DK065872（ENP）、医学研究の卓越性のスミスファミリー財団賞（ENP）、F31 DA023760によってサポートされています。

まとめとキシラジン（10mgを、2ヶ月の古い平均年齢のC57BL/6Jマウスまたは同等以上の三ヶ月の平均年齢Sprague Dawleyラットまたは同等のケタミン（マウスでは100 mg / kgをIPラットで60 mg / kgを腹腔内）で麻酔/ kgのいずれかの種のためのIP）。鎮静は反射がで示した撤退穏やかなつま先のピンチを使用して監視されます<a href="/index/details.stp?ID=236">。Walantusら</a> （Joveの、6、2007）と<a href="/index/details.stp?ID=404">Szotら</a> （Joveの、9、2007）。体温調節はthermostatregulated加熱パッド（ALAインスツル株式会社）を介して提供し、直腸の温度計で監視することができます。ヘッドは、毛皮の剃毛と切開の前にヨウ素で洗浄される。皮膚切開（マウスの場合は1センチのラット、2センチ）と頭蓋骨の表面からすべての軟組織を除去した後、ガイドカニューレの配置は、ブレグマとの関係で決定されます。 6mmの穴はげっ歯類の手術のために設計されたバッテリ駆動ドリラー（ファイン科学のツール、（株））で頭蓋骨貫通されています。ケアは、ドリルビットが髄膜や血管を貫通しないように取られます。頭蓋骨は、後側坐核、背側線条体や前頭前野に至る二国間の5ミリメートル21ゲージステンレス製のガイドシャフトに注入される。定位座標はフランクリンとPaxinos、1997年（定位座標でのマウスの脳、アカデミックプレス）またはPaxinosとワトソン、2006（定位座標でのラットの脳、ア​​カデミックプレス）あたりとして確立されています。インプラントは歯科用セメントで固定されている。 0.9％生理食塩水の乳酸リンゲルのボーラスは、脱水を防ぐために、手術の終わり（ラットにおける5mls SCとマウスの1ミリリットルSC流体は通常の体温に温めされた後）で与えられる。動物が苦痛にあると表示される場合ブプレノルフィン（0.1-0.5mg/kg SC）は、必要に応じて、その後、1日2回投与とされています。術後感染症が発生した場合、ローカル抗生物質治療（バシトラシン軟膏）と抗生物質の全身治療（最初の48時間術後12時間ごとにIMペニシリン100,000 IU / kg）を投与されています。 手術後、動物は個別に食料と水利用可能な自由に摂取して収容されています。少なくとも一週間は、マイクロダイアリシスと安楽死の前に回復のために許可されています。手術から回復後、動物は、マイクロダイアリシスケージとマイクロダイアリシスプローブに転送され、手術中にインストールされているガイドシャフトに挿入して接合されている。プローブは、ガイド軸を介して皮膚、筋肉および髄膜の組織をバイパスされているため、プローブの挿入は、痛みや不快感が発生することはありません。そのため、プローブの挿入は、麻酔なしで行われ、神経化学または行動上の任意の麻酔誘発性の影響が回避されています。我々は、プローブは12時間安定化し、我々が実験に応じて別の8〜12時間30分ごとのサンプリングを開始させて。我々はフォトセルまたは実験による移動の手動録画を介して動物の運動行動を監視する。 Microdialysateサンプルは神経化学的検出と解析のための電気化学検出（HPLC - EC）の測定器で高速液体クロマトグラフィーに注入されています。我々は、基礎神経化学と運動行動への影響を探します。実験の終了時に動物が全身ケタミン（200 mg / kgを腹腔内）とキシラジン（20 mg / kgを、IP）の過剰投与により安楽死されています。その後、心臓を4％パラホルムアルデヒドに続いて0.9％生理食塩水で灌流される。脳を取り出し、凍結と正確なプローブの配置を確認するためにマイクロダイアリシスプローブの管に沿ってカットされています。 手順<ol><li>定位固定装置と必要なすべてのマテリアルを設定します。エリアや楽器のクリーニングと滅菌されていることを確認します。 </li><li>電気かみそりで毛を剃り落とす。耳からだけで、間に目に移動し、毛皮の領域を効果的にきれいに異なる方向にカミソリを動かす。剃毛面積にpovidine /ヨウ素を適用するが、それから目を保護する。 </li><li>外耳道に耳のバーを配置し、所定の位置に締めて、定位固定装置の上に動物をマウントします。