌纱布的污染物（ 例如 ，血液或组织）。之后，放置在消毒袋缝合和消毒它。 </li><li>重复步骤6.1 - 6.4和监视动物的恢复。 </li></ol> 8.组织分析<ol><li>为了监测每搏输出量，剖析了大脑。 <ol><li>使用两步骤过程安乐死鼠标。首先，暴露动物异氟醚过量，直到呼吸停止。对于TTC染色，立即斩首鼠标进行脑部提取。脑切片（ 如用于免疫分析），通过前一个斩首心脏穿刺灌注执行。 </li><li>切断在颅底头和切断从颈部开口的头骨顶部的头皮到眼睛之间。与颅骨外露，切沿两侧从颅底开始开骨到眼窝。使沿着头骨的一部分，广本晋级。要小心，不要达到用剪刀剪，以免损坏大脑太深。 </li><li>此后，切两眼窝之间的骨头。 </li></ol></li><li>使用镊子，抬起头颅顶部，露出大脑。应该只有一个小的阻力。如果不轻松举起，头骨切口可能不完整。撬出轻轻用钝弯钳大脑。大脑仍可能由几个神经被连接。继续慢慢地，沿途移除附件。 </li><li>放置新鲜脑在1毫米大脑基质，加入一对的PBS滴，并沿着用剃刀刀片或低温恒温器叶片基质切片。 </li><li>脑转移到100mm培养皿，并添加染色溶液（2％2,3,5-氯化三苯四唑溶解在PBS [TTC]）以覆盖培养皿的底部。使用两个镊子，独立的所有脑切片（正面朝下），并将其放置，以便在培养皿的底部。 注:染色会变成活组织的红色，与非活组织剩余的白色。温热该溶液至37℃将加速染色。中风面积（白色）的症状可以迅速视为90分钟后中风且持续超过7天可见。成像应做连同一把尺子，使每搏输出量计算。 </li><li>过程解剖大脑用于切片，免疫染色，蛋白质分离，或各种其他程序17，18，19。 </li></ol>

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Authors have no competing or conflicts of interest.

所描述的MCAO方法的成功应用是高度依赖于脑血流解剖学的理解。由于缝合线的正确位置是难以辨别，由于缺乏直接的视觉线索，反复练习很重要，将其用于调查研究之前掌握的过程。行程容积应分析，以确保一致的结果。增加了一个激光多普勒系统可以帮助确定血流的成功闭塞和应定期使用，以确保过程正确进行。路由MCAO缝合到闭塞区域可以通过操纵动脉来促进。为了帮助在缝线引导至MCA，一?...

心血管疾病的研究中，如中风，依赖于使用的体内模型。理解局部缺血，药物毒性，和/或治疗的可能含义，有必要使用该疾病，从而使治疗组之间的比较研究的一个合适的，标准化的，可靠的和可再现的模型。在这个手稿中，我们使用的是老鼠，给予了大量的转基因小鼠和标准化的评估模型的有效性。耙分数来评估以下的实验缺血性卒中及以下恢复运动和行为障碍已经开发的1，2。 几个缺血性中风型号可供选择，如完整的全球脑缺血，不完全性缺血，多灶性脑缺血和脑缺血。后一组也是中风患者最常见的种类。大多数前夕NTS由栓塞或血栓闭塞的形成在或接近大脑中动脉（MCA）发起。鉴于这些参数，紧紧提出的模型模拟人类中风的疾病的病因，使获得高度相关的3个结果。尽管如此，从动物模型在人类疾病治疗的发现的翻译已被证明是具有挑战性的。截至目前，只有使用溶栓组织型纤溶酶原激活物已被批准用于治疗急性缺血性脑卒中4。 间在小鼠脑缺血，大脑后循环中风模型和脑静脉血栓形成模型的模型是高度侵入性，减少它们的适用性，并限制能够进行分析的范围。然而，其他技术如栓塞模型，photothrombosis模型，内皮素-1诱导的中风模型，和管腔内缝合大脑中动脉闭塞（MCAO）模型，可用于使用没有这些限制。在缺血模型在本协议中所述的技术。它提供了诱导可以容易地再灌注，并在高通量的方式进行局部脑缺血的可靠方法。有两种方法这一模式，即玉米，龙格和小泉的方法。它们在闭塞缝合插入脉管的方式略有不同。在玉米-隆加技术，缝线经由外颈动脉5插入。这里提出的技术是从在其中封闭缝合经由颈总动脉6插入小泉方法改性。 该MCAO模型已成功地应用于评估缺血性中风期间发生的不同事件。以下再灌注，脑水肿可以与血脑屏障的击穿可以观察到沿。山顶神经元死亡通常在24小时观察;然而，将其重新后7天7转动到基线水平。在人类中，性别和年龄的确定行程结果时，这也将在小鼠和大鼠8，9，10观察的重要变量。若干出版物已经使用了MCAO模型以证明处理效率11，12，13，14。

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			<td height="21" style="height:21px;width:276px;">MCAO suture 0.23mm</td>
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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.

为闭塞缝线的插入路径是表现在图1中 。在MCAO缝合要被路由到闭塞区域，在ICA中分叉。马华的成功闭塞会导致组织损伤，TTC染色可见。 图2呈现从假治疗的动物（ 图2A）和从60分钟缺血缺血再灌注动物染色的图像（染色，在90分钟或24小时后的闭塞， 图2B）。为了确定每搏量，首先计算行程区的每个部分中，使用包括在成像...

Watch this Scientific Journal Video about Induction of Ischemic Stroke and Ischemia-reperfusion in Mice Using the Middle Artery Occlusion Technique and Visualization of Infarct Area at JoVE.com

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

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 ...

induction-ischemic-stroke-ischemia-reperfusion-mice-using-middle

Research

JoVE Journal

Medicine

16.2K 次观看.  University of Miami. 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.

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

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 ...