マークマッコーリーは、NIHメンター訓練助成金5T32HL066991 - 07でサポートされていますザンダーWehrensは1R01HL091947 - 01A209と3R01HL089598 - 03S109 NIH / NHLBIの助成金によってサポートされています

1。外科的移植のためのテレメーターの調製<ol><li>マウスに心電図テレメータを挿入する前に、それは、心電図テレメータは無菌と良好な状態にあることを確認することが重要です。新しいテレメトリ装置は、通常、製造元によって無菌状態で提供されています。心電図テレメーターは、デバイスが少なくとも4時間Tergazyme 1％溶液を用いて洗浄されていれ再利用することができます。あなたがTergazymeで洗浄した後、滅菌水でテレメータを洗うことができる。さらに、心電図テレメータを殺菌するために一晩Wavicide - 01消毒剤を使用。 48時間のマウスに移植する前に、滅菌水で消毒剤を洗い落とすようにしてください。滅菌容器に保管する。 </li><li>テレメータをチェックし、両方の導線の整合性と絶縁シースにつながります。磁石テレメータの5cm以内に手を振ってを使用してテレメトリ送信機の電源を入れ、AMラジオ、周波数530との信号をテストします。信号は強いと明確にする必要がありますし、強度とワイヤの操作に基づいてピッチに変化するはずです。モデル番号、テレメータシリアル番号、および心電図の較正値を記録します。メーカーはしばしば、テレメーターは、移植前に24時間の上にあることをお勧めします、詳細については、特定のテレメータの取扱説明書を参照してください。この例では、我々は、移植のためのデータサイエンスインターナショナル（DSI）テレメータを使用します。 </li><li>テレメータリードの準備は約3.5 cmおよび2.5センチメートルに正（赤）リード線にマイナス（白）のリード線を切断することによって達成することができる。これらは、右胸の負極リードの注入、及び左腹部のプラス側のリード線（図2参照）のための最適な長さです。次に、ワイヤーの7 mmを公開するために絶縁被覆を除去。そのような獣医-ボンドのような無菌瞬間接着剤、金属製のリードの端をペイントして、金属の先端に鉛ダストキャップを取り付け。これらのキャップは、配置を導くために起因する皮膚の浸食を避けることができます。約2〜3mmの金属線は、ネイティブの心臓のリズムの電気的検出用に公開される必要があります。 </li></ol> 2。テレメータの外科的移植<ol><li>マウス手術で無菌操作を使用するためには、滅菌手袋、滅菌ドレープ、および6から0 Prolene縫合が必要になります。また、滅菌ピンセット、先の尖っていないハサミの1ペア、メス、および針のドライバの2ペアが必要になります。手術器具は、摂氏250度に加熱ガラスビーズ滅菌器で滅菌することができる。以前、使用前に説明したようにテレメータを殺菌し、洗浄してください。 </li><li>最初の0.5 L /分100％O2でイソフルラン3％使用してインダクションボックスにマウスを麻酔による手術のためのマウスを準備します。麻酔マウスの温度を維持するために、そのような暖かい水を循環させるT -ポンプとして、加熱パッドを使用してください。必要に応じて温度は、直腸プローブによって監視されることがあります。マウスが十分に鎮静されている場合、完全に電気バリカンでマウスの腹部と胸部を削る。頭が離れて直面しているとあなたに向かって尾で、仰臥位、一定のフローチューブにマウスを転送する前に、イソフルランでマウスを再度麻酔。加熱された手術台にマウスの足ダウンテープ。剃毛毛を削除すると術野をきれいにアルコール綿で腹部を拭きます。腹部と胸部を消毒するために綿棒でbetadineの3つの別々のコートを適用します。マウスの爪床に圧力を適用することによって、麻酔の正しいレベルを確認してください。マウスが手術のために準備された後、滅菌手袋を使用し、手術野に滅菌ドレープを適用する。 </li><li>腹腔内テレメータの注入は、運動生理学の実験の利点を提供しています。腹腔内挿入のために、長さの腹部約2.5センチメートルを覆う皮膚に垂直正中切開を作成するためにメスを使用して手術を始めます。先の尖っていないハサミを使用して、基になる結合組織から慎重に別の皮膚。その後、、長さが約1.5 cmの腹膜を覆う白線の垂直正中切開を作成します。また、皮下のリードのための出口となる腹膜切開、に（脳神経）のすぐ上腹膜に小さな穴を作成します。手術を行っている間、時折動作フィールドに滅菌生理食塩水を滴下して手術部位の潤いを保つ。 </li><li>右腹腔内にテレメーターを挿入します。テレメータに優れた穴に鉗子を挿入し、それらが腹膜から突出しているように、リード穴に通し、両方のリード線をプルアップ。 1.5センチメートル腹膜垂直正中切開を閉じるために、連続6から0 Prolene縫合糸を使用してください。 </li></ol> 3。注入と腹部閉鎖をリード<ol><li> ECGのリードは、リードIIの構成で配置されます。白/透明シースと鉛は負であり、左上腹部に置かれます。最初に、マウスの右胸に0.5 cmの皮膚切開を作成します。次に、腹部切開に戻ってトンネルを作成する鈍はさみを使用してください。