その後、最初のマウント外耳道の片方の耳のバー、および他の耳のバーの位置とスライドでそれを保持。あなたは頭がもう左右に移動することができる正しい場所にある知っている。定位の前部マウントで口を固定し、ヘッドが定規と水平であることを確認してください。定位固定装置のプラットフォームに対して垂直の位置に定規を置くと）動物の頭皮の定規と真ん中の間に90 °の角度を確認してください。それはそのような機能を提供する場合定位固定装置を介してこれを確認してください。 </li><li>ラムダからわずか動物の眼の間に延びる滅菌メスで頭皮上に前部/後部の切開を加えます。皮膚をピンチオフと切開を開いたままにして滅菌止血を使用してください。いくつかの滅菌綿棒を使用して、露出頭蓋骨の表面をふいて乾かす。 <li>この場所をガイドカニューレの右側頭骨上ブレグマ、および位置を見つけ、そのマウントの上にガイドカニューレを置く。後方/前方と横方向の座標を書き留めておきます。ブレグマから、脳地図の助けを借りて、あなたのプローブの配置に必要な正しい座標を見つける。追加またはブレグマから減算することによって、正しい座標にガイドカニューレを置きます。それがこの腹側の座標を記録して頭蓋骨に触れ、そしてされるまで、あなたのガイドカニューレをダウンさせる。頭蓋骨上のこの場所で滅菌鉛筆で鉛筆のマークを付ける、これが掘削される場所です。 </li><li>ガイドカニューレを外し、ドリルビットを殺菌する。あなたが頭蓋骨の幅を介して取得されるまで慎重に鉛筆のマークに穴を開ける。それは側面に触れることなく穴をクリアするだろうかどうかを確認するためのガイドカニューレに確認してください。カニューレはストレートパスでクリアできるようになるまで掘削し、定期的にチェックしてください。穴が行われると、静かにカニューレの遮るもののない挿入を可能にするために、髄膜を開ける滅菌針を使用してください。 </li><li>カニューレの穴に2つの前方に、二つのカニューレの穴、側面の後方に2〜横：次に、ハンドドリルを使用すると、頭蓋骨のネジのための6つの穴を作成します。 6本のネジを滅菌し、それらがしっかりと上に固定されるまで、頭蓋骨の上に置きます。 </li><li>エタノールおよび生理食塩水、マウント付きガイドカニューレを清掃し、適切な腹側の座標にゆっくりそれを下げて。双方が接触していないこと、それが完全に垂直に起きていることを確認してください。 </li><li>内側および後頭骨のネジの後ろにアンカーボルトを配置し、ピンセットで所定の位置に固定します。液体歯科用セメントの薄いバッチを混合し、ガイドカニューレ、ネジ、および滅菌スパチュラで頭蓋骨の残りをカバーしています。 、別のバッチを作る今回は厚く、そして完全にエリアとそれを確保するのに十分なカニューレとアンカーネジをカバーする。 </li><li>セメントが厚く、それが固まる前になると、セメントのカップから皮膚を分離し、セメントのキャップは、すべての周りに滑らかであると後で皮膚を刺激しないことを確認するためにスパチュラでセメントのカップを成形。 </li><li>歯科用セメントが完全に装置から動物を削除する前に乾燥することができます。止血を削除します。バシトラシンのセメントのキャップの周りのすべての方法を適用する。 </li><li>動物は、定位固定装置をオフになると、生理食塩水SC（皮下）を1ml、続いペニシリンIM（筋肉内）0.25mlのでそれを注入する。 </li><li>独自のケージに動物を配置し、それを回復するために、その部屋にそれを返す前に意識的になるまでそれを監視する。 </li><li>彼らは手術と術後日常の日に麻酔から回復するまで痛み/苦痛の感染と評価の兆候を、実験の終わりまで、動物を監視します。これは、週末および休日を含んでいる。低自発運動、取り扱い時に遭難発声、猫背の姿勢、下痢、headmountの領域の腫れと放電、および摂食/飲料の欠如は、痛みや苦痛のすべての兆候です。動物が苦痛にあると表示される場合ブプレノルフィン（0.1-0.5mg/kg SC）は、必要に応じて、毎日二度、そしてその後に投与される。術後感染症が発生した場合、ローカル抗生物質治療（バシトラシン軟膏）と抗生物質の全身治療（最初の48時間術後12時間ごとにIMペニシリン100,000 IU / kg）を投与されています。これらの症状のいずれかが手術の12時間以内にブプレノルフィンの投与後、補足的な流体、および抗生物質治療を継続する場合は、動物を安楽死されている。 </li></ol>

