著者らは、トリクローム染色および分析のヘルプのためにワシントン大学の組織学とイメージングコアのブライアン・ジョンソンに感謝します。この作品は、NIHの助成金HL098067とHL089455によってサポートされていました。

ワシントンとシーダーズ・サイナイ医療センターの大学の施設内動物管理使用委員会（IACUC）は、これらの研究のために必要な動物の作業を承認しました。 1.準備<ol><li>平滑末端鉗子と降圧介してオートクレーブの両方を滅菌します。 </li><li>生物学的安全キャビネットを使用して、凍結乾燥粉末からPBSでBLMの作業ストックを準備します。偶数混合を確実にするために35 kHzで10分間この溶液を超音波処理します。 注：μlの30〜45の総容量は、より大容量で、ピペットローエンドのバリエーション、および窒息を防ぐために推奨されます。 </li><li>手順自体は約1メートル2を備えてクリーンなワークスペースを準備し、同様の手順の前と後の両方のケージの場所を指定。 </li><li>ベースとunderl渡って実験用テープの2または3のストリップを置くことによって、直ちに研究者の前でベンチに手続きボードのベースを修正しました。英ベンチ。ボードの作成 ​​に関する更なる仕様については、図1を参照してください。 </li><li>手順ボードの2つの位置決めねじのサイズ4.0縫合糸の単一の長さを接続します。 </li><li>削除し、3 1ミリリットルのシリンジからプランジャーを破棄し、気密シールを形成するために、各バレルの上部にPBSを60μlを堆積させることにより、その場しのぎの肺活量計を生成します。シリンジの1に緩くカテーテルのハブを固定し、基板の一方の側に配置します。 </li><li> 1mlシリンジ内への空気の吸引し、300μlのとは、基板の一方の側に配置します。 </li><li>一辺までの長さと場所で約6インチテープの追加部分をカット。これは、ステップ2.4​​でボードに動物を保護するために使用されます。 </li><li>イソフルラン室を設定します。露光室とクリアランス真空の両方で適切なポートに、O 2、イソフルラン、真空を取り付けます。また、イソフルラン互換生物学的に麻酔薬を投与します安全キャビネット。 </li></ol> 2.挿管<ol><li>それは意識を失い、呼吸が適切な速度に減速するまで、チャンバ内でイソフルランでマウスを麻酔。典型的な曝露は、3〜4分間、4％イソフルランおよび2％O 2を含み、理想的な結果は、鎮静の2〜2.5分です。これは、すべての2秒1呼吸の呼吸速度に対応しています。 </li><li>片側にピペッター、所定の位置にBLMの30と45μlの間に設定するには、鎮静、吸引を待っている間。 </li><li>準備ができたら、手順プラットフォームの位置決めネジに取り付けられたスレッドからその上顎切歯によって鎮静させたマウスを一時停止します。動物の背側は、プラットフォーム表面に対して平らにすることを確認してください。 </li><li>換気を制限しないように注意しながら、ちょうど横隔膜の上、緩く胸腔の下（尾）部分を横切ってテープ片を配置します。配置は、procの中に適切な位置合わせを維持するのに十分タイトであるべきですedureが、それは呼吸を制限することは非常にタイトではありません。 </li><li> 80％の間と100％の強度に照明をオンにして、それは太陽神経叢の近くに、皮膚の表面から1〜2cm程度になるように、グース​​ネックを向けます。定期的にマウスを傷つけることがないように、熱のためのグースネックの先端を確認してください。 </li><li>プラットフォームの後ろに立ち、舌を見つけるために、滅菌、平滑末端の鉗子を使用しています。下の切歯、軽くグリップを回避し、口腔外に舌を描くように注意しながら。 </li><li>残りの手を使用して、圧子を挿入し、口腔内の床面に対して舌を平らにするためにそれを使用。鉗子を解放しますが、次の2つの手順のための場所で圧子を残します。 </li><li>オリエント光気管は、それがmainstem気管支のレベルに達するまで太陽神経叢のレベルから近位グースネックを案内することにより、表示されるように。 注：気管を簡単に呼吸の作用によって区別することができ、WHIchが放出された光は強度が変動する原因となります。正しく配置すると、この構造体は、口腔自体に最小限の環境光の光の中心に位置するピンとして軸平面に識別可能になります。 </li><li>角度シリンジそれは気管の自然な経路をたどる、とストレート内腔に、液滴を含む添付のシリンジを用いて、22-Gのカテーテル先端を下げるようにします。 PBSバブルが成功した配置時に各呼吸と共に上昇し、秋に開始されます。 注：このアクションは、深い鎮静の結果として数秒遅れで表示することができます。 </li><li>追加の5ミリメートルにカテーテルを養います。舌を取り外し圧子。 </li><li>反対側の手に注射器をシフトし、ハブを把持し、静かに注射器を取り外します。 </li><li>カテーテルハブの内部の中心部へのBLMの30と45μlの間の預金は、第2のシリンジを接続し、ハブへの空気の300μLを分注します。 </li><li>第二を交換しますPBSの気泡を含有する第一と注射器。バブルが上昇し、プロシージャが正常に実行された場合に落下していきます。 </li></ol> 3.ポスト手続きケア<ol><li>カテーテルとテープを取り外し、それが意識を取り戻すまで、乾燥した暖かい場所に動物を置く - 通常、数分以内に。 </li></ol>

