Name: an improved method for rapid intubation of the trachea in mice
Uploaded: 2016-02-22T14:00:00.000+00:00
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Description: Watch this Scientific Journal Video about An Improved Method for Rapid Intubation of the Trachea in Mice at JoVE.com

作者感谢组织学的布赖恩·约翰逊和影像核心在华盛顿大学与三色染色和分析的帮助。这项工作是由美国国立卫生研究院资助HL098067和HL089455支持。

机构的动物护理和使用委员会（IACUC）在华盛顿和雪松 - 西奈医学中心大学的必要批准这些研究动物的工作。 1.准备<ol><li>既消毒钝端钳和降压通过高压灭菌。 </li><li>使用生物安全柜，从冻干粉末制备BLM的工作原液在PBS中。超声处理10分钟，该溶液在35千赫到保证均匀混合。 注:建议的30和45微升之间的总体积，以防止与较大体积在低端移液变异，和窒息。 </li><li>准备一个干净的工作区，其包括用于该过程本身约1米2，以及两个前和后的程序笼指定地点。 </li><li>通过在底座和underl铺设2或3条实验室胶带固定的程序董事会立即在研究者面前的板凳基地颖替补。 图 1所示为创造一个板进一步规范。 </li><li>领带的过程板的两个定位螺丝之间尺寸4.0缝合线的单根长度。 </li><li>产生通过拆卸和从三个1毫升注射器丢弃柱塞，以及沉积60微升PBS中到每个筒的顶部上以形成气密密封的一个临时的肺活量计。固定导管的枢纽松散到注射器之一，它放置在电路板的一侧。 </li><li>吸300微升空气进入1毫升的注射器，并将其放置在电路板的一侧。 </li><li>切割胶带在长度和地点约6英寸的附加片一侧。这将被用于动物固定在板步骤2.4。 </li><li>设置一个异氟醚室。附加O 2，异氟醚，真空到相应的端口上都曝光室和真空间隙。另外，施用麻醉剂在异氟醚兼容的生物安全柜。 </li></ol> 2.插管<ol><li>麻醉异氟烷鼠标在室内，直到它失去意识，呼吸减慢，以适当的速度。典型的曝光包括3至4分钟的4％的异氟醚和2％O 2，和理想的结果是2〜2.5分钟的镇静。这对应于1呼吸每2秒的呼吸速率。 </li><li>在等待镇静设置在，吸BLM的30和45微升之间成移液器和地点到一侧。 </li><li>当准备好后，从连接于程序平台的定位螺钉的螺纹暂停其上门牙的镇静鼠标。确保动物的背部靠在台表面平坦。 </li><li>小心不要限制通风，放置一块胶布松散整个胸腔下部（尾）部分，只是隔膜上方。位置应足够紧PROC过程中保持正确对准edure，但不是那么紧，它限制呼吸。 </li><li>打开照明灯％和100％的强度80之间和定向鹅颈，以便它是从皮肤的表面1至2厘米，靠近太阳穴。定期检查鹅颈管的尖端热，以确保它不伤害老鼠。 </li><li>站在平台背后，用无菌，钝端钳定位舌头。小心，以避免下切牙，轻轻夹住并绘制舌头口腔。 </li><li>使用剩余的手，插入抑制剂并用它来压平舌针对口腔的楼层。松开钳，但留下的降压到位接下来的两个步骤。 </li><li>定向光使气管是由从太阳穴的水平向近侧引导鹅颈管，直到它到达主支气管的水平可见的。 注意:气管可通过呼吸的作用而容易地分开，WHICH导致发出的光的强度波动。当正确定位时，该结构将在轴向平面内的光的一个位于中心的销与在口腔中本身最小环境光可辨别。 </li><li>角的注射器，使得它跟随气管的自然路径，并降低22-G的导管末端，与含有微滴，直入管腔所附注射器。 PBS的泡沫将开始上升，并在成功安置每次呼吸下降。 注意:此动作可通过几秒钟作为深度镇静的结果被延迟。 </li><li>喂导管在额外的5毫米。取出压舌板。 </li><li>移注射器相反的手，抓住中心，轻轻取下注射器。 </li><li> BLM的30和45微升之间存到导管毂的内部的中心，连接第二注射器和分配300μl的空气进入中心。 </li><li>更换第二与第一含PBS的气泡注射器。泡沫将继续上升，如果程序已经成功地进行下降。 </li></ol> 3.后护理程序<ol><li>取下导管和胶带，然后将动物在干燥温暖的地方，直到它恢复意识 - 通常在一两分钟。 </li></ol>

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The authors have nothing to disclose.

