私たちは、ビデオ編集のための畜産用Eleutヘルナンデス、顕微鏡の専門知識のためのゲイブGaidosh、およびKhueトランとZhenyang趙にお世話になっています。国立眼研究所によって資金を供給されたこの研究は、UMにUCSDとEY014801にJLGとP30の助成金EY022589にR01-EY022129を付与。米国心臓協会、ジェームズとエスター王財団、復旦大学大学院の博士課程の学生交流プログラム基金（第2010033）、及び失明防止社への調査から無制限の助成金

すべての動物の手順は、カリフォルニア州サンディエゴ、マイアミ機関動物のケアと使用委員会（IACUC）の大学によって承認され、眼科およびVisual研究における動物の使用に関するARVO声明に従って行われました。すべての試薬及び外科的処置に使用される器具は、無菌です。 1.手術のためにラットを麻酔し、準備します<ol><li>手順の前に、ラットを体重に応じて、ケタミン（60mgの/ kg）およびキシラジン（8mg / kg）の腹腔内注射で麻酔します。麻酔の適切な深さはつま先のピンチ刺激に対する否定応答によって決定されるべきです。 </li><li>麻酔をかけた後、窒息を防止し、手術中に角膜の乾燥を防ぐために、両眼に軟膏を潤滑適用するために前方に舌を引っ張ります。 </li><li>バリカンを使用して、手術部位を剃り、面積10％ - イオジン洗剤溶液、70％エタノールで3回拭きます。 </li><li>ドレープトン無菌のフィールド内の彼は動物。滅菌手袋と手術器具は、生存手術中に使用されています。動物間のホットビーズ滅菌器を使用して、機器の先端を再滅菌します。 </li></ol> 2.外科的アプローチ<ol><li> パイオン誘導 <ol><li> 大腿静脈カテーテル法 <ol><li>手術部位を準備し、清掃してください。バリカンを使用して、右太ももの内側を剃る、10％ - イオジン洗剤溶液、70％エタノールでエリアを3回ずつを拭きます。 </li><li>チューブを準備します。 70％エタノールで滅菌ポリエチレンチューブの長さ40センチメートル（PE 10）を切断します。 （20 mg / kg体重の用量をもたらす、1μL/ mg）を生理食塩水でチューブをフラッシュし、2％のエリスロシンB色素の予め測定された溶液を含む1 mlの注射器に接続します。 600μL/分の速度に設定し、フットスイッチ制御注入ポンプにシリンジをマウントします。 </li><li> 15番ブレードを使用して、ある小さな水平切開を作ります右腿の付け根。カットし、内側の膜を広げ、滅菌綿棒でエリアを清掃してください。 </li><li>大腿静脈の枝が表示されるまで、鉗子で筋肉を区切ります。シースは、動脈、静脈や神経を囲んでいます。 Vannas春のはさみと三角形のくさびのベースの近く（2-4 mmは通常十分である）つまみ、鉗子（ファインチップデュモン鉗子）と上方このシースを引っ張って、小さな切開をカット。必要に応じてカットを展開します。 </li><li>静脈の方向に鈍いマイクロ手術用フック平行な静脈と動脈を分離します。繊細な膜と静脈枝を傷つけないように注意してください。そして、そっと静脈を持ち上げ、下にある結合組織から分離。 </li><li>ニードルレスナイロン縫合糸を入手して、大腿静脈の横に置きます。マイクロ手術用フックを使用して、静脈を上げ、先端領域の下の先の細いピンセットを渡します。縫合糸の一方の端をつかみ、下にそれを引っ張ります静脈。しっかり遠位静脈を結紮。近位静脈の下で同様の方法で、第2の縫合糸をパスし、緩い結び目を作ります。 </li><li> Vannasスプリングはさみで遠位結紮に近い静脈の小さなカットを行います。先の細いピンセットで、必要に応じて穴を展開します。一部の血液は、カットを通って漏れることがあります。冷たい、滅菌BSSおよび滅菌綿棒で手術領域を清掃してください。 </li><li>カットの縁に静脈壁を持ち、針ホルダを用いて調製し、生理食塩水で洗い流しチューブを容器にカテーテルを挿入。静脈とチューブの周りの近位結び目を締めます。次に、遠位縫合糸にそれを結びつけることによって、チューブを固定します。 </li><li> 1ミリリットルが適切である&lt;、生理食塩水を注入するために、フットスイッチを押すことにより、カテーテル法の品質を確認してください。チューブが遮られていないと漏れを持っていないことを確認します。一時的カテーテル法と組織を保護するために縫合糸で切開を閉じます。 </li></ol></li><li> 視神経の暴露 <ol><li> PreparE 10％ - イオジン洗剤溶液、70％エタノールで左目の上に各3回の事前坊主頭の領域を拭いて手術部位。 </li><li>第15ブレードで目の後ろの皮膚2〜3ミリメートルに沿って切開してください。ピンチと鋸歯状の鉗子で結合組織を持ち上げて、Vannasスプリングハサミで小さな切開を行います。この小さな切開は、一般的に長さが約5mmであるが、訓練中の外科医のための大きい露出を提供するために長くすることができます。