目前的研究由美国国家研究院​​卫生部授予R01 - HL092188，R01 - DK083385和R01HL098294，麻醉教育及研究基金会（FAER）港灯资助，FAER授予JYD和东风集团（德意志研究联合会）研究的支持奖学金（GR2121/1-1）公司。

总论所有操作应执行下一个堂堂正正的解剖显微镜（Leica），并通过使用手术凝固剂。应在实验组小鼠在年龄和体重相匹配，以确保结果的可比性。在实验过程中，应该是稳定的，体温，血压，麻醉和流体管理和监测。 1。麻醉和手术<ol><li>肾缺血实验中，可以同时申请两个肾缺血再灌注后，或单肾切除术相结合，与其余肾缺血治疗。由于双边的两个肾脏缺血治疗是增加技术上的困难，更复杂的实验设置和较长的手术，我们将执行一个正确的悬重系统利用剩余的左肾治疗缺血性肾切除术。我们也利用这个吊重系统在心脏冠状动脉阻塞。但是，该过程是不同的，因为肾动脉，可以从周围组织中分离，而在于心脏肌肉的冠状动脉内。 </li><li>小鼠（8-14周龄，雌雄不限）将被麻醉的剂量70mg/kg戊巴比妥。我们将保持约麻醉。 10毫克/公斤/小时戊巴比妥钠。过量可显着降低血压，从而可以改变的结果。在我们最初的研究中，我们也使用氯氨酮和甲苯噻嗪。然而，这些化合物诱导强大的心动过缓（150 /分）。的观察心动过缓旁边也有证据表明，这两种化合物是在缺血的情况下保护机制所涉及的。挥发性麻醉剂不能使用或是由于他们的能力，也诱发肾脏保护和低血压。因此，我们最终选择麻醉的是苯巴比妥。此外，氯胺酮和甲苯噻嗪主要由肾脏排出体外。肾切除术和缺血可能延长氯胺酮麻醉。 </li><li>一旦疼痛反射缺席（镊子挤压脚趾测试）将被置于小鼠在加热温度控制表（RT，埃芬博格，德国慕尼黑，体温保持在37.0 ° C）在左边侧卧位置。 </li><li>用胶带将较低和上肢表。 </li><li>对于非生存的手术，如视频演示中，用矿物油覆盖切口区，以防止切割皮肤吸入鼠毛，从而防止鼠毛过敏的风险。在求生存实验的情况下，适当的无菌技术，包括裁剪的头发，皮肤消毒用碘伏（如优碘），蒸压仪器的无菌处理和使用口罩一个外科医生的无菌手套是必需的。生存手术中不使用矿物油。为了生存手术使用无菌氯化钠</li><li>用剪刀和凝血电极（ERBE，ICC50，德国），以防止肌肉和皮肤血管出血，右侧切口将执行。在通过中线开腹访问肾脏，这种技术不需要进入腹腔。 </li><li>使用4 / 0丝线缝合，右肾剪刀将被削减，而不会干扰与肾上腺皮质血管，包括肾动脉和肾静脉，肾蒂结扎。 </li><li>手术伤口是使用肌壁和皮肤用4 / 0缝线连续缝合两针封闭。 </li><li>动物，然后放置在右侧卧的位置和左翼切口将被执行。 </li><li>左肾小心取出的结缔组织，避免肾上腺和船只。 </li><li>接下来，打开肾脏腹侧下降到一个有机玻璃杯。有机玻璃杯应消毒的生存手术。与周围组织的一个钳，肾极小心推入有机玻璃杯。此后，极是仔细有机玻璃杯推到一个镊子。肾脏将保持潮湿和温暖，用湿棉签用清水浸泡在37.0 ° C。 </li><li>肾者，暴露左肾和安置到一个有机玻璃杯克制腹侧肾血管紧张。一个足够的限制是很重要的一个肾动脉下成功就业的缝合。此后，肾脏的人是固定的，可以很容易地确定肾动脉，肾静脉上运行。 </li><li>肾动脉解剖从​​邻近组织，从腹主动脉用细镊子起飞。从最高层的一个弯钳的基本静脉动脉剥离。 </li><li>被放置在肾动脉，通过一个8 / 0丝线缝合（爱惜康，日本东京，德国）作为一个领先的指南钳。这种技术允许在不中断只有动脉血流肾肾静脉受压，较肾蒂夹紧。 </li><li>缝合是放置在一个小杆（可事先准备好的安全别针），重量约。 1G是连接到每个（Eppendorf管）。肾脏大小或添加到Eppendorf管中的水的重量，可适应。然而，动脉应稍微拉向上，使没有血液通过动脉，但没有对动脉的主要舒展运行。主要拉伸应力对动脉可引起动脉壁的增厚和减少再灌注血管壁损伤。 </li><li>虽然重量是不受支持的，立即肾动脉闭塞。相反，当的权重支持，立即恢复血流的器官。成功闭塞证实的颜色从红色变为白色。 </li><li>不同缺血时间可以进行实验的要求。一个完整的再灌注缺血时间的长短取决于将发生。缺血时间长达45分钟让可收回肾损伤。在我们的实验设置中，我们用20高达30分钟，缺血与一个中等的肾损伤，这让我们看到了改善，以及根据小鼠品系和治疗的肾脏损害减值。 </li><li>完成再灌注后肾脏可以更换的有机玻璃杯。因此肾脏的持有人是不固定的，对肾脏的紧张被释放。颅极轻轻的杯子中剔除出去，不愠不火的镊子和尾杆紧随。 </li><li>可以释放鼠标的固定（磁带）和鼠标就可以放置在仰卧姿势如FITC -菊粉标记间隙。 </li><li>在实验的子集，血压和心率，可以记录整个手术过程。血压测量得到不断通过颈内动脉导管。总之，颈内动脉通过钝性分离气管肌肉暴露。经过进一步的接触和小心避免组织创伤（尤其是迷走神经），导管插入使用两个缝线和一个小钳的船只。测量血压是一个statham元素（280 WK，王氏港建，Kaltbrunn，瑞士）。 </li><li>对于FITC标记的菊粉间隙导管将放在左侧颈内静脉滴注氯化钠和FITC标记的菊粉。此外，膀胱导管插入尿液收集在一个随时间变化的的方式将在下面详细解释。 </li></ol> 2。肾损伤的测量<ol><li>一般建议:下面的结果参数，以确定肾损伤，肾缺血: 执行FITC标记的菊粉的间隙，以评估肾小球滤过率（GFR）。作为替代方案确定通过代谢笼调查24小时，肾功能损伤后的肌酐清除率和血清肌酐。执行肾H＆E染色和一个过氧化物酶（MPO）ELISA法，以确定肾功能损害和炎症24小时，肾缺血。 </li><li>代谢笼调查: <ol><li>左肾缺血转身进入腹膜后的位置，如上所述，伤口闭合。 </li><li>小鼠在手术结束时，收到0.3毫升生理盐水IP和下加热灯2小时的恢复。 </li><li>此后，老鼠将被放入代谢笼（Tecniplast德国，Hohenpeissenberg，德国），肾功能参数测定24小时的2倍。 </li><li>每只小鼠的取水口已确定称重水空和填充瓶供应前鼠标比24小时后再次称重满瓶。鼠标的重量以及待定。 </li><li> 24小时后约75μl血从眼睛后方的静脉丛通过短isoflorane麻醉下的毛细管。 </li><li>通过测量肾使用市售的colorimtric方法，根据制造商的协议（LT - SYS的，劳动+技术公司，德国柏林）缺血后血浆和尿肌酐24小时的肾功能评估。血浆和尿钠 ，钾浓度+决心与火焰发射光度计（6361 ELEX的Eppendorf公司，汉堡，德国）。肾脏的排泄和血液动力学的价值观是使用 15-17标准的公式计算。 </li></ol></li><li>菊粉间隙:FITC -菊粉清除实验将执行以下肾缺血。 <ol><li>左侧颈内静脉插管连续的Infusion。解剖后从邻近组织的静脉用一备剪刀（杜蒙，WPI），放置两个缝线周围静脉（4 / 0）。放置之前，阻止血液流向proximale缝合静脉钳。此后，关闭远端结和重量放在远端缝合结束应变静脉紧张。静脉导管，这是非常重要的。使用微型剪刀切一个小口进入静脉。保持开放与罚款钳（杜蒙，WPI），并放入静脉导管。为了推进导管钳断。确保没有导管的张力，让你可以无需拿着它下来。随后关闭近端缝合静脉导管在第二。 </li><li>导管放置在定时收集尿液的膀胱。因此，你切一个小切口，在皮肤，膀胱。通过皮肤孔，小心地取出膀胱。用湿纸巾修复膀胱。放置在膀胱（4 / 0）准备缝合。 </li><li>剪下一小段导管（1.5厘米）结束的热量，这使得它更容易在膀胱修复导管周围的一环。 </li><li>削减其极顶一个孔进入膀胱。请削减之间的船只，以防止出血。插入导管进入膀胱的大底和关闭通过导管和膀胱的缝合。 </li><li>所有的老鼠收到0.85％氯化钠溶液中静脉推注，金额相等于体重的20％。连续输液维持在800ul/h/25g体重和FITC标记的菊粉是评价整个肾脏肾小球滤过率（GFR）的输液率。 20分钟的动物稳定后，20分钟定时尿液集合进行测定尿流率和FITC -菊粉（四次）。血液是在每测量FITC -菊粉期间中获得。 </li><li>当开始20分钟收集期间使用一个计时器。收集尿液，由一个小的Eppendorf管放入膀胱导管结束。测量前后收集尿液，使你能确定尿量管重量。 retroorbital血液收集的形式，通过在每个收集期中的毛细管静脉丛通过采取血液样本。 </li></ol></li><li>后进一步分析，如MPO ELISA法，细胞因子的EISA，组织学的每个程序，可采集血液和肾脏。 </li></ol> 3。代表性的成果: <img src="/files/ftp_upload/2549/2549fig1.jpg" alt="图1"/> 图1吊重系统为肾缺血模型:一个悬重系统已经建立，导致左肾肾缺血。因此8.0尼龙缝线划归肾动脉和一个小，重量轻（1G）附加在每个缝合结束。这个实验设计允许申请和释放的重量负荷可逆的肾动脉闭塞。