在雾化是不切实际由于有限的试剂可用性，安全性或成本的情况下，直接气管施用是用于递送外源剂的进入肺部优良的方法16经气管滴注已被广泛用于完成此。然而，与所有的外科手术，它也与它携带用于引起该过程本身并发症的可能性，并且不一定剂被灌输。13 由于这些原因，已经变得越来越普遍，直接经由插管16,17,18,19,20-服用一种物质进入气管。然而，这些方法也遭受限制，包括关于该导管的位置，由于利用强大的镇静剂像氯胺酮/赛拉嗪延长的恢复时间，不确定性（即，进入食道而不是气管），在组织中的不必要的操作口腔，和相关的成本与购买专用仪器，如耳镜/喉镜。 一旦建立，这里列出的方法提供了比其他插管协议的几个有意义的好处。一方面，使用异氟醚的允许在每只小鼠花费的总时间的减少。这主要是由于以恢复知觉以下相比像氯胺酮受控物质的异氟烷镇静所需动物的减少时间（约5分钟与有时大于3小时）14另外，临时肺活量计通过允许研究者保证成功的信心以可视化的振荡PBS泡沫，这表明之前和之后滴注给药物质的正确位置。最后，通过利用外部光源，而不是一个笨重耳镜或喉镜和导丝，研究者能够减少他们手中设备的数量，同时提高其查看OF中的口腔和减少其损伤周围气管软组织的可能性。 与此协议相关的具体问题不多，但严重。例如，虽然使用异氟醚的减少与从镇静诱导和恢复相关联的时间，它也需要在动物的镇静状态的严格观察。未能从异氟醚室取出的动物及时可导致死亡，尤其是在动物已经在上一次干预复苏。相反，异氟醚不足剂量和延长程序时间可以导致动物的过程本身，这也可能导致人身伤害中恢复意识。因此，当务之急是为动物两者的安全性和使动物在意识的第一个迹象返回到异氟烷室实验的完整性。自学习这种方法时这可以是通用的，所以建议在经验丰富的研究证实麻醉下一个之前每个动物的成功管理。此外，有困难的研究人员还可以选择延长异氟醚曝光率以增加可用的过程时，只要他们密切监视动物濒死。 这种方法的另一个问题是，与处理相关的损害。这是在三个步骤尤其可能:从口中取出时舌头，插入导管时，并撞出从轮毂治疗后的注射器时。该导管的放置是迄今为止最危险的这些步骤。重要的是，直到气管的观点是理想的导管不应进入口腔。如果气管不能通过任一光源或降压的操作被带入视图中，研究者应松开舌和重新尝试的过程。围绕气管i上的软组织的操纵不是必需的或推荐。然而，一旦导管已被正确地放置，应注意也可采取保持毂从太远进入气管前进和射孔主茎支气管。这通常可以通过确保导​​管仅松散地附着到注射器筒体被避免，并且通过支撑含有对程序板的导管的手使之保持静止。这些细微之处，在需要的及时性和精准在一起，突出完善这一过程的准备和实践的重要性。然而，随着训练它应该有可能对于一个新手研究者超过一个小时的过程中处理20个或更多的动物，和有经验的研究人员来完成两倍多或更多。 通过直接气管沉积传授的一致性使得它的管理是近年来最佳途径。事实上，类似的方法已经报道了完全不同的队友众多里亚尔这种方式19,25。与这样的协议相一致，在这里所概述的方式博莱霉素的递送导致两肺强效纤维化反应，并在所有波瓣，这表明深度和均匀性分布的类似水平。在此基础上，很可能，这个协议提供了可比的实验优势的范围内，依赖于接触来介导其作用的其它材料（例如，药品，抗体，细菌剂， 和体内蛋白质修饰系统，诸如慢病毒载体）和，像博莱霉素，它将允许这样的试验也可与信心，小的费用，和最小的术后护理进行。