トンネルを経由してリードを引いて、胸筋にリードを固定するために6から0 Proleneの縫合糸を使用してください。縫合糸は、リードの露出部分の上にある、鉛及び基本的な筋肉の間の良好な接触を作成することを確認してください。筋肉にリードを固定するために、前述のものに近位第二縫合糸を使用してください。 6から0 Proleneの縫合糸を使用して皮膚切開を閉じます。 </li><li>プラス側のリード線（赤シース）は左横隔膜下と心臓の下の左腹部に配置されます。リードは6から0 Proleneの縫合によって下にある腹膜組織に固定されており、腹膜組織との良好な接触を持つ必要があります。鉛注入のサイトはこのように簡単に既存の手術野でアクセスすることが、垂直正中切開に近いため、追加の切開は、このステップでは必要ありません。腹膜組織の基盤となる腸の穿孔を避けるために、解除する必要があることに注意してください。 </li><li>層の6から0 Proleneで順次腹部筋膜と皮膚を閉じます。 </li></ol> 4。術後ケア<ol><li> [1 mg / mLの]手術直後鎮痛のためのブプレノルフィンのマウス0.1mLを与える。マウスは、加熱されたパッドの手術から回復することができます。マウスは長時間手術&gt; 30分、またはマウスが乾いた表示される場合は、水分補給のための腹膜に0.2から0.3 mLの滅菌生理食塩水を注入する可能性を持っている場合。継続的な術後の鎮痛のためには、3日間1日2回ブプレノルフィンとマウスを提供する標準的な手法であり、その後8時間ごとに必要に応じて。 </li><li>手術後の炎症を避けるために、アルコール綿棒でbetadineを拭き取ります。手術後の健康のその後の決定のための外科手術後の体重を記録してください。 </li><li>手術後8〜24時間、必要な時にブプレノルフィンとマウスの追加鎮痛を与える、これは痛みを軽減し、マウスのclawing後の手術部位でのリードの配置の失敗を削減するとして。 </li></ol> 5。代表的な結果皮下の皮膚の下にリードを正しく実行するとき、マウスは腹部切開を閉じているはずです。通常、マウスは麻酔から回復するために10〜30分かかります。マウスは完全に手術から回復している前にそれは、モビリティの術後の体重減少と正規化の回復によって証明される、約7〜10日かかる。 手術から最も一般的な合併症は、手術後の最初の10日間の皮膚からの手足のリード線の浸食です。この一般的な合併症は、しっかりと皮下に皮下組織や腹膜につながる付着することで回避することができます。この手法は、マウスがambulates時に皮膚への圧力と摩擦を作成スラックリード線を、避けることができます。あまり一般的で、感染症や敗血症は無菌操作でエラーが発生し、または使用するテレメータの不完全な洗浄から発生する可能性がありますが。 心電図の出力は、データサイエンスインターナショナル（DSI）のようなデータ収集ソフトウェアと結合受信機の行列、記録することができる。波形の結果は、明確に定義されたP波、心房脱分極を示す、また、下の図3に示すように、心室脱分極を、意味するQRS波を含める必要があります。良質ECGトレースが1:1の割合で明確に定義されたQRS波の前に明確に定義されたP波を持つ必要があります。低いバックグラウンドシグナルとRR間隔が野生型マウスでは定期的にあるべき存在するはずです。質の悪いECGトレースは、その後のコンピュータ解析を困難にすることが不明瞭なPやQRS波、またはバックグラウンド信号を、可能性があります。貧しいECGトレースは通常、再注入するテレメータのリード、一度で固定してもよい。テレメータリードを再移植する、麻酔下でマウスを置いて、手術手技を使用して、直接問題の鉛を覆う皮膚に切開を行います。リード線がアクセスされると、リードをアンカー縫合糸を切断し、再びリードを固定する前に目的の場所にリードを移動する。最後に、6から0 Proleneで手術部位を閉じてください。 <img src="/files/ftp_upload/1739/1739fig1.jpg" alt="図1" /> 図1：正と負のリード線と小動物のテレメーターの例。一般的には、赤いリード線は、正極リードであり、そして白のリード線は負のリードです。各リード線はむき出しになったワイヤの絶縁被覆、5-7 mmとし、皮膚を通して、鉛の侵食を防止するプラスチックキャップの先端を、持つ必要があります。 <img src="/files/ftp_upload/1739/1739fig2.jpg" alt="図2" /> 図2：マウスで適切なテレメータ注入部位を示す漫画。白（負の）リードは、マウスの右胸の上部に注入される、と赤（正の）リードは、テレメータの感覚その左腹部、&quot;鉛の二つの&quot;構成で配置されます。 &lt;Pクラス=&quot;jove_content&quot;&gt; <img src="/files/ftp_upload/1739/1739fig3.jpg" alt="図3" /> 図3：正常洞調律のマウスの代表波形。 P波は心房脱分極を表し、QRS波は心室の脱分極を表しています。高品質の波形は、1:1の比率でそれぞれのQRS波の前に明確なP波を持つ必要があります。