血管を破壊しないように注意しながら、ぶっきらぼうに眼窩骨の優れたリムに沿って結合組織を通って解剖を続行します。綿棒で手術領域を清掃してください。 </li><li>優れた直筋が表示されるまで結膜を通って下方解剖。ピンチと筋肉を通して解剖。筋肉が軌道内に深くから解放されます。今、周囲の組織を視覚化を容易にするために目の後退及び仰角を助けるために利用することができます。 </li><li>再横方向と下方向に皮膚や結合組織のフラップを管、縫合や止血剤のある場所に保持します。これは、視神経を囲む脂肪含有シースを明らかにするために前進し、外側に目を回転します。 </li><li>慎重に鋭いピンセットを挿入し、シースの周囲の結合組織を分離するために視神経に平行に展開します。鉗子の鋭い先端で視神経を触れないでください。 </li><li>視神経と周囲の鞘の5ミリメートルの長さが表示されるようになるはずです。シース表面の微小血管のネットワークは、視神経を取り囲みます。これらは、レーザー照射時に対象とされます。 </li></ol></li><li> エリスロシンBおよびレーザー照射の静脈内注射 <ol><li>緑色レーザ光から身を遮蔽するためにレーザ照射装置を操作しながら、常にオレンジ色の保護眼鏡を着用してください。 、レーザーをオンにし、シャッターを開いて、ピークと平均ポウを調整必要に応じてレーザーのRS。シャッターを閉じます。 </li><li>レーザ照射装置にラットを置きます。適切なビーム配置を確実にするために、弱い照準ビームは、空間的に閉じたシャッタ羽根に穿設さ100μmの直径の穴を介してレーザをフィルタリングすることによって生成されます。先端の細いピンセットで視神経を再公開すると、3ミリメートルと視神経頭の後ろの4ミリメートルの間で眼窩内視神経に照準ビームを配置します。 </li><li>輸液ポンプの活性化を介して2％のエリスロシンBの溶液を注入します。外科医が視神経の表面を湿らせるため、BSSの小滴を追加しながら、数秒間循環することがあります。 </li><li>照準ビームの位置を確認し、照射を開始するために、フットスイッチをクリックします。顕微鏡の光路に追加されたオレンジ色の安全フィルタは、すぐに1秒の遅延後にシャッターの開口部に続くトリガされます。 </li><li> 9用の視神経を照射135mWのピーク電力および15％のデューティサイクルで250 Hzで回転するビームチョッパーによって生成18mWの平均電力と0秒（これは、熱の影響を最小限に抑えます）。安全フィルタを通して明るいオレンジ色として可視黄色蛍光は、視神経の上面から出射されるビームは対称的に、神経を照射することを保証するのに十分であることになります。 </li><li>照射後、オレンジ色の安全フィルタが自動的に開きます。微小出血がある場合には観察することができます。 </li><li>余分な眼筋の牽引力を軽減し、中立位置に目を戻します。結節縫合で切開を閉じます。その後、カテーテルを撤回し、漏出を防ぐためにしっかりと大腿静脈を縛ります。結節縫合で閉じます。両方の切開に抗生物質軟膏を適用します。網膜中心静脈と動脈の血管の完全性を検証するために眼底を確認してください。 </li></ol></li></ol></li><li> 逆行性標識法。網膜神経節細胞（RGC）の注：RGCの生存を評価するために、フルオロゴールド（FG）との逆行性標識はパイ中間子の一週間前に完了する必要があります。この方法は、Joveのプロトコル 819 13に詳細に記載されています。 <ol><li>簡単に言えば：ケタミン（60 mgの/ kg）およびキシラジン（8mg / kg）で動物を麻酔し、頭を剃ります。 </li><li>外科的切開部位をスクラブし、頭蓋骨を露出するように、ヘッド全体に正中切開を行います。 </li><li>頭蓋骨（φ2×2 mm）の両方の矢状方向および横縫合から0.5ミリメートルを通じて両国間の穴を開けます。 </li><li>慎重に真空ポンプを用いて上丘（SC）の背側表面にある脳のコンテンツを吸引。その後、SCの表面に4％のFGに浸したゲルフォームの小片を配置します。 </li><li>縫合糸で切開を閉じて、標準的な術後のケアを用いて動物の世話。 </li></ol></li><li> 術後ケアと安楽死 <ol><li>動物が回復するまでは手術後、再循環温水​​パッドの上に別々の清潔なケージに動物を配置します。 </li><li>術後鎮痛薬（ブプレノルフィン塩酸は、0.01ミリグラム/ kg）を、不快感を最小限にするために3日間連続して1日2回投与されるべきです。 </li><li>ラットを個別に保持し、彼らは胸骨横臥を維持し、十分な意識を取り戻すことができるようになるまで観察されるはずです。 </li><li>回復と健康の徴候は手術後、または抜糸と最新のいずれか早い方の手術部位の十分な治癒までに少なくとも5日間毎日監視されています。 </li><li>調査の関心に応じて、手術後に科学的に適切な時点で、4％PFAで灌流によって動物を安楽死させます。 </li></ol></li></ol>