尽管某些解剖和生理差异，1鼠模型继续是无价建模人类生物学和疾病的发病机制。2从饲养的角度来看，小鼠是容易处理，具有低的繁殖时间，加速寿命，并且是相对便宜的去家。用不同遗传品系和策略（例如，条件敲奏，记者小鼠谱系追踪方法等），以及在宽范围内可用的试剂（例如，抗体，重组蛋白，抑制剂的开发，等），老鼠已经成为一个重要的脊椎动物模型生物体揭示人类的稳态和疾病过程。3 小鼠已经特别有价值用于研究肺部疾病，包括急性肺损伤（ALI）和肺纤维化。4 ALI在人类可以通过创伤，损伤或败血症引起，其特征是上皮和内皮泄漏（即，水肿），炎症，和新生的纤维化。在许多患者中，急性肺损伤前进到它的严重的形式，急性呼吸窘迫综合征（ARDS），这经常导致纤维化和死亡因呼吸衰竭。5,6-肺纤维化是一个渐进的，致命的病理学特征在于细胞外基质的过量沉积，最显着的I型胶原，导致肺功能受损。-7,8-管理博莱霉素（BLM）是最广泛使用和最佳表征模型实验动物诱导ALI和纤维化。9虽然博莱霉素诱导的肺纤维化啮齿动物确实不概括完全人类纤维化表型，与该模型10小鼠研究已导致的影响的发生和疾病的进展许多重要因素的发现。11 虽然确切机制（S）的后面博莱霉素诱导的纤维化是未知的，则发起伤被认为是从在上皮细胞接触依赖性的DNA链断裂衬传导气道和肺泡，并且特别地，1型肺泡产生。12的必要性BLM和肺上皮之间的直接接触突出一个健壮的递送途径的重要性和这些问题也有密切关系广泛的治疗靶向远端气道，包括重组蛋白，抗体，siRNA的，病毒，细菌，微粒，等等。口咽抽吸（OPA）已被广泛地用于此目的13，但OPA的一个主要缺点是，递送剂的某些部分可以被吞咽到胃肠道，从而导致不精确的给药剂量。另一广泛使用的方法是经气管滴注，涉及强麻醉下气管到代理的气管和滴注暴露直接进入呼吸道。14然而，不仅可以这样一个过程是不希望的，由于它的invasivity，但它也是耗时的，需要的培训公平位，同时产生强效伤到呼吸道15,16若干协议已被开发涉及的试剂的直接施用进而不需要手术干预气管，引起强大麻醉药16,17,18,19,20-但这些方法包括延长的恢复时间，使用昂贵的设备（即，耳镜/喉镜，市售程序板，光纤线等），在口腔中的过量操纵，和不确定性有关的剂量。 本文描述的给药通过插管相对容易的方法，其允许研究者快速，廉价和可靠地灌输试剂进鼠肺与对周围组织的残余破坏的风险有限。

<table><tbody><tr><td>Bleomycin For Injection, 30 units/vial</td><td>APP Pharmaceuticals, LLC</td><td>103720</td><td>For best results, BLM should be suspended in PBS, aliquoted, and stored as single use lyophilzed aliquots</td></tr><tr><td>Blunt End Forceps</td><td></td><td></td><td></td></tr><tr><td>Tongue Depressor (i.e. bent Valleylab Blade Electrode, 2.4&quot;)&amp;nbsp;</td><td>Covidien</td><td>E1551G</td><td>Before use, create a 45 degree bend 1.5 cm&amp;nbsp; from the blade tip. A suitable depressor can also be created from any metal implement of similar dimensions.&amp;nbsp;</td></tr><tr><td>Exel Safelet Catheter 22G X 1&quot;</td><td>Exel International</td><td>26746</td><td></td></tr><tr><td>1 ml Slip-tip Disposable Tuberculin Syringe (200/sp, 1600/ca)</td><td>BD</td><td>309659</td><td></td></tr><tr><td>0.2 ml Pipettor and Filter Tips</td><td></td><td></td><td></td></tr><tr><td>Fiber-Lite Illuminator. Model 181-1: Model 180 mated with Standard Dual Gooseneck illuminator</td><td>Dolan Jenner Industries, Inc.</td><td>181-1</td><td>Lower output LED illuminators are not recommended as they fail to suficiently illuminate the trachea.</td></tr><tr><td>Intubation Board</td><td></td><td></td><td>See Diagram 1.</td></tr><tr><td>Colored Label Tape: 0.5&amp;nbsp;in. Wide</td><td>Fisherbrand</td><td>15-901-15A</td><td></td></tr><tr><td>Oxygen&amp;nbsp;</td><td></td><td></td><td></td></tr><tr><td>Phosphate-Buffered Saline, 1X</td><td>Corning</td><td>21-040-CV</td><td>Product should be sterile</td></tr><tr><td>Non-Sterile Silk Black Braided Suture Spool, 91.4 m, Size 4-0</td><td>Harvard Apparatus</td><td>517698</td><td></td></tr><tr><td>Table Top Anesthesia Machine Isoflurane</td><td>Highland Medical Equipment</td><td></td><td>http://www.highlandmedical.net/</td></tr><tr><td>Slide Top Induction Mouse Isoflurane Chamber</td><td>MIP / Anesthesia Technologies</td><td>AS-01-0530-SM</td><td></td></tr><tr><td>FORANE (isoflurane, USP) Liquid For Inhalation 100 mL&amp;nbsp;</td><td>Baxter</td><td>1001936040</td><td></td></tr><tr><td>&amp;nbsp;Nanozoomer Digital Pathology system</td><td>Hamamatsu</td><td></td><td></td></tr><tr><td>IgM ELISA Quantification Kit&amp;nbsp;</td><td>Bethyl Laboratories</td><td>E90-101</td><td></td></tr></tbody></table>