<ol>
	<li>Cerrone, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Bidirectional+ventricular+tachycardia+and+fibrillation+elicited+in+a+knock-in+mouse+model+carrier+of+a+mutation+in+the+cardiac+ryanodine+receptor.">Bidirectional ventricular tachycardia and fibrillation elicited in a knock-in mouse model carrier of a mutation in the cardiac ryanodine receptor.</a> Circ Res. 96 (10), e77-e77 (2005).</li><li>Chelu, M. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Calmodulin+kinase+II-mediated+sarcoplasmic+reticulum+Ca2++leak+promotes+atrial+fibrillation+in+mice.">Calmodulin kinase II-mediated sarcoplasmic reticulum Ca2+ leak promotes atrial fibrillation in mice.</a> J Clin Invest. 119 (7), 1940-1940 (1940).</li><li>Sood, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intracellular+calcium+leak+due+to+FKBP12.6+deficiency+in+mice+facilitates+the+inducibility+of+atrial+fibrillation.">Intracellular calcium leak due to FKBP12.6 deficiency in mice facilitates the inducibility of atrial fibrillation.</a> Heart Rhythm. 5 (7), 1047-1047 (2008).</li><li>Swoap, S. J., Gutilla, M. J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiovascular+changes+during+daily+torpor+in+the+laboratory+mouse.">Cardiovascular changes during daily torpor in the laboratory mouse.</a> Am J Physiol Regul Integr Comp Physiol. 297 (3), R769-R769 (2009).</li><li>Costa-Goncalves, d. a., C, A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+the+multidomain+protein+spinophilin+in+blood+pressure+and+cardiac+function+regulation.">Role of the multidomain protein spinophilin in blood pressure and cardiac function regulation.</a> Hypertension. 52 (4), 702-702 (2008).</li><li>Wehrens, X. H., Kirchhoff, S., Doevendans, P. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mouse+electrocardiography:+an+interval+of+thirty+years.">Mouse electrocardiography: an interval of thirty years.</a> Cardiovasc Res. 45 (1), 231-231 (2000).</li></ol>

この手順の重要なステップは、生存の手術前にマウスの適切な洗浄と準備です。テレメータとマウス手術部位が適切に洗浄されている場合、より良い手術後の転帰があります。さらに、配置を導くために細心の注意は、外科手術後の皮膚の侵食を軽減。技術への可能な変更は、ECG波形解析のためだけでなく、温度、さらには血圧の解析のためだけでなく、横方向のLV壁の機能を（私の構成を?...