<ol>
	<li>Wang, Y., Brown, D. P., Duan, Y., Kong, W., Watson, B. D., Goldberg, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+rodent+model+of+posterior+ischemic+optic+neuropathy..">A novel rodent model of posterior ischemic optic neuropathy..</a> JAMA Ophthalmology. 131, 194-204 (2013).</li><li>Rucker, J. C., Biousse, V., Newman, N. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Ischemic+optic+neuropathies.">Ischemic optic neuropathies.</a> Curr Opin Neurol. 17, 27-35 (2004).</li><li>Hayreh, S. 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The authors have nothing to disclose.

Here we describe in detail a method for inducing PION in a rat model. The most critical part of the protocol is the exposure and irradiation of the optic nerve &#8211; to expose the nerve as long as possible while avoiding damage caused by the sharp fine tip forceps or from stretching. In rats, the ophthalmic artery enters the optic nerve &#8804;1 mm from the optic nerve head. Therefore, irradiation of the optic nerve 3-4 mm away from the optic nerve head should only result in ischemia of the capillaries feeding the nerv...

50歳以上の患者では、虚血性視神経症（ION）は、急性視神経2の最も一般的なタイプです。条件は、特定の影響を受けた血液供給および臨床プレゼンテーションのソースに応じて2つのサブタイプの一つとして提示することができます：前方（AION）または後部（パイオン）3。 AIONの病因とコースが広く4-7研究されてきたが、パイ中間子が不十分、その低い有病率、変数プレゼンテーション、不明確な診断基準と動物モデルの不足のために理解し続けています。また、何の治療が効果的に防止またはAIONやパイオンから失明を逆転することが証明されていません。したがって、パイ中間子の再現性と信頼性の高い動物モデルは、in vivoでの疾患プロセスを研究し、神経保護および軸索再生のための新しい治療法をテストするために非常に価値があります。 光化学vasogその結果微小血管系への虚血性損傷を誘発しましたENIC浮腫および血栓症を効果的に地域の組織虚血8-12を作成します 。血管循環系に注入した後、感光性色素エリスロシンBは、標的血管のレーザー照射による活性化の際に反応性一重項酸素分子を生成します。一重項酸素は、直接血小板の接着/凝集を刺激し、血栓形成を閉塞性につながる、血管内皮をperoxidizes。虚血性損傷は、近隣地域に広がり、さらにによる血管原性浮腫に微小血管の圧縮によって悪化します。このプロトコルの全体的な目標は、光化学パイ中間子による被害をミラーリングするために球後視神経に虚血を誘導することです。 我々の知る限り、これは後部視神経1における虚血性傷害の最初のモデルです。物理的な外傷を回避しながら、このモデルは虚血を生成するように、後部の虚血性視神経症の生理学的プロセスは、より良い模倣と検討されています。また、このモデルでは、視神経障害および他のCNS虚血性疾患の候補治療薬のスクリーニングのための新たなプラットフォームを提供しています。ここで、ラットパイオンモデルにおける大腿静脈カテーテル、視神経曝露、エリスロシンBの静脈内注射とレーザー照射のための詳細な手順を説明します。