an improved method for rapid intubation of the trachea in mice

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity is a commonly used model to study both acute lung injury and fibrosis, and multiple methods have been developed for administering bleomycin (and other toxic agents) into the lungs. However, many of these approaches, such as transtracheal instillation, have inherent drawbacks, including the need for strong anesthetics and survival surgery. This paper reports a quick, reproducible method of intratracheal intubation that involves mild inhaled anesthesia, visualization of the trachea, and the use of a surrogate spirometer to confirm exposure. As a proof of concept, 8-12 week old C57BL/6 mice were administered either 2.0 U/kg of bleomycin or an equivalent volume of PBS, and both damage and fibrotic endpoints were measured post-exposure. This procedure allows researchers to treat a large cohort of mice in a relatively short period with little expense and minimal post-procedure care.

如别处所述21插管小鼠每天监测体重减轻和窘迫，处死4,10或17天后通过2.5％2,2,2- 三溴腹腔注射。 支气管肺泡灌洗 （BAL）收集在PBS中洗涤三次和右肺固定在10％福尔马林，石蜡包埋，并用Masson三色由华盛顿组织学和成像核心22的大学染色。 在既定的数据一致，博莱霉素处理的小鼠经历7和10天暴露后23（图2A）之间的峰的重量损失。此外，在BAL IgM的水平升高从处死博莱霉素处理的小鼠表现出显著，时间依赖性增加在肺通透性，指示上皮细胞和/或内皮屏障功能障碍（ 图2B）的。 ="jove_content"FO:保together.within页="1"&gt;使用马松染色测量纤维化反应-总胶原蛋白含量24的行之有效的标记。右边肺部进行染色，并且每右肺部分三色染色的平均总面积进行定量（图3A）。代表性的部分显示在纤维化病变 （ 图3B）的肺间质和增加的所得治疗依赖性增厚 。 <img alt="图1" src="/files/ftp_upload/53771/53771fig1.jpg" /> 图1.生产工序的板常规参数，给出的长度表明有必要创建工序设备的大致尺寸。在这些实验中，标准铝金属片切成适当的尺寸，然后用"T"型支架加强。ftp_upload / 53771 / 53771fig1large.jpg"目标="_空白"&gt;点击此处查看该图的放大版本。 <img alt="图2" src="/files/ftp_upload/53771/53771fig2.jpg" /> 图下面的博来霉素的施用2.体重减轻和屏障功能障碍。小鼠施用2.0单位/ kg的BLM或PBS的等量的。 （A）减肥表示为0天的重量百分比（BLM:N = 21; PBS:5）。 （B）的小鼠在第4天或10天暴露后处死，从收获的肺中BAL IgM的蛋白通过ELISA测量。 （PBS:N = 5;第4天:N = 3;第10天:N = 5）。数据表示为平均值±SEM表示。 * P &lt;0.05相比，PBS的控制。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/53771/53771fig2large.jpg" target="_blank">请点击此处查看该图的放大版本。</a> <img alt="图3"SRC ="/文件/ ftp_upload / 53771 / 53771fig3.jpg"/&gt; 以下博莱霉素管理图3.肺纤维化。（A）总胶原蛋白，量化为从WT控制总倍数变化马松染色。 （B）以 5倍放大倍数下拍摄三色染色的肺切片的代表性图像。 PBS控制包括5至14天之间采取后PBS对待动物（PBS:N = 5;第10天:N = 5 17天:N = 3）。数据表示为平均值±SEM表示。 * P &lt;0.05相比，PBS的控制。 <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/53771/53771fig3large.jpg" target="_blank">请点击此处查看该图的放大版本。</a>