<!DOCTYPE html PUBLIC "-//W3C//DTD XHTML 1.0 Transitional//EN" "http://www.w3.org/TR/xhtml1/DTD/xhtml1-transitional.dtd">
<html xmlns="http://www.w3.org/1999/xhtml" xml:lang="en" lang="en">	
		
		<div>
		<table border="1">
		<thead>
		<tr>
		<th>Material Name</th>
		<th>Type</th>
		<th>Company</th>
		<th>Catalogue Number</th>
		<th>Comment</th>
		</tr>
		</thead>
		<tbody>
		
<tr>
<td>Steri 250 Bead Sterilizer</td>
<td>&nbsp;</td>
<td>Inotech</td>
<td>IS-250</td>
<td>&nbsp;</td>
<tr>
<td>Blunt-Ended Scissors</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5980</td>
<td>&nbsp;</td>
<tr>
<td>Blunt-Ended Forceps (x2)</td>
<td>&nbsp;</td>
<td>Roboz</td>
<td>RS-5138</td>
<td>&nbsp;</td>
<tr>
<td>Veterinary Anesthesia Vaporizer</td>
<td>&nbsp;</td>
<td>Surgivet</td>
<td>Isotec 4</td>
<td>&nbsp;</td>
<tr>
<td>T/Pump</td>
<td>&nbsp;</td>
<td>Gaymar</td>
<td>P/N11184-000</td>
<td>&nbsp;</td>
<tr>
<td>Propylene Blue Monofilament Suture</td>
<td>&nbsp;</td>
<td>Ethicon</td>
<td>8718</td>
<td>&nbsp;</td>
<tr>
<td>Povodone Iodine, 10%</td>
<td>&nbsp;</td>
<td>Equate</td>
<td>N/A</td>
<td>&nbsp;</td>
<tr>
<td>Alcohol Swabs</td>
<td>&nbsp;</td>
<td>PDI</td>
<td>B339</td>
<td>&nbsp;</td>
<tr>
<td>Tergazyme</td>
<td>&nbsp;</td>
<td>Alconox</td>
<td>21837-118</td>
<td>&nbsp;</td>
<tr>
<td>Wavidicde-01</td>
<td>&nbsp;</td>
<td>Medical Chemical Corporation</td>
<td>Wavicide-01</td>
<td>&nbsp;</td>
<tr>
<td>ECG Telemeter</td>
<td>&nbsp;</td>
<td>DataSciences International</td>
<td>EA-F20</td>
<td>&nbsp;</td>		</tbody>
		</table>
		</div>

ambulatory ecg recording in mice

マウスの遠隔計測心電図記録は、不整脈、伝導障害、そして心臓突然死の背後にあるメカニズムを理解するために不可欠です。表面心電図は、波形の間隔の短期的な測定のために利用されていますが、それは、心拍変動や不整脈の珍しいエピソードのキャプチャの長期的研究のために実用的ではありません。埋め込み型心電図テレメーターは、単純な外科的移植、外来マウスのelectrogramsの長期的な記録、および複数の動物の同時録画とスケーラビリティの利点があります。ここで、我々は、マウスのホルター心電図記録用テレメータの注入へのステップバイステップガイドを紹介。無菌操作に細心の注意を順守は数週間から数ヶ月に注入し、記録の可能性と良好な生存率の結果が必要です。このように、埋め込み型心電図テレメトリは、マウスなどの外来動物モデルにおける心臓電気生理学に関する重要な情報を検出するための貴重なツールです。

Watch this Scientific Journal Video about Ambulatory ECG Recording in Mice at JoVE.com

Ambulatory ECG Recording in Mice

遠隔測定心電図は心​​臓の不整脈や心臓突然死のために動物モデルを評価する上で不可欠なツールとして浮上している。ここで、我々は、マウスの長期的、ホルター心電図モニタリングのアプリケーションのための測定ECG記録への段階的なガイドを提示する。

マウスにおけるホルター心電図の記録

Telemetric ECG recording in mice is essential to understanding the mechanisms behind arrhythmias, conduction disorders, and sudden cardiac death. Although ...