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综上所述，本研究介绍了一种在小鼠模型使用一个吊重系统为肾动脉闭塞的肾缺血的新技术。事实上，这项研究表明高度重复性的伤害，从而最大限度地减少钳夹肾蒂的变异。

<table border="1"><tbody><tr><th>试剂名称</th><th>公司</th><th>目录编号</th><th>评论</th></tr><tr><td>戊巴比妥钠</td><td> Vortech药业LS，有限公司</td><td> VPL 9372 </td><td> 4mg/mL在生理盐水</td></tr><tr><td>缝合，丝绸4.0 </td><td>哈佛器械</td><td> 517698 </td><td></td></tr><tr><td>缝合后，普理灵8.0 </td><td>美国爱惜康， </td><td> M8739 </td><td>可重复使用</td></tr><tr><td>解剖显微镜</td><td>徕卡</td><td> N / A </td><td>考虑慷慨的工作距离</td></tr><tr><td>暖气表</td><td> RT，Effenberger，德国</td><td> N / A </td><td>只和单一供应商</td></tr><tr><td>手术器械</td><td> WPI的，杜蒙</td><td></td><td>罚款微血管剪刀</td></tr></tbody></table>

use of a hanging-weight system for isolated renal artery occlusion

在住院的病人中，超过50％的急性肾损伤（AKI）的情况下引起的肾缺血1-3。一个住院的病人最近的一项研究显示，只有血清肌酐水平轻度增加（0.3〜0.4毫克/升）大于70％，在没有任何增加 1人死亡风险。沿着这些路线，手术过程中，需要交叉夹紧的主动脉和肾血管与肾功能衰竭率高达30％，4 。同样，超过10％的患者在正常情况下，心脏手术后AKI的发生与死亡率大幅增加有关。 AKI的是肝移植术后常见的并发症。至少有8-17％的患者最终需要肾脏替代治疗5。此外，由于肾小管细胞损伤在肾移植肾功能延迟是经常与缺血相关AKI的6。此外，AKI的发生在约20％的患者患败血症 6。AKI的发生是与急剧增加的发病率和死亡率 1关联。治疗方法是非常有限的，多数在AKI的介入试验，在人类的失败。因此，迫切需要额外的治疗方式，以防止缺血肾损伤3，7-9。 为了阐明肾损伤的机制因缺血和小鼠模型可能的治疗策略是集中需要7-13。小鼠模型提供利用不同的遗传模型，包括基因靶向小鼠和组织特异性的基因靶向小鼠（CRE - flox系统）的可能性。然而，小鼠肾缺血是在技术上具有挑战性和实验细节显着影响的结果。我们进行了系统评价的一种新型模式，为分离出小鼠肾动脉闭塞，特别是避免使用夹紧或缝合肾蒂 14 。这种模式需要一个肾切除右肾，因为缺血，可只有一个肾，由于实验设置执行。事实上，通过使用吊重系统，肾动脉只有仪表曾经在整个手术过程。此外，无静脉或尿道梗阻发生这种技术。通过测量血清肌酐，我们能证明与缺血时间剂量依赖性和高度重复性的肾损伤。此外，缺血预处理与传统的夹紧整个蒂，肾保护这种新的模式进行比较时，，是更深刻，更可靠。因此，他的新技术可能会在急性肾损伤领域工作的其他研究人员的有用。

Watch this Scientific Journal Video about Use of a Hanging-weight System for Isolated Renal Artery Occlusion at JoVE.com

Use of a Hanging-weight System for Isolated Renal Artery Occlusion

因缺血而精确的小鼠模型为急性肾损伤（AKI）是一个重要的工具，调查急性肾损伤，并有可能找到治疗肾损伤的治疗手段。悬重系统提供了一个即时和可靠的肾动脉闭塞再灌注的工具，没有引起肾充血。

一个悬重系统使用隔离肾动脉闭塞

In hospitalized patients, over 50% of cases of acute kidney injury (AKI) are caused by renal ischemia 1-3. A recent study of hospitalized patients ...

use-of-a-hanging-weight-system-for-isolated-renal-artery-occlusion

Nanyang Technological University

Research

JoVE Journal

Medicine

15.9K 次观看.  University of Colorado. 因缺血而精确的小鼠模型为急性肾损伤（AKI）是一个重要的工具，调查急性肾损伤，并有可能找到治疗肾损伤的治疗手段。悬重系统提供了一个即时和可靠的肾动脉闭塞再灌注的工具，没有引起肾充血。

