この作品は、ベルギー連邦政府（IUAP P6/28）、研究財団·フランダース（FWO）（G.0565.07とG.0686.09）、アステラス欧州財団賞2009とKU Leuvenの研究評議会からの補助金によって支えられて（GOA 2009/07、EF/95/010とPFV/10/006）。 PUと我々は、研究財団フランダース（FWO）のポスドクです。 MBは、マリー·キュリー·フェローでもある。 DDR FWOの基本的な臨床の仲間。

1。実験動物<ol><li>動物（マウス、ラット）が12時間明暗サイクルと水と標準食物ペレットへの不断のアクセス権を持つ専門の動物施設に収容されています。動物の年齢や性別はどちらもニーズに応じて標準化されるべき重要なパラメータである。 12週齢の雌の動物5,6 -我々は、典型的には10で膀胱内圧測定を行います。 </li><li>全ての動物実験は、欧州連合コミュニティ協議会のガイドラインに従って実施され、地域の倫理委員会によって承認されています。 </li></ol> 2。麻酔<ol><li>それはその短い半減期のために投与量に容易であるのでイソフルレン麻酔は、小さな外科的処置を行うために使用されます。イソフルランを使用するときに適切な清掃、廃ガスが必要です。 <ol class="lalpha"><li>動物は純粋な酸素でイソフルラン5％（1リットル/分）でガス、密閉ボックス内に配置されます。 </li><li>適切なlの誘導後、麻酔1と1リットル/分の酸素 - - イソフルランの1.5％を0.8 evelによって小さなマスク（カット10mlシリンジ）によって維持されています。 </li></ol></li><li>マウスの場合、ウレタン麻酔は機能録音（cfr.赤外線）中に使用されます。ほとんどの他の麻酔薬とは対照的に、排尿反射がこの麻酔剤4によって抑制されていない、ので、ウレタン麻酔は、機能録音時に使用されます。ウレタン1.2グラム/ kg体重は60分の録音が開始される前に皮下（sc）投与される。必要に応じて、適切な麻酔を得るために必要なウレタンの用量を皮下に追加投与（0.1 g / kg）を投与することにより、滴定することができる。それは注意すべきである、しかし、その追加の麻酔は排尿効率ひいては排尿頻度7,8を変更することがあります。したがって、cystometric録音が密接に監視および制御動作からの有意な偏差が観察されたときに破棄されるべきである。麻酔薬は、試験中に投与してはならない化合物は、データの誤解釈を避けるために適用されます。 </li></ol> 3。外科手術 - 膀胱カテーテル注入<ol><li>外科用器具は、使用前にオートクレーブで滅菌されています。 、動物の腹部を剃る70％エタノールで消毒し、膀胱を露出させるために低正中開腹を行う。 </li><li>手術用顕微鏡下では、非吸収性モノフィラメント縫合糸（サイズ6-0）を使用して膀胱に巾着縫合を置きます。 </li><li>巾着の中に小さな膀胱を（我々が優先的に18Gの針を使用）を実行し、この貫通孔の端の小さなカフ（慎重にチューブを加熱することによって得られる）と、PE 50ポリエチレンカテーテルを挿入します。我々の手で、より薄いチューブ（PE 10）は、より高く、より可変抵抗を持っています。チューブはエタノール70％溶液に入れ、オートクレーブで滅菌することによって消毒することができます。注入前に、チューブを生理食塩水でフラッシュする必要があります。 </li><li>財布strを確保外科医の結び目で管の周囲ING。釣鐘型の先端が右縫合下に配置されるまでゆっくりと外側にカテーテルを引き抜きます。漏れを防ぐために、縫合糸に向かって18Gのカテーテルの部分を押し込みます。 </li><li>漏れがないか確認するために静かに - 生理食塩水を少量（200μlの100）を吹き込む。漏れがある場合は、追加の縫合糸は、配置する必要があります。 </li><li>切開の上部にカテーテルの通路を残して、モノフィラメント縫合糸を用いて腹部の筋肉を閉じます。 </li><li>トンネル中空の金属棒を使って肩甲間部​​にチューブが、それによって実験中にチューブを噛まないように動物を防ぐ。 </li><li>皮膚は非吸収性モノフィラメント縫合糸を使用して閉じられます。移植されたカテーテルは、その透過性を確認するために、生理食塩水でフラッシュされます。 </li><li>動物が目覚める前に、鎮痛薬（ブプレノルフィン、0.05 mg / kg）を皮下投与されています。 </li></ol> 4。セットアップと膀胱内圧測定<ol><li>実験クエスティに応じて（そして、種が使用される）に膀胱内圧測定は、目を覚まして（拘束またはunrestraint）またはウレタン麻酔動物で行われます。膀胱内圧測定が拘束環境で冷静さを保つ傾向にある意識下ラットにおいて、完全に可能であるのに対し、マウス3,5,6,9を使用しているとき、我々は通常、ウレタン麻酔（1.2 mg / kgの皮下）の下で膀胱内圧測定を行います。 </li><li>麻酔したマウスでは、体温を常に監視し、36.5に保たれている℃±0.2℃の温度プローブと加熱ランプを用いた。 </li><li>セットアップは、製造元の指示に従って、すべての実験の前に校正されています。 </li><li>移植されたカテーテルは、一方の側の圧力変換器（エドワーズライフサイエンス）およびその他（ハーバード装置）上の輸液ポンプに接続されている3ウェイタップにルアースタブによって接続されています。圧力トランスデューサは、データ収集システム（DI-730-USB Dataq楽器）と共同にアンプ（78534cモニター、ヒューレットパッカード）を介して接続されているmputer。 