<ol>
	<li>Bungay, P. M., Newton-Vinson, P., Isele, W., Garris, P. A., Justice, J. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Microdialysis+of+dopamine+interpreted+with+quantitative+model+incorporating+probe+implantation+trauma.">Microdialysis of dopamine interpreted with quantitative model incorporating probe implantation trauma.</a> J. Neurochem. 86, 932-946 (2003).</li><li>Chen, K. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effects+of+tissue+trauma+on+the+characteristics+of+microdialysis+zero-net-flux+method+sampling+neurotransmitters.">Effects of tissue trauma on the characteristics of microdialysis zero-net-flux method sampling neurotransmitters.</a> Journal of Theor. Biology. 238, 863-881 (2006).</li><li>Geiger, B. M., Behr, G. G., Frank, L., Caldera-Siu, A. D., Beinfeld, M. C., Kokkotou, E. G., Pothos, E. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Evidence+for+defective+mesolimbic+dopamine+exocytosis+in+obesity-prone+rats.">Evidence for defective mesolimbic dopamine exocytosis in obesity-prone rats.</a> FASEB Journal. 8, 2740-2746 (2008).</li><li>Pothos, E. N., Creese, I., Hoebel, B. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Restricted+eating+with+weight+loss+selectively+decreases+extracellular+dopamine+in+the+nucleus+accumbens+and+alters+dopamine+response+to+amphetamine,+morphine+and+food+intake.">Restricted eating with weight loss selectively decreases extracellular dopamine in the nucleus accumbens and alters dopamine response to amphetamine, morphine and food intake.</a> The Journal of Neuroscience. 15, 6640-6650 (1995).</li></ol>

The authors have nothing to disclose.

生体内微小透析で生きている動物の異なる脳部位で複数の神経伝達物質や代謝産物を測定するための最適なツールです。しかし、それは唯一の神経化学物質の細胞外レベルを監視し、リアルタイムで神経伝達物質のエキソサイトーシスを監視するための時間分解能を提供していません。 &quot;ネットフラックス&quot;と呼ばれるテクニックのバージョンを通じて、与えられたサイトでの実際の神経伝達物質の濃...

材料は、プロトコルドキュメントで説明されています

survivable stereotaxic surgery in rodents 

目を覚まし、動物の脳内の神経伝達物質の細胞外基底レベルを測定する能力は、中枢神経系へのさまざまな全身性の課題の影響（薬理学的または生理学）の測定が可能になります。例えば、一つは、直接動物の中脳のドーパミンの予測は、食品のようなd -アンフェタミンまたは天然の刺激のようなドーパミン放出薬に反応方法測定することができます。このビデオでは、我々はマイクロダイアリシスプローブを挿入して移植する方法、ラットの脳内の特定のサイトをターゲットにガイドカニューレを注入する方法を説明し、細胞外レベルのを測定する電気化学検出と相まって、高性能液体クロマトグラフィー（HPLC - EC）を使用する方法酸化神経伝達物質と代謝物。マイクロダイアリシスプローブを介して薬物のローカル正確な導入は、アクションの化合物のメカニズムの部位特異性で洗練された作業が可能になります。この手法は、優れた解剖学的および化学的分解能を持つが、マイクロダイアリシスサンプルなどのわずかな時間分解能は、通常、検出可能な神経伝達物質のレベルを確保するために毎20-30分を処理されます。補完的な生体外でのツールは（すなわち、スライスや細胞培養、電気生理学）リアルタイムの神経伝達を監視を支援することができます。