Research

JoVE Journal

Biology

23.5K ビュー.  Baylor College of Medicine (BCM). 遠隔測定心電図は心​​臓の不整脈や心臓突然死のために動物モデルを評価する上で不可欠なツールとして浮上している。ここで、我々は、マウスの長期的、ホルター心電図モニタリングのアプリケーションのための測定ECG記録への段階的なガイドを提示する。

ビデオ: Ambulatory ECG Recording in Mice

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

Patch Clamp Recording of Ion Channels Expressed in  Xenopus Oocytes

Since its development by Sakmann and Neher 1, 2, the patch clamp has become established as an extremely useful technique for electrophysiological measurement of single or multiple ion channels in cells. This technique can be applied to ion channels in both their native environment and expressed in heterologous cells, such as oocytes harvested from the African clawed frog, Xenopus laevis. Here, we describe the well-established technique of patch clamp recording from Xenopus oocytes. This technique is used to measure the properties of expressed ion channels either in populations (macropatch) or individually (single-channel recording). We focus on techniques to maximize the quality of oocyte preparation and seal generation. With all factors optimized, this technique gives a probability of successful seal generation over 90 percent. The process may be optimized differently by every researcher based on the factors he or she finds most important, and we present the approach that have lead to the greatest success in our hands.

This is intended as an introduction to patch clamp recording from Xenopus laevis oocytes. It covers vitelline membrane removal, formation of a gigaohm seal (gigaseal), and the optional conversion of the patch to the outside-out topology.

アフリカツメガエルの卵母細胞で発現イオンチャネルのパッチクランプ記録

Since its development by Sakmann and Neher 1, 2, the patch clamp has become established as an extremely useful technique for electrophysiological ...

Micro-drive Array for Chronic in vivo Recording: Drive Fabrication

Chronic recording of large populations of neurons is a valuable technique for studying the function of neuronal circuits in awake behaving rats. Lightweight recording devices carrying a high density array of tetrodes allow for the simultaneous monitoring of the activity of tens to hundreds of individual neurons. Here we describe a protocol for the fabrication of a micro-drive array with twenty one independently movable micro-drives. This device has been used successfully to record from hippocampal and cortical neurons in our lab. We show how to prepare a custom designed, 3-D printed plastic base that will hold the micro-drives. We demonstrate how to construct the individual micro-drives and how to assemble the complete micro-drive array. Further preparation of the drive array for surgical implantation, such as the fabrication of tetrodes, loading of tetrodes into the drive array and gold-plating, is covered in a subsequent video article.

Micro-drive Array for Chronic in vivo Recording: Drive Fabrication

In this protocol we demonstrate how to fabricate a micro-drive array for chronic electrophysiological recordings in rats.

慢性のためのマイクロドライブアレイ in vivoで録音：ドラ​​イブの作製

Chronic recording of large populations of neurons is a valuable technique for studying the function of neuronal circuits in awake behaving rats. ...

Micro-drive Array for Chronic in vivo Recording: Tetrode Assembly

The tetrode, a bundle of four electrodes, has proven to be a valuable tool for the simultaneous recording of multiple neurons in-vivo.  The differential amplitude of action potential signatures over the channels of a tetrode allows for the isolation of single-unit activity from multi-unit signals.  The ability to precisely control the stereotaxic location and depth of the tetrode is critical for studying coordinated neural activity across brain regions.  In combination with a micro-drive array, it is possible to achieve precise placement and stable control of many tetrodes over the course of days to weeks.  In this protocol, we demonstrate how to fabricate and condition tetrodes using basic tools and materials, install the tetrodes into a multi-drive tetrode array for chronic in-vivo recording in the rat, make ground wire connections to the micro-drive array, and attach a protective cone onto the micro-drive array in order to protect the tetrodes from physical contact with the environment.



Micro-drive Array for Chronic in vivo Recording: Tetrode Assembly

In this protocol we demonstrate how to fabricate and condition tetrodes for use with a micro-drive array, which was designed for chronic electrophysiological recordings in rats. In addition, we illustrate the final stages of micro-drive array construction, which includes installing ground wires and a protective cone.

慢性のためのマイクロドライブアレイ in vivoで録音：四極管アセンブリ

The tetrode, a bundle of four electrodes, has proven to be a valuable tool for the simultaneous recording of multiple neurons in-vivo.  The differential ...