Windaq / Liteソフトウェアは、記録のために使用することができます。 </li><li>輸液ポンプは、例えば、生理食塩水またはPBSの注入を可能にし、開始され、100μl/分（ラット）または20μl/分（マウス）。膀胱内圧が記録されている間などの膀胱は、一定の割合で満たされるでしょう。 </li><li> 図5は、標準的な圧力のトレースを表しています特定のボリューム/圧力に到達するまで、輸液中、遅い圧力上昇は、排尿閾値は、膀胱内にあります。このしきい値に達すると、膀胱が収縮し、その後尿道括約筋は尿道を通って尿の通過を可能にする、オープンします。このように、膀胱が空になると、収縮が停止し、圧力が &quot;基礎&quot;レベルに再び低下します。 </li><li>ラットを代謝ケージ内とデジタルのバランスの上に置かれたように、排尿量は排尿量についての情報を与え、同時に測定することができます。マイルの小さなボイドボリュームに起因するceが、それは、このシステムを使用して非常に困難な場合があります。したがって、排尿量は排尿の前後に計量し、小さなろ紙を用いて決定される。 </li><li>通常、我々は、システムが30分間の記録期間が続く30分間平衡化しましょう​​。その後、薬は全身的または膀胱内投与することができ、さらに30分が記録されている。 </li><li>記録されたデータは、定量分析に特化したソフトウェアにインポートすることができます。csvファイルとしてエクスポートすることができます。我々は一般的に（OriginLabのコーポレーション、米国マサチューセッツ州）原点を使用しています。 </li><li>実験の後、動物を頸椎脱臼（マウス）またはCO 2中毒（ラット）で屠殺する。 </li></ol> 5。代表的な結果<img alt="図1" src="/files/ftp_upload/3869/3869fig1.jpg" /> 図1：開腹概要。 A）は仰臥位でラットを置きます。 B）を剃ると殺菌して手術部位を準備します。皮膚のC）を切開。 D）を切開OF腹筋、膀胱暴露。 <img alt="図2" src="/files/ftp_upload/3869/3869fig2.jpg" /> 図2：巾着縫合。 <img alt="図3" src="/files/ftp_upload/3869/3869fig3.jpg" /> （3）カテーテル注入図 。 <img alt="図4" src="/files/ftp_upload/3869/3869fig4.jpg" /> 図4カテーテルのトンネリング。 覚醒ラット及び麻酔マウスに生理食塩水の膀胱内灌流の間に得 ​​られた圧力測定の例を図5に示す。複数のパラメータが圧力信号（intercontractile間隔、基準圧力と閾値圧力など ）から抽出することができます。これらのパラメータの包括的な説明については、。Anderssonらを参照してください（参考文献4）と吉山ら （<html> REF 10）。 我々は最近、マスタードオイル（MO）で、長い間、膀胱11,12などの内臓器官の炎症や痛覚過敏の実験モデルで使用されてきた非常に反応性の高い化合物の分子標的を同定するために膀胱内圧測定を使用している。 10mMのMOの膀胱内注入は、野生型マウスにおける排尿頻度の大幅な上昇（intercontractile間隔の減少）（ 図6A、B）及び排尿量6の減少を誘発した。興味深いことに、MOはMOのTRPA1受容体の欠損マウスでも同様の変更を誘導した。これとは対照的に、MOはWTマウスに比べてTRPV1 KOマウスにおいてcystometricパラメータのはるかに弱い変化を誘導し、Trpa1/Trpv1 KOマウスで任意の効果がなかった。一緒にペプチド（CGRP）6カルシトニン遺伝子関連のリリースの測定値と、これらのデータは、TRPV1は、MOにより誘発される内臓炎症に重要な役割を果たしている可能性があることを示していることを示します。 ve_content &quot;&gt; <img alt="図5" src="/files/ftp_upload/3869/3869fig5.jpg" /> 図5に配慮した雌ラット（）で、麻酔雌マウス（B）に記録された膀胱内圧の代表痕跡。最低圧力は &quot;ベースライン圧力&quot;（赤矢印）として定義されています。充填段階の終わりでの圧力は青い矢印でマークされています。ベースラインの圧力） &quot;intercontractile間隔&quot;（ICI）の間の時間です - 圧力差で割ったもの、これらの点の間に注入された流体の体積は、膀胱壁のコンプライアンスの計算（コンプライアンス=注入量/が（閾値圧力を可能2排尿収縮。 <img alt="図6" src="/files/ftp_upload/3869/3869fig6.jpg" /> 図6野生型における膀胱内圧測定パターンとTRPA1、TRPV1とTrpa1/Trpv1ノックアウトマウスにマスタードオイルの膀胱内のアプリケーションの影響。（A）は、生理食塩水および10mM MOの注入に応答して、WT、KO TRPA1、TRPV1 KOおよびTrpa1/Trpv1 KOマウスに記録されている膀胱内圧の変化の代表的な例。前とMOの膀胱内注入時の平均排尿瞬時周波数の（B）の時間経過。すべてのマウスでは、データは生理食塩水注入の間に得られた平均周波数に正規化された。これらのデータは、エルゼビアの許可を得て、Everaerts ら （文献6）から適合させられます。