Watch this Scientific Journal Video about Survivable Stereotaxic Surgery in Rodents at JoVE.com

Survivable Stereotaxic Surgery in Rodents

自由に動物を移動させるの異なる脳領域における細胞外神経伝達物質レベルのモニタリングは、神経伝達物質の放出と行動の間のリンク上の洞察を提供しています。および基底神経伝達を薬理学的または生理学的操作によって変更される方法については、電気化学検出と相まって生体微量透析に優れた解剖学的および化学的分解能を提供します。

げっ歯類で存続可能な定位手術

The ability to measure extracellular basal levels of neurotransmitters in the brain of awake animals allows for the determination of effects of different ...

survivable-stereotaxic-surgery-in-rodents

Research

JoVE Journal

Biology

Survivable Stereotaxic Surgery in Rodents 

68.3K ビュー.  Tufts University. 自由に動物を移動させるの異なる脳領域における細胞外神経伝達物質レベルのモニタリングは、神経伝達物質の放出と行動の間のリンク上の洞察を提供しています。および基底神経伝達を薬理学的または生理学的操作によって変更される方法については、電気化学検出と相まって生体微量透析に優れた解剖学的および化学的分解能を提供します。

ビデオ: Survivable Stereotaxic Surgery in Rodents

Heterotopic Heart Transplantation in Mice

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable for studying rejection and immune response now that newer transgenic and gene knockout mice are available, and a large number of immunologic reagents have been developed. The heart transplant model is less stringent than the skin transplant models, although technically more challenging. We have developed a modified technique and have completed over 1000 successful cases of heterotopic heart transplantation in mice. When making anastomosis of the ascending aorta and abdominal aorta, two stay sutures are placed at the proximal and distal apexes of recipient abdominal aorta with the donor s ascending aorta, then using 11-0 suture for anastomosis on both side of aorta with continuing sutures. The stay sutures make the anastomosis easier and 11-0 is an ideal suture size to avoid bleeding and thrombosis.
When making anastomosis of pulmonary artery and inferior vena cava, two stay sutures are made at the proximal apex and distal apex of the recipient s inferior vena cava with the donor s pulmonary artery. The left wall of the inferior vena cava and donor s pulmonary artery is closed with continuing sutures in the inside of the inferior vena cava after, one knot with the proximal apex stay suture the right wall of the inferior vena cava and the donor s pulmonary artery are closed with continuing sutures outside the inferior vena cave with 10-0 sutures. This method is easier to perform because anastomosis is made just on the one side of the inferior vena cava and 10-0 sutures is the right size to avoid bleeding and thrombosis. In this article, we provide details of the technique to supplement the video.

The mouse heterotopic heart transplantation model has been proven by many investigators to be an important method for studying mechanisms of rejection and immune response. However, the techniques involved are still challenging. By modifying standard techniques we have had success with more than 1000 transplants.

マウスにおける異所心臓移植

The mouse heterotopic heart transplantation has been used widely since it was introduced by Drs. Corry and Russell in 1973. It is particularly valuable ...

生物学

Small Bowel Transplantation In Mice

Since 1990, the development of tacrolimus-based immunosuppression and improved surgical techniques, the increased array of potent immunosuppressive medications, infection prophylaxis, and suitable patient selection helped improve actuarial graft and patient survival rates for all types of intestine transplantation. Patients with irreversible intestinal failure and complications of parenteral nutrition should now be routinely considered for small intestine transplantation. However, Survival rates for small intestinal transplantation have been slow to improve compares increasingly favorably with renal, liver, heart and lung. The small bowel transplantation is still unsatisfactory compared with other organs. Further progress may depend on better understanding of immunology and physiology of the graft and can be greatly facilitated by animal models. A wider use of mouse small bowel transplantation model is needed in the study of immunology and physiology of the transplantation gut as well as efficient methods in diagnosing early rejection. However, this model is limited to use because the techniques involved is an extremely technically challenging. We have developed a modified technique. When making anastomosis of portal vein and inferior vena cava, two stay sutures are made at the proximal apex and distal apex of the recipient s inferior vena cava with the donor s portal vein. The left wall of the inferior vena cava and donor s portal vein is closed with continuing sutures in the inside of the inferior vena cava after, after one knot with the proximal apex stay suture the right wall of the inferior vena cava and the donor s portal vein are closed with continuing sutures outside the inferior vena cave with 10-0 sutures. This method is easier to perform because anastomosis is made just on the one side of the inferior vena cava and 10-0 sutures is the right size to avoid bleeding and thrombosis.  In this article, we provide details of the technique to supplement the video.