Watch this Scientific Journal Video about An Improved Method for Rapid Intubation of the Trachea in Mice at JoVE.com

An Improved Method for Rapid Intubation of the Trachea in Mice

在小鼠气管插管快速的改进方法

本文介绍了通过插管直接施用博莱到小鼠气管一种快速，简便的方法。这种方法的主要优点是，它是高度可再现的，容易掌握，并且不需要专门的设备或冗长的恢复时间。

Despite some anatomical and physiological differences, mouse models continue to be an essential tool for studying human lung disease. Bleomycin toxicity ...

an-improved-method-for-rapid-intubation-of-the-trachea-in-mice

Nanyang Technological University

Research

JoVE Journal

Medicine

25.1K 次观看.  University of Washington. 本文介绍了通过插管直接施用博莱到小鼠气管一种快速，简便的方法。这种方法的主要优点是，它是高度可再现的，容易掌握，并且不需要专门的设备或冗长的恢复时间。 

视频: An Improved Method for Rapid Intubation of the Trachea in Mice

Pressure Controlled Ventilation to Induce Acute Lung Injury in Mice

Murine models are extensively used to investigate acute injuries of different organs systems (1-34). Acute lung injury (ALI), which occurs with prolonged mechanical ventilation, contributes to morbidity and mortality of critical illness, and studies on novel genetic or pharmacological targets are areas of intense investigation (1-3, 5, 8, 26, 30, 33-36). ALI is defined by the acute onset of the disease, which leads to non-cardiac pulmonary edema and subsequent impairment of pulmonary gas exchange (36). We have developed a murine model of ALI by using a pressure-controlled ventilation to induce ventilator-induced lung injury (2). For this purpose, C57BL/6 mice are anesthetized and a tracheotomy is performed followed by induction of ALI via mechanical ventilation. Mice are ventilated in a pressure-controlled setting with an inspiratory peak pressure of 45 mbar over 1 - 3 hours. As outcome parameters, pulmonary edema (wet-to-dry ratio), bronchoalveolar fluid albumin content, bronchoalveolar fluid and pulmonary tissue myeloperoxidase content and pulmonary gas exchange are assessed (2). Using this technique we could show that it sufficiently induces acute lung inflammation and can distinguish between different treatment groups or genotypes (1-3, 5). Therefore this technique may be helpful for researchers who pursue molecular mechanisms involved in ALI using a genetic approach in mice with gene-targeted deletion.

A murine model for ventilator induced lung injury is an important tool to study an acute lung injury in vivo. Here, we report an easy applicable in situ model for acute lung injury using high-pressure mechanical ventilation to induce acute failure of the lung.

压力控制通气诱导小鼠急性肺损伤

Murine models are extensively used to investigate acute injuries of different organs systems (1-34). Acute lung injury (ALI), which occurs with prolonged ...

科研

医学

Endotracheal Intubation in Mice via Direct Laryngoscopy Using an Otoscope

Mice, both wildtype and transgenic, are the principal mammalian model in biomedical research currently. Intubation and mechanical ventilation are necessary for whole animal experiments that require surgery under deep anesthesia or measurements of lung function. Tracheostomy has been the standard for intubating the airway in these mice to allow mechanical ventilation. Orotracheal intubation has been reported but has not been successfully used in many studies because of the substantial technical difficulty or a requirement for highly specialized and expensive equipment. Here we report a technique of direct laryngoscopy using an otoscope fitted with a 2.0 mm speculum and using a 20 G&#160;intravenous catheter as an endotracheal tube. We have used this technique extensively and reliably to intubate and conduct accurate assessments of lung function in mice. This technique has proven safe, with essentially no animal loss in experienced hands. Moreover, this technique can be used for repeated studies of mice in chronic models.

Endotracheal Intubation in Mice via Direct Laryngoscopy Using an Otoscope

We have developed a simple, reliable, and relatively inexpensive method for endotracheal intubation in mice via direct laryngoscopy using an otoscope with a 2.0 mm speculum. This technique is atraumatic and can be used for repeated measurements in chronic experiments. We find it superior to tracheostomy or previously reported nonsurgical techniques.

气管插管小鼠<em&gt;通过</em&gt;直接喉镜使用耳镜

Mice, both wildtype and transgenic, are the principal mammalian model in biomedical research currently. Intubation and mechanical ventilation are ...