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特別な利害関係は宣言されません。

ここに提示され膀胱内圧測定技術は、動物モデルにおける膀胱機能の in vivo測定で行うことができます。ラットは、おそらく最もよく使われる動物モデルである。マウスのほうが扱いは難しいですが、遺伝子操作された動物を使用することの利点を提供します。なぜなら埋め込まれたカテーテルと膀胱内圧に影響を与える可能性が腹腔内圧力の変化の緩み、その結果非常に活発?...

<table border="1"><tbody><tr><td> 試薬の名称 </td><td> 商品名、会社名、 </td><td> カタログ番号 </td><td> 注釈 </td></tr><tr><td>ウレタン</td><td>ウレタン、シグマアルドリッチ</td><td> 315419 </td><td>グループ2B発がん物質</td></tr><tr><td>イソフルレン</td><td> Isoba、シェリング·プラウアニマルヘルス</td><td></td><td></td></tr><tr><td>ポリエチレンカテーテル</td><td> IntramedicポリエチレンチューブPE50、ベクトン·ディッキンソン</td><td> 427411 </td><td></td></tr><tr><td>外科用顕微鏡</td><td> OP-MI 6、カールツァイス</td><td> OP-MI 6 </td><td></td></tr><tr><td>巾着縫合</td><td>プロレン6/0、エチコン</td><td> 8610H </td><td></td></tr><tr><td>筋膜と皮膚の縫合</td><td> Ethilon 4/0または5/0、エチコン</td><td> 662Gまたは661G </td><td></td></tr><tr><td>術後の鎮痛薬</td><td> Temgesic、シェリング·プラウアニマルヘルス</td><td></td><td>ラットの投与量：0.05 mg / kgで</td></tr><tr><td>アンプ</td><td> 78534cモニター、ヒューレットパッカード</td><td></td><td></td></tr><tr><td>分析天びんとバランスデータ収集ソフト</td><td> FZ 300iは、A＆D </td><td> FZ-300I </td><td></td></tr><tr><td>輸液ポンプ</td><td>ポンプ33、ハーバード装置</td><td> HA33 </td><td></td></tr><tr><td>膀胱内圧測定記録システム</td><td> Dataq楽器、DI-730シリーズとWindaq /ライト</td><td> DI-730-USB Windaq /ライト</td><td></td></tr><tr><td>温度の登録</td><td>フルーク52 KJ体温計</td><td> 52 KJ </td><td></td></tr><tr><td>圧力トランスデューサ</td><td>エドワーズライフサイエンス、圧力監視セット</td><td> T322247A </td><td></td></tr></tbody></table>