The mouse small bowel transplantation model has been recognized as an important tool to study mechanismes of immune rejection and screen new immunosuppressive drugs. However, this model is limited to use because the techniques involved is an extremely technically challenge. Now we introduce the modified technique.

マウスにおける小腸移植

Since 1990, the development of tacrolimus-based immunosuppression and improved surgical techniques, the increased array of potent immunosuppressive ...

A Craniotomy Surgery Procedure for Chronic Brain Imaging

Imaging techniques are becoming increasingly important in the study brain function. Among them, two-photon laser scanning microscopy has emerged as an extremely useful method, because it allows the study of the live intact brain. With appropriate preparations, this technique allows the observation of the same cortical area chronically, from minutes to months. In this video, we show a preparation for chronic in vivo imaging of the brain using two-photon microscopy. This technique was initially pioneered by Dr. Karel Svoboda, who is now a Howard Hughes Medical Institute Investigator at Janelia Farm. Preparations like the one shown here can be used for imaging of neocortical structure (e.g., dendritic and axonal dynamics),  to record neuronal activity using calcium-sensitive dyes, to image cortical blood flow dynamics, or for intrinsic optical imaging studies. Deep imaging of the neocortex is possible with optimal cranial window surgeries. Operating under the most sterile conditions possible to avoid infections, together with using extreme care to do not damage the dura mater during the surgery, will result in successful and long-lasting glass-covered cranial windows.


This video and protocol demonstrate how to implant a glass-covered cranial window in rodents. These preparations can be used for chronic in vivo two-photon imaging of the neocortex over time scales of months. It may also be used for other types of imaging, including optical intrinsic signal imaging.

慢性脳イメージングのための開頭手術の手順

Imaging techniques are becoming increasingly important in the study brain function. Among them, two-photon laser scanning microscopy has emerged as an ...

Dissecting the Non-human Primate Brain in Stereotaxic Space

The use of non-human primates provides an excellent translational model for our understanding of developmental and aging processes in humans1-6. In addition, the use of non-human primates has recently afforded the opportunity to naturally model complex psychiatric disorders such as alcohol abuse7. Here we describe a technique for blocking the brain in the coronal plane of the vervet monkey (Chlorocebus aethiops sabeus) in the intact skull in stereotaxic space. The method described here provides a standard plane of section between blocks and subjects and minimizes partial sections between blocks. Sectioning a block of tissue in the coronal plane also facilitates the delineation of an area of interest. This method provides manageable sized blocks since a single hemisphere of the vervet monkey yields more than 1200 sections when slicing at 50&mu;m. Furthermore by blocking the brain into 1cm blocks, it facilitates penetration of sucrose for cyroprotection and allows the block to be sliced on a standard cryostat.

The non-human primate is an important translational species for our understanding of the normal processing of the brain. The anatomical organization of the primate brain can provide important insights into normal and pathological conditions in humans.

定位空間における非ヒト霊長類の脳を解剖

The use of non-human primates provides an excellent translational model for our understanding of developmental and aging processes in humans1-6. In ...