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The authors have nothing to disclose.

エアロゾル化が原因で制限された試薬の利用可能性、安全性、またはコストに実用的ではない例において、直接気管内投与は、肺への外因性薬剤の送達のための優れた方法である16経気管点滴注入が広くこれを達成するために使用されています。しかしながら、すべての外科的介入と同様に、それはまた、それを使用して手順自体に起因する合併症の可能性を運び、そして必ずしも薬...

畜産観点からいくつかの解剖学的および生理学的な違いにもかかわらず、1マウスモデルは、ヒトの生物学と疾患の病因をモデル化するための非常に貴重であり続ける。2、マウスは、取り扱いが容易で低繁殖時間を持って、加速寿命、かつ比較的安価です家へ。多様な遺伝的系統と戦略（ 例えば 、条件付きノックアウト、レポーターマウス、系統追跡手法など）、ならびに入手可能な試薬の広い範囲（ 例えば 、抗体、組換えタンパク質、阻害剤の開発と、等）、マウスは、ヒト恒常性と疾患過程を明らかにするために不可欠なモデル脊椎動物の生物となっている。3 マウスはヒトで4 ALIは、外傷、傷害、または敗血症によって引き起こされ得る。急性肺損傷（ALI）および肺線維症などの肺の状態を研究するために特に有益であったと上皮を特徴とし内皮リーク（ すなわち、浮腫）、炎症、および新生線維症。多くの患者では、ALIは、その重篤な形態、しばしば線維症、呼吸不全のために死をもたらす急性呼吸窮迫症候群（ARDS）、へと進行する。5,6肺線維症は、細胞外マトリックスの過剰沈着を特徴とする進行、致命的な病態であります、最も顕著に私が損なわ肺機能につながる、型コラーゲン。ブレオマイシンの7,8管理局（BLM）は、実験動物でALIおよび線維症を誘導するための最も広く使用されていると最もよく特徴付けられたモデルである。9げっ歯類におけるBLM誘発肺線維症はありませんが、完全にヒトの線維性の表現型を再現ではない、このモデルと10マウス研究は、疾患の発症と進行に影響を与える多くの重要な要因の発見につながっている。11 BLM誘発性線維形成の背後にある正確なメカニズム（s）は不明であるが、開始けが誘導気道および肺胞、特に、1型肺の内側を覆う上皮細胞における接触依存性DNA鎖切断に起因すると考えられる。12 BLMおよび肺上皮との間の直接接触の必要性は、頑強な送達経路の重要性を強調、およびこれらの懸念はまた、組換えタンパク質、抗体、siRNAを、ウイルス、細菌、微粒子、および多くを含む遠位気道を標的とする治療法の広い範囲に密接な関係がある。口腔咽頭吸引（OPA）が広く、この目的の13のために使用されてきたが、OPAの主な欠点は、送達される薬剤の一部がそれによって投与量に不正確さをもたらす、胃腸管に飲み込むことができることです。もう一つの広く使用されているアプローチは、直接気道への薬剤の気管や点滴を露出するために強力な麻酔下で気管切開を伴う経気管点滴、である。14しかし、だけでなく、そのようなよいです手順、その浸 ​​潤性に望ましくないこと、それはまた、時間がかかり、訓練の公平なビットを必要とし、気道に対して強力な障害を引き起こします。15,16、いくつかのプロトコルが開発されているに薬剤の直接投与を含みます外科的介入を必要とせずに気管、16,17,18,19,20が、これらの方法は、強力な麻酔薬によって引き起こされる長期の回復時間、高価な装置（ すなわち 、耳鏡/喉頭鏡、市販の手順ボード、光ファイバの使用を含みます投与量に関しては、ワイヤなど）、口腔内での操作の過剰、および不確実性。 本論文では、研究者が迅速、安価に、かつ確実に周囲の組織への残留損傷の限定されたリスクとマウスの肺に試薬を植え付けることができます挿管による投与の比較的容易な方法を記載しています。

<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.