<table>
	<tbody>
		<tr>
			<td>532-nm Nd:YAG laser&#160;</td>
			<td>Laser glow</td>
			<td>LRS-532-KM-200-3</td>
			<td></td>
		</tr>
		<tr>
			<td>Beam chopper</td>
			<td>custom-made</td>
			<td>custom-made</td>
			<td></td>
		</tr>
		<tr>
			<td>Mechanical shutter and corresponding shutter drive timer</td>
			<td>AAM Vincent Associates</td>
			<td>SD-10</td>
			<td></td>
		</tr>
		<tr>
			<td>25-cm focal length spherical lens</td>
			<td>CVI/Mellles-Griot</td>
			<td>01 LPX 293</td>
			<td>plano-convexBK7 glass lens with HEBBARTM antireflection coating</td>
		</tr>
		<tr>
			<td>Erythrosin B&#160;</td>
			<td>MP Biomedicals</td>
			<td>190449</td>
			<td></td>
		</tr>
		<tr>
			<td>Fluorogold</td>
			<td>Fluorochrome,LLC</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Gelfoam</td>
			<td>Cardinal Health</td>
			<td>CAH1203421</td>
			<td></td>
		</tr>
		<tr>
			<td>Polyethylene tubing (PE10)</td>
			<td>BD Intramedic</td>
			<td>427400</td>
			<td></td>
		</tr>
		<tr>
			<td>No. 10 Blade</td>
			<td>Miltex</td>
			<td>4-110</td>
			<td></td>
		</tr>
		<tr>
			<td>Fine Forceps</td>
			<td>F.S.T.</td>
			<td>91150-20</td>
			<td>DUMONT #5 RUSTLESS NON-MAGNETIC</td>
		</tr>
		<tr>
			<td>Forceps with Teeth</td>
			<td>F.S.T.</td>
			<td>11153-10</td>
			<td>Germany stainless</td>
		</tr>
		<tr>
			<td>Forceps</td>
			<td>F.S.T.</td>
			<td>18025-10&#160;</td>
			<td>Germany stainless</td>
		</tr>
		<tr>
			<td>Vannas spring scissors</td>
			<td>F.S.T.</td>
			<td>2-220&#160;</td>
			<td>JJECK Stainless</td>
		</tr>
		<tr>
			<td>Polyglactin suture</td>
			<td>Ethicon</td>
			<td>J488G</td>
			<td>7-0 suture</td>
		</tr>
		<tr>
			<td>hemostat</td>
			<td>F.S.T.</td>
			<td>12075-12&#160;</td>
			<td>Germany stainless</td>
		</tr>
	</tbody>
</table>