Establishing Primary Adult Fibroblast Cultures From Rodents

The importance of using primary cells, rather than cancer cell lines, for biological studies is becoming widely recognized. Primary cells are preferred in studies of cell cycle control, apoptosis, and DNA repair, as cancer cells carry mutations in genes involved in these processes. Primary cells cannot be cultured indefinitely due to the onset of replicative senescence or aneuploidization. Hence, new cultures need to be established regularly. The procedure for isolation of rodent embryonic fibroblasts is well established, but isolating adult fibroblast cultures often presents a challenge. Adult rodent fibroblasts isolated from mouse models of human disease may be a preferred control when comparing them to fibroblasts from human patients. Furthermore, adult fibroblasts are the only available material when working with wild rodents where pregnant females cannot be easily obtained. Here we provide a protocol for isolation and culture of adult fibroblasts from rodent skin and lungs. We used this procedure successfully to isolate fibroblasts from over twenty rodent species from laboratory mice and rats to wild rodents such as beaver, porcupine, and squirrel.

This article describes a protocol for isolation and maintenance of primary fibroblast cultures from skin and lung tissue of wild rodents.

げっ歯類からプライマリ大人の線維芽細胞培養の確立

The importance of using primary cells, rather than cancer cell lines, for biological studies is becoming widely recognized.  Primary cells are preferred ...

Principles of Rodent Surgery for the New Surgeon

For both scientific and animal welfare reasons, training in basic surgical concepts and techniques should be undertaken before ever seeking to perform surgery on a rodent. Students, post-doctoral scholars, and others interested in performing surgery on rodents as part of a research protocol may not have had formal surgical training as part of their required coursework. Surgery itself is a technical skill, and one that will improve with practice. The principles of aseptic technique, however, often remain unexplained or untaught. For most new surgeons, this vital information is presented in piecemeal fashion or learned on the job, neither of which is ideal. It may also make learning how to perform a particular surgery difficult, as the new surgeon is learning both a surgical technique and the principles of asepsis at the same time. This article summarizes and makes recommendations for basic surgical skills and techniques necessary for successful rodent surgery. This article is designed to supplement hands-on training by the user's institution.

Before attempting surgery, a new surgeon should have training in basic surgical techniques and concepts. This article will present basic surgical considerations with an emphasis on rodents.

新外科医のための齧歯類の外科の原則

For both scientific and animal welfare reasons, training in basic surgical concepts and techniques should be undertaken before ever seeking to perform ...

Intracavernosal Pressure Recording to Evaluate Erectile Function in Rodents

Erectile dysfunction (ED) is defined as the inability to attain or keep an erection of the penis, and this has become a prevalent male sexual disorder. Rodents are employed by many studies to research the physiology/pathology of erectile function. Erectile function in rodents can be evaluated by measuring the intracavernosal pressure (ICP). In practice, ICP can be monitored following electrical stimulation of the cavernous nerves (CNs). The arterial pressure of the carotid artery (the mean arterial pressure) is used as the reference for ICP. Using ICP recording protocols, many key parameters of erectile function can be measured from the ICP response curve. The ICP measurement provides more information than the apomorphine-induced penile erection test, and is cheaper than telemetric monitoring of the corpus spongiosum penis, making this method the most popular one to evaluate erectile function. However, compared to the easily-performed APO-induced erectile function test, successful ICP recordings require attention to detail, practice, and adherence to the operation method. In this work, an introduction to ICP recording in rats is provided to complement the procedure efficiently.

Intracavernosal pressure recording (ICP) is an important method to evaluate the erectile function of experimental animals. Here, a detailed protocol is demonstrated for the recording procedure of ICP by catheterizing the crura penis and then electrically stimulating the cavernous nerves in rats.

齧歯動物で勃起機能評価 Intracavernosal 圧記録

Erectile dysfunction (ED) is defined as the inability to attain or keep an erection of the penis, and this has become a prevalent male sexual disorder. ...