A Simple Method of Mouse Lung Intubation

A simple procedure to intubate mice for pulmonary function measurements would have several advantages in longitudinal studies with limited numbers or expensive animal. One of the reasons that this is not done more routinely is that it is relatively difficult, despite there being several published studies that describe ways to achieve it. In this paper we demonstrate a procedure that eliminates one of the major hurdles associated with this intubation, that of visualizing the trachea during the entire time of intubation. The approach uses a 0.5 mm fiberoptic light source that serves as an introducer to direct the intubation cannula into the mouse trachea. We show that it is possible to use this procedure to measure lung mechanics in individual mice over a time course of at least several weeks. The technique can be set up with relatively little expense and expertise, and it can be routinely accomplished with relatively little training. This should make it possible for any laboratory to routinely carry out this intubation, thereby allowing longitudinal studies in individual mice, thereby minimizing the number of mice needed and increasing the statistical power by using each mouse as its own control.

This paper describes a striaghforward and efficient method of intubating mice for pulmonary function measurements or pulmonary instillation, that allows the mice to recover and be studied at later times. The procedure involves an inexpensive fiberoptic light source that directly illuminates the trachea.

一个简单的方法小鼠肺插管

A  simple procedure to intubate mice for pulmonary function measurements would  have several advantages in longitudinal studies with limited numbers or  ...

Noninvasive Intratracheal Intubation to Study the Pathology and Physiology of Mouse Lung

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular disease and for developing therapeutic intervention. With the increasing availability of knockout and transgenic derivatives, together with a vast amount of genetic information, mice provide one of the best models to study the molecular mechanisms underlying the pathology and physiology of lung diseases. Inhalation, intranasal instillation, intratracheal instillation, and intratracheal intubation are the most widely used techniques by a number of investigators to administer materials of interest to mouse lungs. There are pros and cons for each technique depending on the goals of a study. Here a noninvasive intratracheal intubation method that can directly deliver exogenous materials to mouse lungs is presented. This technique was applied to administer bleomycin to mouse lungs as a model to study pulmonary fibrosis.

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular disease and for developing therapeutic intervention. Here a noninvasive intratracheal intubation method that can directly deliver exogenous materials to mouse lungs is presented.

无创性气管内插管，以研究小鼠肺的病理生理

The use of a model that mimics the condition of lung diseases in humans is critical for studying the pathophysiology and/or etiology of a particular ...

A Reversible, Non-invasive Method for Airway Resistance Measurements and Bronchoalveolar Lavage Fluid Sampling in Mice

Airway hyperreactivity (AHR) measurements and bronchoalveolar lavage (BAL) fluid sampling are essential to experimental asthma models, but repeated procedures to obtain such measurements in the same animal are generally not feasible.  Here, we demonstrate protocols for obtaining from mice repeated measurements of AHR and bronchoalveolar lavage fluid samples.  Mice were challenged intranasally seven times over 14 days with a potent allergen or sham treated.  Prior to the initial challenge, and within 24 hours following each intranasal challenge, the same animals were anesthetized, orally intubated and mechanically ventilated.  AHR, assessed by comparing dose response curves of respiratory system resistance (RRS) induced by increasing intravenous doses of acetylcholine (Ach) chloride between sham and allergen-challenged animals, were determined.  Afterwards, and via the same intubation, the left lung was lavaged so that differential enumeration of airway cells could be performed.  These studies reveal that repeated measurements of AHR and BAL fluid collection are possible from the same animals and that maximal airway hyperresponsiveness and airway eosinophilia are achieved within 7-10 days of initiating allergen challenge.  This novel technique significantly reduces the number of mice required for longitudinal experimentation and is applicable to diverse rodent species, disease models and airway physiology instruments.  

Repeated measurements of rodent respiratory physiology and sampling of airway inflammatory cells are desirable, but generally not feasible. Here we describe a repeatable method for orally intubating mice that permits repeated measurements of airway hyperreactivity and sampling of airway inflammatory cells.

一个可逆的，非侵入性方法，气道阻力测量和支气管肺泡灌洗小鼠流体取样

Airway hyperreactivity (AHR) measurements and bronchoalveolar lavage (BAL) fluid sampling are essential to experimental asthma models, but repeated ...