视频: Use of a Hanging-weight System for Isolated Renal Artery Occlusion

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

Use of a Hanging Weight System for Coronary Artery Occlusion in Mice

Murine studies of acute injury are an area of intense investigation, as knockout mice for different genes are becoming increasingly available 1-38. Cardioprotection by ischemic preconditioning (IP) remains an area of intense investigation. To further elucidate its molecular basis, the use of knockout mouse studies is particularly important 7, 14, 30, 39. Despite the fact that previous studies have already successfully performed cardiac ischemia and reperfusion in mice, this model is technically very challenging. Particularly, visual identification of the coronary artery, placement of the suture around the vessel and coronary occlusion by tying off the vessel with a supported knot is technically difficult. In addition, re-opening the knot for intermittent reperfusion of the coronary artery during IP without causing surgical trauma adds additional challenge. Moreover, if the knot is not tied down strong enough, inadvertent reperfusion due to imperfect occlusion of the coronary may affect the results. In fact, this can easily occur due to the movement of the beating heart.
Based on potential problems associated with using a knotted coronary occlusion system, we adopted a previously published model of chronic cardiomyopathy based on a hanging weight system for intermittent coronary artery occlusion during IP 39. In fact, coronary artery occlusion can thus be achieved without having to occlude the coronary by a knot. Moreover, reperfusion of the vessel can be easily achieved by supporting the hanging weights which are in a remote localization from cardiac tissues.
We tested this system systematically, including variation of ischemia and reperfusion times, preconditioning regiments, body temperature and genetic backgrounds39. In addition to infarct staining, we tested cardiac troponin I (cTnI)
as a marker of myocardial infarction in this model. In fact, plasma levels of cTnI correlated with infarct sizes (R2=0.8). Finally, we could show in several studies that this technique yields highly reproducible infarct sizes during murine IP and myocardial infarction6, 8, 30, 40, 41. Therefore, this technique may be helpful for researchers who pursue molecular mechanisms involved in cardioprotection by IP using a genetic approach in mice with targeted gene deletion. Further studies on cardiac IP using transgenic mice may consider this technique.

A murine model for myocardial ischemia and ischemic preconditioning is an important tool study cardioprotective mechanisms in vivo. Here, we report an easy applicable in situ model for cardiac IP using a hanging-weight system for coronary artery occlusion.

悬重系统的使用在小鼠冠状动脉闭塞

Murine studies of acute injury are an area of intense investigation, as knockout mice for different genes are becoming increasingly available 1-38. ...

Use of a Hanging-weight System for Liver Ischemia in Mice

Acute liver injury due to ischemia can occur during several clinical procedures e.g. liver transplantation, hepatic tumor resection or trauma repair and can result in liver failure which has a high mortality rate1-2. Therefore murine studies of hepatic ischemia have become an important field of research by providing the opportunity to utilize pharmacological and genetic studies3-9. Specifically, conditional mice with tissue specific deletion of a gene (cre, flox system) provide insights into the role of proteins in particular tissues10-13 . Because of the technical difficulty associated with manually clamping the portal triad in mice, we performed a systematic evaluation using a hanging-weight system for portal triad occlusion which has been previously described3. By using a hanging-weight system we place a suture around the left branch of the portal triad without causing any damage to the hepatic lobes, since also the finest clamps available can cause hepatic tissue damage because of the close location of liver tissue to the vessels. Furthermore, the right branch of the hepatic triad is still perfused thus no intestinal congestion occurs with this technique as blood flow to the right hepatic lobes is preserved. Furthermore, the portal triad is only manipulated once throughout the entire surgical procedure. As a result, procedures like pre-conditioning, with short times of ischemia and reperfusion, can be easily performed. Systematic evaluation of this model by performing different ischemia and reperfusion times revealed a close correlation of hepatic ischemia time with liver damage as measured by alanine (ALT) and aspartate (AST) aminotransferase serum levels3,9. Taken together, these studies confirm highly reproducible liver injury when using the hanging-weight system for hepatic ischemia and intermittent reperfusion. Thus, this technique might be useful for other investigators interested in liver ischemia studies in mice. Therefore the video clip provides a detailed step-by-step description of this technique.

We established a novel murine model of a hanging weight system for portal triad occlusion. This technique may be useful for future investigations of ischemia in murine hepatic models.

使用吊重系统在小鼠肝缺血

Acute liver injury due to ischemia can occur during several clinical procedures e.g. liver transplantation, hepatic tumor resection or trauma repair and ...

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 ...