挿管マウスは2.5％2,2,2- トリブロモエタノールの腹腔内注射を介して、10または17日後に、体重減少及び苦痛を毎日モニターし、4を屠殺した。他の場所21に記載のように気管支肺胞洗浄 （BAL）は、PBSの3回の洗浄で回収しました、および右肺は、10％ホルマリンで固定し、パラフィン包埋し、ワシントン組織学とイメージングコア22の大学の?...

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

マウスの気管の迅速な挿管のための改善された方法

この記事では、挿管を経由して、マウスの気管に直接ブレオマイシンを投与するための迅速かつ簡単な方法を提示します。この方法の主な利点は、習得が容易で、それは非常に再現可能であることであり、特殊な装置または長い回復時間を必要としません。

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

Research

JoVE Journal

Medicine

25.1K ビュー.  University of Washington. この記事では、挿管を経由して、マウスの気管に直接ブレオマイシンを投与するための迅速かつ簡単な方法を提示します。この方法の主な利点は、習得が容易で、それは非常に再現可能であることであり、特殊な装置または長い回復時間を必要としません。 

ビデオ: 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.

マウスにおける急性肺障害を誘発するために圧力制御された換気

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

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.

マウスの気管内挿管<em&gt;経由</emオトスコープの使用&gt;直接喉頭鏡検査

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.

マウス肺挿管の簡単な方法

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.

マウス肺の病理生理学を研究するための非侵襲的気管内挿管

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.

気道抵抗測定とマウスにおける気管支肺胞洗浄液のサンプリングのためのリバーシブル、非侵襲的方法

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.

マウスで横方向の大動脈狭窄を誘導するために閉じて胸モデル

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.

口腔気管挿管と人工肺虚血再灌流手術のマウスモデル

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

免疫・感染学

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.

インビボ マウス劣オリーブにおけるカルシウムイメージング

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 ...

神経科学

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.

マウスにおける反復性の耳管挿管

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>非生存ラットモデルのための気管切開術に基づく気管内挿管プロトコル

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>非生存アプリケーションのためのラットにおける気管切開術およびその後の開胸術による気管内挿管術

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.

非生存アプリケーションのためのラットにおける気管切開術およびその後の開胸術による気管内挿管術

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

マウスの気管の迅速な挿管のための改善された方法

要約

さらに動画を探す

この動画の章

マウスにおける急性肺障害を誘発するために圧力制御された換気

マウスの気管内挿管

マウス肺挿管の簡単な方法

マウス肺の病理生理学を研究するための非侵襲的気管内挿管

気道抵抗測定とマウスにおける気管支肺胞洗浄液のサンプリングのためのリバーシブル、非侵襲的方法

マウスで横方向の大動脈狭窄を誘導するために閉じて胸モデル

口腔気管挿管と人工肺虚血再灌流手術のマウスモデル

インビボマウス劣オリーブにおけるカルシウムイメージング

マウスにおける反復性の耳管挿管

非生存アプリケーションのためのラットにおける気管切開術およびその後の開胸術による気管内挿管術

マウスの気管の迅速な挿管のための改善された方法

要約

さらに動画を探す

この動画の章

マウスにおける急性肺障害を誘発するために圧力制御された換気

マウスの気管内挿管

マウス肺挿管の簡単な方法

マウス肺の病理生理学を研究するための非侵襲的気管内挿管

気道抵抗測定とマウスにおける気管支肺胞洗浄液のサンプリングのためのリバーシブル、非侵襲的方法

マウスで横方向の大動脈狭窄を誘導するために閉じて胸モデル

口腔気管挿管と人工肺虚血再灌流手術のマウスモデル

インビボ マウス劣オリーブにおけるカルシウムイメージング

マウスにおける反復性の耳管挿管

非生存アプリケーションのためのラットにおける気管切開術およびその後の開胸術による気管内挿管術

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