the use of cystometry in small rodents: a study of bladder chemosensation

<p class="jove_content"尿を格納するために、効率的に都合の良い時に、それを追放することができる動的な貯水池として&gt;下部尿路（LUT）で機能します。尿を保存しながら、しかし、膀胱が廃棄物に長時間さらされている。タイトな障壁として作用することにより、LUTの上皮、尿路上皮には、有害物質の再吸収を避けることができます。また、有害化学物質は、膀胱の神経支配侵害を刺激し、膀胱の内容を追放排尿収縮を開始する。興味深いことに、有害な化学物質への膀胱の感度は膀胱神経因性膀胱の過活動の治療にTRPV1アゴニストカプサイシンを注入することにより、臨床現場で正常に使用されています<sup&gt; 1</sup&gt;。これは、化学感覚器官として膀胱を見ることの利点を強調し、さらに臨床研究のためのプロンプトが表示されます。しかし、倫理的な問題は深刻なヒト被験者は、LUT臨床薬理学の分子基盤を解明するために必要な侵襲的な測定では、実行する可能性を制限する。この限界を克服する方法は、いくつかの動物モデルを使用することである<sup&gt; 2</sup&gt;。ここでは、マウスとラットの膀胱内圧測定、制御された膀胱灌流の条件で膀胱内圧を測定可能にする技術の実装について説明します。</p&gt;<p class="jove_content"&gt;開腹後、カテーテルを膀胱内に注入し、肩甲間部に皮下にトンネリングされます。尿道は排尿のための無料のままですその後膀胱は、制御された速度で充填することができる。充填と排尿の反復サイクルの間に、膀胱内圧が埋め込まれたカテーテルを介して測定することができます。このように、圧力の変化を定量化して分析することができます。また、排尿量の同時測定により、非排尿収縮を区別排尿収縮<sup&gt; 3</sup&gt;。</p&gt;<p class="jove_content"&gt;重要なのは、げっ歯類とヒトの間に排尿のコントロールの違いにより、これらの動物におけるcystometric測定は、限られた並進値を持っている<sup&gt; 4</sup&gt;。それにもかかわらず、彼らは実験的な前臨床設定で膀胱の病態生理および薬理学の研究において非常に尽力されています。この手法を用いた最近の研究では、膀胱のメカノと化学感覚特性における新規分子の選手の重要な役割を明らかにした。</p

Watch this Scientific Journal Video about The Use of Cystometry in Small Rodents: A Study of Bladder Chemosensation at JoVE.com

The Use of Cystometry in Small Rodents: A Study of Bladder Chemosensation

膀胱内圧測定では、小動物の膀胱機能を測定するための効率的な手法である<em生体内で&gt;</em&gt;。膀胱は、継続的に尿道が排尿のための無料の残っているのに対し、膀胱カテーテルを介して制御された速度で注入されています。これは、膀胱内圧と排尿量が記録されている間、繰り返しのために充填し、膀胱を空にすることができます。

小型げっ歯類における膀胱内圧測定の使用：膀胱Chemosensationの研究

	The lower urinary tract (LUT) functions as a  dynamic reservoir that is able to store urine and to efficiently expel it at a  convenient time. While ...

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Research

JoVE Journal

Medicine

22.3K ビュー.  KU Leuven, Belgium. 膀胱内圧測定では、小動物の膀胱機能を測定するための効率的な手法である。膀胱は、継続的に尿道が排尿のための無料の残っているのに対し、膀胱カテーテルを介して制御された速度で注入されています。これは、膀胱内圧と排尿量が記録されている間、繰り返しのために充填し、膀胱を空にすることができます。

ビデオ: The Use of Cystometry in Small Rodents: A Study of Bladder Chemosensation

Mouse Bladder Wall Injection

Mouse bladder wall injection is a useful technique to orthotopically study bladder phenomena, including stem cell, smooth muscle, and cancer biology. Before starting injections, the surgical area must be cleaned with soap and water and antiseptic solution. Surgical equipment must be sterilized before use and between each animal. Each mouse is placed under inhaled isoflurane anesthesia (2-5% for induction, 1-3% for maintenance) and its bladder exposed by making a midline abdominal incision with scissors. If the bladder is full, it is partially decompressed by gentle squeezing between two fingers. The cell suspension of interest is intramurally injected into the wall of the bladder dome using a 29 or 30 gauge needle and 1 cc or smaller syringe. The wound is then closed using wound clips and the mouse allowed to recover on a warming pad. Bladder wall injection is a delicate microsurgical technique that can be mastered with practice.