Measuring Blood Pressure in Mice using Volume Pressure Recording, a Tail-cuff Method

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. The CODA tail-cuff system uses Volume Pressure Recording (VPR) to measure the blood pressure by determining the tail blood volume. A specially designed differential pressure transducer and an occlusion tail-cuff measure the total blood volume in the tail without the need to obtain the individual pulse signal. Special attention is afforded to the length of the occlusion cuff in order to derive the most accurate blood pressure readings. VPR can easily obtain readings on dark-skinned rodents, such as C57BL6 mice and is MRI compatible. The CODA system provides you with measurements of six (6) different blood pressure parameters; systolic and diastolic blood pressure, heart rate, mean blood pressure, tail blood flow, and tail blood volume. Measurements can be made on either awake or anesthetized mice or rats. The CODA system includes a controller, laptop computer, software, cuffs, animal holders, infrared warming pads, and an infrared thermometer. There are seven different holder sizes for mice as small as 8 grams to rats as large as 900 grams.

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. This tail-cuff system uses Volume Pressure Recording (VPR) to measure the blood pressure by determining the tail blood volume.

ボリューム圧力記録、テールカフ法を用いたマウスにおける測定血圧

The CODA 8-Channel High Throughput Non-Invasive Blood Pressure system measures the blood pressure in up to 8 mice or rats simultaneously. The CODA ...

Electrophysiological Recording in the Drosophila Embryo

Drosophila is a premier genetic model for the study of both embryonic development and functional neuroscience. Traditionally, these fields are quite isolated from each other, with largely independent histories and scientific communities. However, the interface between these usually disparate fields is the developmental programs underlying acquisition of functional electrical signaling properties and differentiation of functional chemical synapses during the final phases of neural circuit formation. This interface is a critically important area for investigation. In Drosophila, these phases of functional development occur during a period of &lt;8 hours (at 25&deg;C) during the last third of embryogenesis. This late developmental period was long considered intractable to investigation owing to the deposition of a tough, impermeable epidermal cuticle. A breakthrough advance was the application of water-polymerizing surgical glue that can be locally applied to the cuticle to enable controlled dissection of late-stage embryos. With a dorsal longitudinal incision, the embryo can be laid flat, exposing the ventral nerve cord and body wall musculature to experimental investigation. Whole-cell patch-clamp techniques can then be employed to record from individually-identifiable neurons and somatic muscles. These recording configurations have been used to track the appearance and maturation of ionic currents and action potential propagation in both neurons and muscles. Genetic mutants affecting these electrical properties have been characterized to reveal the molecular composition of ion channels and associated signaling complexes, and to begin exploration of the molecular mechanisms of functional differentiation. A particular focus has been the assembly of synaptic connections, both in the central nervous system and periphery. The glutamatergic neuromuscular junction (NMJ) is most accessible to a combination of optical imaging and electrophysiological recording. A glass suction electrode is used to stimulate the peripheral nerve, with excitatory junction current (EJC) recordings made in the voltage-clamped muscle. This recording configuration has been used to chart the functional differentiation of the synapse, and track the appearance and maturation of presynaptic glutamate release properties. In addition, postsynaptic properties can be assayed independently via iontophoretic or pressure application of glutamate directly to the muscle surface, to measure the appearance and maturation of the glutamate receptor fields. Thus, both pre- and postsynaptic elements can be monitored separately or in combination during embryonic synaptogenesis. This system has been heavily used to isolate and characterize genetic mutants that impair embryonic synapse formation, and thus reveal the molecular mechanisms governing the specification and differentiation of synapse connections and functional synaptic signaling properties.

Electrophysiological Recording in the Drosophila Embryo

Electrophysiological recordings from Drosophila embryos allow analyses of developing muscle and neuron electrical properties, as well as characterization of functional synaptogenesis at the glutamatergic neuromuscular junction and central cholinergic and GABAergic synapses.

の電気生理学的記録ショウジョウバエ胚

Drosophila is a premier genetic model for the study of both embryonic development and functional neuroscience. Traditionally, these fields are quite ...