rat model of photochemically-induced posterior ischemic optic neuropathy

後方虚血性視神経症（パイ中間子）は、臨床診療における視覚壊滅的な疾患です。しかし、その病因と自然史はあまり理解残っていました。最近、我々は、パイ中間子の信頼性、再現可能な動物モデルを開発し、このモデル1のいくつかの神経栄養因子の治療効果を試験しました。このビデオの目的は、後部虚血性視神経障害の我々の光化学的誘発モデルを実証するために、および網膜神経節細胞の逆行性標識して、その効果を評価することです。後部視神経、光増感色素、エリスロシンBの外科的露出の後に、静脈内に注入されたレーザ光は、視神経の表面上に集束されます。血小板血栓症および浮腫性圧縮によって媒介微小血管閉塞を促すエリスロシンBと照射損傷時のレーザ血管内皮の光化学的相互作用、。結果として虚血性傷害は緩やかなもののpronouncをもたらしますリモート、損傷誘導と臨床的に関連する結果 - 軸索入力の喪失に起因して、網膜神経節細胞の枯死を編。したがって、このモデルは、パイ中間子の病態生理学的経路を研究するための新規のプラットフォームを提供し、さらに、視神経症、ならびに他のCNSの虚血性疾患の治療アプローチを試験するために最適化することができます。

この技術によって誘導された結果として虚血性傷害は、虚血性軸索損傷後の網膜神経節細胞の緩やかなものの顕著な死をもたらします。これは、ヒトの疾患において観察されるものと同様の臨床的に関連性の結果です。 FG逆行性標識はパイオン後のRGC生存を定量するために使用されます。同じ方法を成功したモデルの作成 ​​を検証するだけでなく、異なる治療法の効果を評価するために使...

Watch this Scientific Journal Video about Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy at JoVE.com

Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

このプロトコルの目的は、光化学ラットにおける後部視神経に虚血損傷を誘導することです。このモデルは、後部の虚血性視神経症の病態生理学の研究、およびこれと他の視神経障害のためだけでなく、他のCNSの虚血性疾患の治療的アプローチに重要です。

光化学誘起後方虚血性視神経障害のラットモデル

Posterior Ischemic optic neuropathy (PION) is a sight-devastating disease in clinical practice. However, its pathogenesis and natural history ...

rat-model-photochemically-induced-posterior-ischemic-optic

Research

JoVE Journal

Medicine

8.5K ビュー.  University of Miami Miller School of Medicine. このプロトコルの目的は、光化学ラットにおける後部視神経に虚血損傷を誘導することです。このモデルは、後部の虚血性視神経症の病態生理学の研究、およびこれと他の視神経障害のためだけでなく、他のCNSの虚血性疾患の治療的アプローチに重要です。 

ビデオ: Rat Model of Photochemically-Induced Posterior Ischemic Optic Neuropathy

Creation of Reversible Cholestatic Rat Model

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Cholestasis can cause various physiological changes and affect the nutritional status and surgical outcomes. Study of the pathophysiological changes occurring in the liver and other organs is of importance. Various studies have been done in cholestatic rat models.
We used a reversible cholestatic rat model in our recent study looking at the role of methylprednisolone in the ischemia reperfusion injury. Various techniques for creation of a reversible cholestatic model have been described. Creation of a reversible cholestatic rat model can be challenging in view of the smaller size and unique hepatopancreatobiliary anatomy in rats. This video article demonstrates the creation of a reversible cholestatic model.
This model can be used in various studies, such as looking at the changes in nutritional, physiological, pathological, histological and immunological changes in the gastrointestinal tract. This model can also be used to see the effects of cholestasis and various therapeutic interventions on major hepatic surgeries.

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Creation of a reversible cholestatic rat model can be challenging in view of the smaller size and unique hepatopancreatobiliary anatomy in rats. This video article demonstrates the creation of a reversible cholestatic model.

リバーシブル胆汁うっ滞ラットモデルの作成

Cholestasis is a clinical condition commonly encountered by both surgeons and gastroenterologists. Cholestasis can cause various physiological changes and ...

医学

Gene Transfer for Ischemic Heart Failure in a Preclinical Model

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine their efficacy and safety.
Gene therapy using viral vectors is one of the most promising approaches for treating cardiac diseases. Viral delivery of various different genes by changing the carrier gene has immeasurable therapeutic potential.
In this video, the full process of an animal model of heart failure creation followed by gene transfer is presented using a swine model. First, myocardial infarction is created by occluding the proximal left anterior descending coronary artery. Heart remodeling results in chronic heart failure. Unique to our model is a fairly large scar which truly reflects patients with severe heart failure who require aggressive therapy for positive outcomes. After myocardial infarct creation and development of scar tissue, an intracoronary injection of virus is demonstrated with simultaneous nitroglycerine infusion. Our injection method provides simple and efficient gene transfer with enhanced gene expression. This combination of a myocardial infarct swine model with intracoronary virus delivery has proven to be a consistent and reproducible methodology, which helps not only to test the effect of individual gene, but also compare the efficacy of many genes as therapeutic candidates.