Inhibition of Wound Epidermis Formation via Full Skin Flap Surgery During Axolotl Limb Regeneration

Classical experiments in salamander regenerative biology over the last century have long established that the wound epidermis is a crucial signaling structure that forms rapidly post-amputation and is required for limb regeneration. However, methods to study its precise function at the molecular level over the last decades have been limited due to a paucity of precise functional techniques and genomic information available in salamander model systems. Excitingly, the recent plethora of sequencing technologies coupled with the release of various salamander genomes and the advent of functional genetic testing methods, including CRISPR, makes it possible to re-visit these foundational experiments at unprecedented molecular resolution. Here, I describe how to perform the classically developed full skin flap (FSF) surgery in adult axolotls in order to inhibit wound epidermis formation immediately following amputation. The wound epidermis normally forms via distal migration of epithelial cells in the skin proximal to the amputation plane to seal off the wound from the outside environment. The surgery entails immediately suturing full thickness skin (which includes both epidermal and dermal layers) over the amputation plane to hinder epithelial cell migration and contact with the underlying damaged mesenchymal tissues. Successful surgeries result in the inhibition of blastema formation and limb regeneration. By combining this surgery method with contemporary downstream molecular and functional analyses, researchers can begin to uncover the molecular underpinnings of wound epidermis function and biology during limb regeneration.

This article describes how to perform a surgical method to inhibit wound epidermis formation during axolotl limb regeneration by immediately suturing full thickness skin over the amputation plane. This method allows researchers to investigate the functional roles of the wound epidermis during the early stages of limb regeneration.

アキソロトル四肢再生中のフルスキンフラップ手術による創傷表皮形成の阻害

Classical experiments in salamander regenerative biology over the last century have long established that the wound epidermis is a crucial signaling ...

Application of Passive Head Motion to Generate Defined Accelerations at the Heads of Rodents

Exercise is widely recognized as effective for various diseases and physical disorders, including those related to brain dysfunction. However, molecular mechanisms behind the beneficial effects of exercise are poorly understood. Many physical workouts, particularly those classified as aerobic exercises such as jogging and walking, produce impulsive forces at the time of foot contact with the ground. Therefore, it was speculated that mechanical impact might be implicated in how exercise contributes to organismal homeostasis. For testing this hypothesis on the brain, a custom-designed ''passive head motion'' (hereafter referred to as PHM) system was developed that can generate vertical accelerations with controlled and defined magnitudes and modes and reproduce mechanical stimulation that might be applied to the heads of rodents during treadmill running at moderate velocities, a typical intervention to test the effects of exercise in animals. By using this system, it was demonstrated that PHM recapitulates the serotonin (5-hydroxytryptamine, hereafter referred to as 5-HT) receptor subtype 2A (5-HT2A) signaling in the prefrontal cortex (PFC) neurons of mice. This work provides detailed protocols for applying PHM and measuring its resultant mechanical accelerations at rodents' heads.

The present protocol describes a custom-designed ''passive head motion'' system, which reproduces mechanical accelerations at rodents' heads generated during their treadmill running at moderate velocities. It allows dissecting mechanical factors/elements from the beneficial effects of physical exercise.

げっ歯類の頭部に定義された加速度を生成するための受動的頭部運動の適用

Exercise is widely recognized as effective for various diseases and physical disorders, including those related to brain dysfunction. However, molecular ...

Performing In Vivo and Ex Vivo Electrical Impedance Myography in Rodents

Electrical impedance myography (EIM) is a convenient technique that can be used in preclinical and clinical studies to assess muscle tissue health and disease. EIM is obtained by applying a low-intensity, directionally focused, electrical current to a muscle of interest across a range of frequencies (i.e., from 1 kHz to 10 MHz) and recording the resulting voltages. From these, several standard impedance components, including the reactance, resistance, and phase, are obtained. When performing ex vivo measurements on excised muscle, the inherent passive electrical properties of the tissue, namely the conductivity and relative permittivity, can also be calculated. EIM has been used extensively in animals and humans to diagnose and track muscle alterations in a variety of diseases, in relation to simple disuse atrophy, or as a measure of therapeutic intervention. Clinically, EIM offers the potential to track disease progression over time and to assess the impact of therapeutic interventions, thus offering the opportunity to shorten the clinical trial duration and reduce sample size requirements. Because it can be performed noninvasively or minimally invasively in living animal models as well as humans, EIM offers the potential to serve as a novel translational tool enabling both preclinical and clinical development. This article provides step-by-step instructions on how to perform in vivo and ex vivo EIM measurements in mice and rats, including approaches to adapt the techniques to specific conditions, such as for use in pups or obese animals.