Transverse Aortic Constriction in Mice

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model for pressure overload-induced cardiac hypertrophy and heart failure.1 TAC initially leads to compensated hypertrophy of the heart, which often is associated with a temporary enhancement of cardiac contractility. Over time, however, the response to the chronic hemodynamic overload becomes maladaptive, resulting in cardiac dilatation and heart failure.2 The murine TAC model was first validated by Rockman et al.1, and has since been extensively used as a valuable tool to mimic human cardiovascular diseases and elucidate fundamental signaling processes involved in the cardiac hypertrophic response and heart failure development. When compared to other experimental models of heart failure, such as complete occlusion of the left anterior descending (LAD) coronary artery, TAC provides a more reproducible model of cardiac hypertrophy and a more gradual time course in the development of heart failure. Here, we describe a step-by-step procedure to perform surgical TAC in mice. To determine the level of pressure overload produced by the aortic ligation, a high frequency Doppler probe is used to measure the ratio between blood flow velocities in the right and left carotid arteries.3, 4 With surgical survival rates of 80-90%, transverse aortic banding is an effective technique of inducing left ventricular hypertrophy and heart failure in mice.

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model to study mechanisms underlying cardiac hypertrophy and heart failure development. Here, we describe procedures to constrict the aorta to create a reproducible degree of cardiac hypertrophy in mice.

マウスの横大動脈狭窄

Transverse aortic constriction (TAC) in the mouse is a commonly used experimental model for pressure overload-induced cardiac hypertrophy and heart ...

Recording Multicellular Behavior in Myxococcus xanthus Biofilms using Time-lapse Microcinematography

A swarm of the &delta;-proteobacterium Myxococcus xanthus contains millions of cells that act as a collective, coordinating movement through a series of signals to create complex, dynamic patterns as a response to environmental cues. These patterns are self-organizing and emergent; they cannot be predicted by observing the behavior of the individual cells. Using a time-lapse microcinematography tracking assay, we identified a distinct emergent pattern in M. xanthus called chemotaxis, defined as the directed movement of a swarm up a nutrient gradient toward its source 1.
In order to efficiently characterize chemotaxis via time-lapse microcinematography, we developed a highly modifiable plate complex (Figure 1) and constructed a cluster of 8 microscopes (Figure 2), each capable of capturing time-lapse videos. The assay is rigorous enough to allow consistent replication of quantifiable data, and the resulting videos allow us to observe and track subtle changes in swarm behavior. Once captured, the videos are transferred to an analysis/storage computer with enough memory to process and store thousands of videos. The flexibility of this setup has proven useful to several members of the M. xanthus community.

Recording Multicellular Behavior in Myxococcus xanthus Biofilms using Time-lapse Microcinematography

To study Myxococcus xanthus swarm behavior, we have designed a time-lapse microcinematography protocol that can be modified for different assays. It employs standard growth conditions adapted for microscopy, and yields reproducible results by the use of inexpensive, reusable silicone gaskets. We have used this method to quantify multicellular chemotaxis.

で多細胞の動作を記録する Myxococcusザンザス</emタイムラプスMicrocinematographyを使用して>バイオフィルム

A swarm of the &delta;-proteobacterium Myxococcus xanthus contains millions of cells that act as a collective, coordinating movement through a series of ...

One-channel Cell-attached Patch-clamp Recording

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate depends on properties intrinsic to each channel protein as well as the mechanism by which it is generated and controlled and represents an important area of current research. Information about the operational dynamics of ion channel proteins can be obtained by observing long stretches of current produced by a single molecule. Described here is a protocol for obtaining one-channel cell-attached patch-clamp current recordings for a ligand gated ion channel, the NMDA receptor, expressed heterologously in HEK293 cells or natively in cortical neurons. Also provided are instructions on how to adapt the method to other ion channels of interest by presenting the example of the mechano-sensitive channel PIEZO1. This method can provide data regarding the channel&#8217;s conductance properties and the temporal sequence of open-closed conformations that make up the channel&#8217;s activation mechanism, thus helping to understand their functions in health and disease.

Described here is a procedure for obtaining long stretches of current recording from one ion channel with the cell-attached patch-clamp technique. This method allows for observing, in real time, the pattern of open-close channel conformations that underlie the biological signal. These data inform about channel properties in undisturbed biological membranes.

一チャネルセル付パッチクランプ法

Ion channel proteins are universal devices for fast communication across biological membranes. The temporal signature of the ionic flux they generate ...

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. ...