Mouse bladder wall injection is a useful approach to orthotopically study bladder stem cell and cancer biology. This delicate microsurgical method can be mastered with careful technique and practice.

マウスの膀胱壁インジェクション

Mouse bladder wall injection is a useful technique to orthotopically study bladder phenomena, including stem cell, smooth muscle, and cancer biology. ...

医学

Biomarkers in an Animal Model for Revealing Neural, Hematologic, and Behavioral Correlates of PTSD

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for objective diagnosis but also for the evaluation of therapeutic efficacy and resilience to trauma. Ongoing research is directed at identifying molecular biomarkers for PTSD, including traumatic stress induced proteins, transcriptomes, genomic variances and genetic modulators, using biologic samples from subjects' blood, saliva, urine, and postmortem brain tissues. However, the correlation of these biomarker molecules in peripheral or postmortem samples to altered brain functions associated with psychiatric symptoms in PTSD remains unresolved. Here, we present an animal model of PTSD in which both peripheral blood and central brain biomarkers, as well as behavioral phenotype, can be collected and measured, thus providing the needed correlation of the central biomarkers of PTSD, which are mechanistic and pathognomonic but cannot be collected from people, with the peripheral biomarkers and behavioral phenotypes, which can.
Our animal model of PTSD employs restraint and tail shocks repeated for three continuous days - the inescapable tail-shock model (ITS) in rats. This ITS model mimics the pathophysiology of PTSD 17, 7, 4, 10. We and others have verified that the ITS model induces behavioral and neurobiological alterations similar to those found in PTSD subjects 17, 7, 10, 9. Specifically, these stressed rats exhibit (1) a delayed and exaggerated startle response appearing several days after stressor cessation, which given the compressed time scale of the rat's life compared to a humans, corresponds to the one to three months delay of symptoms in PTSD patients (DSM-IV-TR PTSD Criterian D/E 13), (2) enhanced plasma corticosterone (CORT) for several days, indicating compromise of the hypothalamopituitary axis (HPA), and (3) retarded body weight gain after stressor cessation, indicating dysfunction of metabolic regulation.
The experimental paradigms employed for this model are: (1) a learned helplessness paradigm in the rat assayed by measurement of acoustic startle response (ASR) and a charting of body mass; (2) microdissection of the rat brain into regions and nuclei; (3) enzyme-linked immunosorbent assay (ELISA) for blood levels of CORT; (4) a gene expression microarray plus related bioinformatics tools 18. This microarray, dubbed rMNChip, focuses on mitochondrial and mitochondria-related nuclear genes in the rat so as to specifically address the neuronal bioenergetics hypothesized to be involved in PTSD.

We describe a rat model of post traumatic stress disorder (PTSD) that reveals the persistent alterations in neuroendocrine function and the delayed long-term, exaggerated fear response, characteristic of PTSD patients. The animal model and methods described here are useful for correlating biomarkers in brain nuclei, which are mechanistic but cannot be measured in patients, with biomarkers in peripheral white blood cells, which can.

PTSDの、ニューラル血液学的および行動的相関関係を明らかにするための動物モデルにおけるバイオマーカー

Identification of biomarkers representing the evolution of the pathophysiology of Post Traumatic Stress Disorder (PTSD) is vitally important, not only for ...

Protocol for Long Duration Whole Body Hyperthermia in Mice

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body temperature that is observed during fever. The use of hyperthermia as an adjuvant has emerged as a promising procedure for tumor regression in the field of cancer biology. For this purpose, the most important requirement is to have reliable and uniform heating protocols. We have developed a protocol for hyperthermia (whole body) in mice. In this protocol, animals are exposed to cycles of hyperthermia for 90 min followed by a rest period of 15 min. During this period mice have easy access to food and water. High body temperature spikes in the mice during first few hyperthermia exposure cycles are prevented by immobilizing the animal. Additionally, normal saline is administered in first few cycles to minimize the effects of dehydration. This protocol can simulate fever like conditions in mice up to 12-24 hr. We have used 8-12 weeks old BALB/Cj female mice to demonstrate the protocol.

This paper describes a protocol for whole body hyperthermia in mice that can stimulate fever like conditions up to 12-24 hr.