A method of gene transfer for the treatment of ischemic heart failure is described using a swine model of myocardial infarction. Our simple and reproducible method enables us to readily evaluate the efficacy of various gene transfers with a very simple and reproducible way.

前臨床モデルにおける虚血性心不全のための遺伝子導入

Various emerging technologies are being developed for patients with heart failure. Well-established preclinical evaluations are necessary to determine ...

A Simplified Technique for Producing an Ischemic Wound Model

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work well in non-ischemic wounds may not work in human diabetic wounds because vasculopathy is one major factor that hinders healing of these wounds. We published an article in 2007 describing a rabbit ear ischemic wound model created by a minimally invasive surgical technique. Since then, we have further simplified the procedure for easier operation. On one ear, three small skin incisions were made on the vascular pedicles, 1-2 cm from the ear base. The central artery was ligated and cut along with the nerve. The whole cranial bundle was cut and ligated, leaving only the caudal branch intact. A circumferential subcutaneous tunnel was made through the incisions, to cut subcutaneous tissues, muscles, nerves, and small vessels. The other ear was used as a non-ischemic control. Four wounds were made on the ventral side of each ear. This technique produces 4 ischemic wounds and 4 non-ischemic wounds in one animal for paired comparisons. After surgery, the ischemic ear was cool and cyanotic, and showed reduced movement and a lack of pulse in the ear artery. Skin temperature of the ischemic ear was 1-10 &deg;C lower than that on the normal ear and this difference was maintained for more than one month. Ear tissue high-energy phosphate contents were lower in the ischemic ear than the control ear. Wound healing times were longer in the ischemic ear than in the non-ischemic ear when the same treatment was used. The technique has now been used on more than 80 rabbits in which 23 were diabetic (diabetes time ranging from 2 weeks to 2 years). No single rabbit has developed any surgical complications such as bleeding, infection, or rupture in the skin incisions. The model has many advantages, such as little skin disruption, longer ischemic time, and higher success rate, when compared to many other models. It can be safely used in animals with reduced resistance, and can also be modified to meet different testing requirements.

We have developed a minimally invasive technique to create a rabbit ischemic ear wound model by dividing the central artery and nerve and the cranial neurovascular bundle. A subcutaneous tunnel then cuts all subcutaneous tissues. This procedure causes minimal skin disruption and can be safely used in diabetic animals.

虚血性創傷モデルを作製するための簡略化手法

One major obstacle in current diabetic wound research is a lack of an ischemic wound model that can be safely used in diabetic animals. Drugs that work ...

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.

ラットにおけるレーザードップラー血流計の指針と虚血性脳卒中のエンドセリン-1誘発性中大脳動脈閉塞モデル

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 ...

Angiogenesis in the Ischemic Rat Lung

The adult lung is perfused by both the systemic bronchial artery and the entire venous return flowing through the pulmonary arteries. In most lung pathologies, it is the smaller systemic vasculature that responds to a need for enhanced lung perfusion and shows robust neovascularization. Pulmonary vascular ischemia induced by pulmonary artery obstruction has been shown to result in rapid systemic arterial angiogenesis in man as well as in several animal models. Although the histologic assessment of the time course of bronchial artery proliferation in rats was carefully described by Weibel 1, mechanisms responsible for this organized growth of new vessels are not clear. We provide surgical details of inducing left pulmonary artery ischemia in the rat that leads to bronchial neovascularization. Quantification of the extent of angiogenesis presents an additional challenge due to the presence of the two vascular beds within the lung. Methods to determine functional angiogenesis based on labeled microsphere injections are provided.