生物学

A Closed-chest Model to Induce Transverse Aortic Constriction in Mice

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to perform a partial thoracotomy to visualize the transverse aortic arch. However, the surgical trauma caused by the thoracotomy in open-chest models changes the respiratory physiology as the ribs are dissected and left unattached after chest closure. To prevent this, we established a minimally invasive, closed chest approach via lateral thoracotomy. Herein we approach the aortic arch via the 2nd intercostal space without entering the chest cavities, leaving the mouse with a less traumatic injury to recover from. We perform this operation using standard laboratory settings for open chest TAC procedures with equal survival rates. Apart from maintaining physiological breathing patterns due to the closed chest approach, the mice seem to benefit by showing rapid recovery, as the less invasive technique appears to facilitate a fast healing process and to reduce immune response after trauma.

Here, we present a protocol of Transverse Aortic constriction (TAC) via a lateral thoracotomy. This technique is a minimally invasive, closed chest surgical procedure aiming to simulate pressure overload and heart failure in mice utilizing standard TAC laboratory settings.

诱导小鼠横向主动脉狭窄的闭合胸模型

Research on cardiac hypertrophy and heart failure is frequently based on pressure overload mouse models induced by TAC. The standard procedure is to ...

A Mouse Model of Orotracheal Intubation and Ventilated Lung Ischemia Reperfusion Surgery

Ischemia reperfusion (IR) injury frequently results from processes that involve a transient period of interrupted blood flow. In the lung, isolated IR permits the experimental study of this specific process with continued alveolar ventilation, thereby avoiding the compounding injurious processes of hypoxia and atelectasis. In the clinical context, lung ischemia reperfusion injury (also known as lung IRI or LIRI) is caused by numerous processes, including but not limited to pulmonary embolism, resuscitated hemorrhagic trauma, and lung transplantation. There are currently limited effective treatment options for LIRI. Here, we present a reversible surgical model of lung IR involving first orotracheal intubation followed by unilateral left lung ischemia and reperfusion with preserved alveolar ventilation or gas exchange. Mice undergo a left thoracotomy, through which the left pulmonary artery is exposed, visualized, isolated, and compressed using a reversible slipknot. The surgical incision is then closed during the ischemic period, and the animal is awakened and extubated. With the mouse spontaneously breathing, reperfusion is established by releasing the slipknot around the pulmonary artery. This clinically relevant survival model permits the evaluation of lung IR injury, the resolution phase, downstream effects on lung function, as well as two-hit models involving experimental pneumonia. While technically challenging, this model can be mastered over the course of a few weeks to months with an eventual survival or success rate of 80%-90%.

A mouse surgical model to create left lung ischemia reperfusion (IR) injury while maintaining ventilation and avoiding hypoxia.

口腔插管和通气肺缺血再灌注手术小鼠模型

Ischemia reperfusion (IR) injury frequently results from processes that involve a transient period of interrupted blood flow. In the lung, isolated IR ...

免疫与感染

In vivo Calcium Imaging in Mouse Inferior Olive

Inferior olive (IO), a nucleus in the ventral medulla, is the only source of climbing fibers that form one of the two input pathways entering the cerebellum. IO has long been proposed to be crucial for motor control and its activity is currently considered to be at the center of many hypotheses of both motor and cognitive functions of the cerebellum. While its physiology and function have been relatively well studied on single-cell level in vitro, presently there are no reports on the organization of the IO network activity in living animals. This is largely due to the extremely challenging anatomical location of the IO, making it difficult to subject to conventional fluorescent imaging methods, where an optic path must be created through the entire brain located dorsally to the region of interest.
Here we describe an alternative method for obtaining state-of-the-art -level calcium imaging data from the IO network. The method takes advantage of the extreme ventral location of the IO and involves a surgical procedure for inserting a gradient-refractive index (GRIN) lens through the neck viscera to come into contact with the ventral surface of the calcium sensor GCaMP6s-expressing IO in anesthetized mice. A representative calcium imaging recording is shown to demonstrate the feasibility to record IO neuron activity after the surgery. While this is a non-survival surgery and the recordings must be conducted under anesthesia, it avoids damage to life-critical brainstem nuclei and allows conducting large variety of experiments investigating spatiotemporal activity patterns and input integration in the IO. This procedure with modifications could be used for recordings in other, adjacent regions of the ventral brainstem.

In vivo Calcium Imaging in Mouse Inferior Olive

We present a protocol to expose the brainstem of adult mouse from the ventral side. By using a gradient-refractive index lens with a miniature microscope, calcium imaging can be used to examine the activity of inferior olive neural somata in vivo.