Use of Animal Model of Sepsis to Evaluate Novel Herbal Therapies

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several techniques, including infusion of exogenous bacterial toxin (endotoxemia) or bacteria (bacteremia), as well as surgical perforation of the cecum by cecal ligation and puncture (CLP)1-3. CLP allows bacteria spillage and fecal contamination of the peritoneal cavity, mimicking the human clinical disease of perforated appendicitis or diverticulitis. The severity of sepsis, as reflected by the eventual mortality rates, can be controlled surgically by varying the size of the needle used for cecal puncture2. In animals, CLP induces similar, biphasic hemodynamic cardiovascular, metabolic, and immunological responses as observed during the clinical course of human sepsis3. Thus, the CLP model is considered as one of the most clinically relevant models for experimental sepsis1-3.
Various animal models have been used to elucidate the intricate mechanisms underlying the pathogenesis of experimental sepsis. The lethal consequence of sepsis is attributable partly to an excessive accumulation of early cytokines (such as TNF, IL-1 and IFN-&gamma;)4-6 and late proinflammatory mediators (e.g., HMGB1)7. Compared with early proinflammatory cytokines, late-acting mediators have a wider therapeutic window for clinical applications. For instance, delayed administration of HMGB1-neutralizing antibodies beginning 24 hours after CLP, still rescued mice from lethality8,9, establishing HMGB1 as a late mediator of lethal sepsis. The discovery of HMGB1 as a late-acting mediator has initiated a new field of investigation for the development of sepsis therapies using Traditional Chinese Herbal Medicine. In this paper, we describe a procedure of CLP-induced sepsis, and its usage in screening herbal medicine for HMGB1-targeting therapies.

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection, and can be simulated by a surgical technique termed cecal ligation and puncture (CLP). Here we describe a method to use CLP-induced animal model to screen medicinal herbs for therapeutic agents.

脓毒症动物模型，用于评估新型草药疗法

Sepsis refers to a systemic inflammatory response syndrome resulting from a microbial infection. It has been routinely simulated in animals by several ...

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 ...

An Isolated Working Heart System for Large Animal Models

Since its introduction in the late 19th century, the Langendorff isolated heart perfusion apparatus, and the subsequent development of the working heart model, have been invaluable tools for studying cardiovascular function and disease1-15. Although the Langendorff heart preparation can be used for any mammalian heart, most studies involving this apparatus use small animal models (e.g., mouse, rat, and rabbit) due to the increased complexity of systems for larger mammals1,3,11. One major difficulty is ensuring a constant coronary perfusion pressure over a range of different heart sizes &#8211; a key component of any experiment utilizing this device1,11. By replacing the classic hydrostatic afterload column with a centrifugal pump, the Langendorff working heart apparatus described below allows for easy adjustment and tight regulation of perfusion pressures, meaning the same set-up can be used for various species or heart sizes. Furthermore, this configuration can also seamlessly switch between constant pressure or constant flow during reperfusion, depending on the user&#8217;s preferences. The open nature of this setup, despite making temperature regulation more difficult than other designs, allows for easy collection of effluent and ventricular pressure-volume data.

Most studies involving the Langendorff apparatus use small animal models due to the increased complexity of systems for larger mammals. We describe a Langendorff system for large animal models that allows for use across a range of species, including humans, and relatively easy data acquisition.

一个孤立的工作心脏系统的大型动物模型

Since its introduction in the late 19th century, the Langendorff isolated heart perfusion apparatus, and the subsequent development of the working heart ...

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

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 ischemic in origin and occur in the territory of the middle cerebral artery (MCA). Currently the only Food and Drug Administration (FDA) approved drug for the treatment of this devastating disease is tissue plasminogen activator (tPA). However, tPA has a small therapeutic window for administration (3 - 6 hr), and is only effective in 4% of the patients who actually receive it. Current research focuses on understanding the pathophysiology of stroke in order to find potential therapeutic targets. Thus, reliable models are crucial, and the MCA occlusion (MCAo) model (also termed the intraluminal filament or suture model) is deemed to be the most clinically relevant surgical model of ischemic stroke, and is fairly non-invasive and easily reproducible. Typically the MCAo model is used with rodents, especially with mice due to all the genetic variations available for this species. Here we describe (and present in the video) how to successfully perform the MCAo model (with reperfusion) in mice to generate reliable and reproducible data.

In order to understand the pathophysiology of stroke, it is important to use reliable models. This paper will describe one of the most frequently used stroke models in mice, termed the middle cerebral artery occlusion (MCAo) model (also termed the intraluminal filament or suture model) with reperfusion.

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 ...

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 ...