The lung is perfused by both the systemic bronchial artery and pulmonary arteries. In most lung pathologies, it is the smaller systemic vasculature that shows robust neovascularization. Cessation of pulmonary blood flow promotes brisk bronchial angiogenesis. We provide surgical details of inducing left pulmonary artery ischemia that promotes bronchial neovascularization.

虚血ラットの肺の血管新生

The  adult lung is perfused by both the systemic bronchial artery and the entire  venous return flowing through the pulmonary arteries. In most lung ...

A Piglet Model of Neonatal Hypoxic-Ischemic Encephalopathy

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the world&#8217;s 2.9 million neonatal deaths. Animal models of HIE have contributed to the understanding of the pathophysiology in HIE, and have highlighted the dynamic process that occur in brain injury due to perinatal asphyxia. Thus, animal studies have suggested a time-window for post-insult treatment strategies. Hypothermia has been tested as a treatment for HIE in pdiglet models and subsequently proven effective in clinical trials. Variations of the model have been applied in the study of adjunctive neuroprotective methods and piglet studies of xenon and melatonin have led to clinical phase I and II trials1,2. The piglet HIE model is further used for neonatal resuscitation- and hemodynamic studies as well as in investigations of cerebral hypoxia on a cellular level. However, it is a technically challenging model and variations in the protocol may result in either too mild or too severe brain injury. In this article, we demonstrate the technical procedures necessary for establishing a stable piglet model of neonatal HIE. First, the newborn piglet (&#60; 24 hr old, median weight 1500 g) is anesthetized, intubated, and monitored in a setup comparable to that found in a neonatal intensive care unit. Global hypoxia-ischemia is induced by lowering the inspiratory oxygen fraction to achieve global hypoxia, ischemia through hypotension and a flat trace amplitude integrated EEG (aEEG) indicative of cerebral hypoxia. Survival is promoted by adjusting oxygenation according to the aEEG response and blood pressure. Brain injury is quantified by histopathology and magnetic resonance imaging after 72 hr.

Hypoxic-ischemic encephalopathy following perinatal asphyxia can be studied using animal models. We demonstrate the procedures necessary for establishing a piglet model of neonatal hypoxic-ischemic encephalopathy.

新生児低酸素性虚血性脳症の子豚モデル

Birth asphyxia, which causes hypoxic-ischemic encephalopathy (HIE), accounts for 0.66 million deaths worldwide each year, about a quarter of the ...

Demonstration of the Rat Ischemic Skin Wound Model

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and unrelieved pressure due to immobility. Animal models have been developed that attempt to mimic these conditions for the purpose of furthering our understanding of the complexity of chronic wounds. The model described herein is a rat ischemic skin flap model that permits a prolonged reduction of blood flow resulting in wounds that become ischemic and resemble a chronic wound phenotype (reduced vascularization, increased inflammation and delayed wound closure). It consists of a bipedicled dorsal flap with 2 ischemic wounds placed centrally and 2 non-ischemic wounds lateral to the flap as controls. A novel addition to this ischemic skin flap model is the placement of a silicone sheet beneath the flap that functions as a barrier and a splint to prevent revascularization and reduce contraction as the wounds heal. Despite the debate of using rats for wound healing studies due to their quite distinct anatomic and physiologic differences compared to humans (i.e., the presence of a panniculus carnosus muscle, short life-span, increased number of hair follicles, and their ability to heal infected wounds) the modifications employed in this model make it a valuable alternative to previously developed ischemic skin flap models.

The rat, due to its size, availability, and rather docile behavior, has been utilized as a research model for many years. The goal of this protocol is to utilize the rat as an ischemic skin wound healing model to provide valuable insight into the pathophysiology of chronic wounds.

ラット虚血皮膚創傷モデルの実証

The propensity for chronic wounds in humans increases with ageing, disease conditions such as diabetes and impaired cardiovascular function, and ...