在体内 小鼠劣质橄榄中的钙成像

Inferior olive (IO), a nucleus in the ventral medulla, is the only source of climbing fibers that form one of the two input pathways entering the ...

神经科学

Repeated Orotracheal Intubation in Mice

The literature describes several methods for mouse intubation that either require visualization of the glottis through the oral cavity or incision in the ventral neck for direct confirmation of cannula placement in the trachea. The relative difficulty or the tissue trauma induced to the subject by such procedures can be an impediment to an investigator&#8217;s ability to perform longitudinal studies. This article illustrates a technique in which physical manipulation of the mouse following the use of a depilatory to remove hair from the ventral neck permits transcutaneous visualization of the trachea for orotracheal intubation regardless of degree of skin pigmentation. This method is innocuous to the subject and easily achieved with a limited understanding of murine anatomy. This refined approach facilitates repeated intubation, which may be necessary for monitoring progression of disease or instillation of treatments. Using this method may result in a reduction of the number of animals and technical skill required to measure lung function in mouse models of respiratory disease.

The goal of this article is to describe a refined method of intubation of the laboratory mouse. The method is noninvasive and, therefore, ideal for studies that require serial monitoring of respiratory function and/or instillation of treatments into the lung.

小鼠重复的奥内切管插管

The literature describes several methods for mouse intubation that either require visualization of the glottis through the oral cavity or incision in the ...

<meta charset="utf8"></head><body>基于气管切开术的气管插管方案，用于非存活大鼠模型

Endotracheal Intubation via Tracheotomy and Subsequent Thoracotomy in Rats for Non-Survival Applications

Endotracheal intubation and subsequent ventilation are often basic requirements for translational research in rat models for various interventions that require controlled or high ventilation pressures or access to the thoracic cavity and organs. Conventional endoorotracheal intubation using the anatomically existing route through the mouth is well suited for survival experiments. However, this procedure poses some challenges, including generally higher levels of the required experience and technical skill, more advanced equipment, and greater time effort with relevant intubation failure rates and complications such as tracheal perforation, temporary systemic hypooxygenation, and relevant aerial leakage.
This manuscript, therefore, presents a detailed step-by-step protocol for endotracheal intubation through tracheotomy in non-survival rat models when guaranteed intubation success, constant oxygenation levels, high ventilation pressures, or open thoracotomy are required.
The protocol emphasizes the importance of meticulous surgical technique to ensure consistent and reliable outcomes, especially for researchers who are inexperienced or lack routine in the technique of endoorotracheal intubation via direct laryngoscopy. This procedure is, therefore, expected to minimize animal suffering and unnecessary animal losses.

<meta charset="utf8"></head><body>大鼠气管切开术气管插管和随后的开胸术用于非存活应用

Here we introduce a standardized procedure for endotracheal intubation via tracheotomy followed by thoracotomy in rats, aimed at enhancing the precision and reproducibility of non-survival applications requiring invasive ventilation and exposure of thoracic organs in in vivo rat models.

大鼠气管切开术气管插管和随后的开胸术用于非存活应用

Endotracheal intubation and subsequent ventilation are often basic requirements for translational research in rat models for various interventions that ...

在小鼠气管插管快速的改进方法

摘要

探索更多视频

此视频中的章节

压力控制通气诱导小鼠急性肺损伤

气管插管小鼠

一个简单的方法小鼠肺插管

无创性气管内插管，以研究小鼠肺的病理生理

一个可逆的，非侵入性方法，气道阻力测量和支气管肺泡灌洗小鼠流体取样

诱导小鼠横向主动脉狭窄的闭合胸模型

口腔插管和通气肺缺血再灌注手术小鼠模型

在体内小鼠劣质橄榄中的钙成像

小鼠重复的奥内切管插管

大鼠气管切开术气管插管和随后的开胸术用于非存活应用

在小鼠气管插管快速的改进方法

摘要

探索更多视频

此视频中的章节

压力控制通气诱导小鼠急性肺损伤

气管插管小鼠

一个简单的方法小鼠肺插管

无创性气管内插管，以研究小鼠肺的病理生理

一个可逆的，非侵入性方法，气道阻力测量和支气管肺泡灌洗小鼠流体取样

诱导小鼠横向主动脉狭窄的闭合胸模型

口腔插管和通气肺缺血再灌注手术小鼠模型

在体内 小鼠劣质橄榄中的钙成像

小鼠重复的奥内切管插管

大鼠气管切开术气管插管和随后的开胸术用于非存活应用

在体内小鼠劣质橄榄中的钙成像