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy (rNAION)

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their lifetime. NAION results in optic nerve edema, selective loss of the retinal ganglion cell neurons (RGCs) and atrophy of the optic nerve. A rodent model of NAION that expresses most NAION features and sequelae has been developed, which is applicable to both rats and mice. This model utilizes a focal laser application of 532 nm wavelength to illuminate a photoactive dye, Rose Bengal (RB), to cause capillary damage and leakage at the targeted anterior optic nerve (the laminar region). After rNAION induction, there is an early optic nerve ischemia, optic nerve edema, and intraneural inflammation, followed by selective RGC and axonal loss. Since the optic nerve is a CNS white matter tract, the rNAION model is applicable to mechanistic studies of selective white matter ischemia, as well as neuroprotective analyses and short and long-term mechanisms of glial and neuronal response to ischemia.

The Rodent Model of Nonarteritic Anterior Ischemic Optic Neuropathy rNAION

The following report describes how to replicate the rodent model of nonarteritic anterior ischemic optic neuropathy (rNAION), using the appropriate dye, contact lens and laser parameters. We also reveal the appropriate steps for evaluating the rNAION lesion in vivo.

Nonarteritic前部虚血性視神経症の齧歯類モデル（rNAION）

Nonarteritic anterior ischemic optic neuropathy (NAION) is a focal ischemic lesion of the optic nerve that affects 1/700 individuals throughout their ...

Isometric Contractility Measurement of the Mouse Mesenteric Artery Using Wire Myography

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and drugs. It is widely used in many studies on the physiological functions of vascular smooth muscle, the pathogenesis of vascular diseases such as hypertension, and the development of smooth muscle relaxant drugs. The mouse is a widely used model animal with a large pool of disease models and genetically modified strains. We introduced this method to measure the isometric contraction of mouse mesenteric artery in detail. A 1.4-mm segment of mouse mesenteric resistance artery was isolated and mounted on a myograph chamber by passing two steel wires through its lumen. After equilibration and normalization steps, the vessel segment was potentiated by a high-K+ solution twice prior to the contraction assay. As an example of the application of this method in drug development, we measured the relaxant effect of a novel natural substance, neoliensinine, isolated from a Chinese herb, embryos of the lotus seed (Nelumbo nucifera Gaertn.) on mouse mesenteric arteries. The vessel segments mounted on the myograph chamber were stimulated with a high-K+ solution. When the force tension reached a stable sustained phase, cumulative doses of neoliensinine were added to the chamber. We found that neoliensinine had a dose-dependent relaxant effect on smooth muscle contraction, thus suggesting that it bears potential activity against hypertension. In addition, as the vessel segment can survive at least 4 hours after mounting and maintain contractility induced by the high-K+ solution for many times, we suggest that the wire myograph system may be used for the time-consuming process of drug screening.

The wire myograph technique is used to investigate vascular smooth muscle functions and screen new drugs. We report a detailed protocol for measuring the isometric contractility of the mouse mesenteric artery and for screening new relaxants of vascular smooth muscle.

ワイヤー図法を使用してマウスの腸間膜動脈の等尺性収縮力測定

The wire myograph technique is used to assess the contractility of vascular smooth muscles in response to depolarization, GPCR agonists/inhibitors and ...

Oxygen-Induced Retinopathy Model for Ischemic Retinal Diseases in Rodents

One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR model induction and its readouts in both mice and rats. Retinal neovascularization is induced in OIR by exposing rodent pups either to hyperoxia (mice) or alternating levels of hyperoxia and hypoxia (rats). The primary readouts of these models are the size of neovascular (NV) and avascular (AVA) areas in the retina. This preclinical in vivo model can be used to evaluate the efficacy of potential anti-angiogenic drugs or to address the role of specific genes in the retinal angiogenesis by using genetically manipulated animals. The model has some strain and vendor specific variation in the OIR induction which should be taken into consideration when designing the experiments.

Oxygen-induced retinopathy (OIR) can be used to model ischemic retinal diseases such as retinopathy of prematurity and proliferative diabetic retinopathy and to serve as a model for proof-of-concept studies in evaluating antiangiogenic drugs for neovascular diseases. OIR induces robust and reproducible neovascularization in the retina that can be quantified.

げっ歯類における虚血性網膜症の酸素誘導網膜症モデル

One of the commonly used models for ischemic retinopathies is the oxygen-induced retinopathy (OIR) model. Here we describe detailed protocols for the OIR ...