マウスの長い持続時間の全身温熱療法のためのプロトコル

Hyperthermia is a general term used to define the increase in core body temperature above normal. It is often used to describe the increased core body ...

Human Neuroendocrine Tumor Cell Lines as a Three-Dimensional Model for the Study of Human Neuroendocrine Tumor Therapy

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human malignancies.1 Other than surgery for the minority of patients who present with localized disease, there is little or no survival benefit of systemic therapy. Therefore, there is a great need to better understand the biology of NETs, and in particular define new therapeutic targets for patients with nonresectable or metastatic neuroendocrine tumors. 3D cell culture is becoming a popular method for drug screening due to its relevance in modeling the in vivo tumor tissue organization and microenvironment.2,3 The 3D multicellular spheroids could provide valuable information in a more timely and less expensive manner than directly proceeding from 2D cell culture experiments to animal (murine) models.
To facilitate the discovery of new therapeutics for NET patients, we have developed an in vitro 3D multicellular spheroids model using the human NET cell lines. The NET cells are plated in a non-adhesive agarose-coated 24-well plate and incubated under physiological conditions (5% CO2, 37 &deg;C) with a very slow agitation for 16-24 hr after plating. The cells form multicellular spheroids starting on the 3rd or 4th day. The spheroids become more spherical by the 6th day, at which point the drug treatments are initiated. The efficacy of the drug treatments on the NET spheroids is monitored based on the morphology, shape and size of the spheroids with a phase-contrast light microscope. The size of the spheroids is estimated automatically using a custom-developed MATLAB program based on an active contour algorithm. Further, we demonstrate a simple method to process the HistoGel embedding on these 3D spheroids, allowing the use of standard histological and immunohistochemical techniques. 
This is the first report on generating 3D spheroids using NET cell lines to examine the effect of therapeutic drugs. We have also performed histology on these 3D spheroids, and displayed an example of a single drug's effect on growth and proliferation of the NET spheroids. Our results support that the NET spheroids are valuable for further studies of NET biology and drug development.

We present a simple agarose overlay platform to grow 3D multicellular spheroids using neuroendocrine cancer cell lines. This method provides a very convenient way to examine the effect of therapeutic drugs on the neuroendocrine tumor cells. It could also help us establish human neuroendocrine tumor spheroids for cancer therapy.

人間の神経内分泌腫瘍療法の研究のための三次元モデルとしての人間の神経内分泌腫瘍細胞株

Neuroendocrine tumors (NETs) are rare tumors, with an incidence of two per 100,&#x200A;000 individuals per year, and they account for 0.5% of all human ...

Urinary Bladder Distention Evoked Visceromotor Responses as a Model for Bladder Pain in Mice

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition characterized by increased urgency and frequency of urination, as well as nocturia and general pelvic pain, especially upon bladder filling or voiding. Despite years of research, the cause of IC/BPS remains elusive and treatment strategies are unable to provide complete relief to patients. In order to study nervous system contributions to the condition, many animal models have been developed to mimic the pain and symptoms associated with IC/BPS. One such murine model is urinary bladder distension (UBD). In this model, compressed air of a specific pressure is delivered to the bladder of a lightly anesthetized animal over a set period of time. Throughout the procedure, wires in the superior oblique abdominal muscles record electrical activity from the muscle. This activity is known as the visceromotor response (VMR) and is a reliable and reproducible measure of nociception. Here, we describe the steps necessary to perform this technique in mice including surgical manipulations, physiological recording, and data analysis. With the use of this model, the coordination between primary sensory neurons, spinal cord secondary afferents, and higher central nervous system areas involved in bladder pain can be unraveled. This basic science knowledge can then be clinically translated to treat patients suffering from IC/BPS.

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition characterized in part by pelvic pain. In order to study nervous system contributions to the condition, a physiological model of urinary bladder pain is used in mice and rats.

マウスの膀胱痛に対するモデルとしての膀胱膨満誘発内臓運動応答

Approximately 3-8 million people in the United States suffer from interstitial cystitis/bladder pain syndrome (IC/BPS), a debilitating condition ...

Remote Limb Ischemic Preconditioning:  A Neuroprotective Technique in Rodents

Sublethal ischemia protects tissues against subsequent, more severe ischemia through the upregulation of endogenous mechanisms in the affected tissue. Sublethal ischemia has also been shown to upregulate protective mechanisms in remote tissues. A brief period of ischemia (5-10 min) in the hind limb of mammals induces self-protective responses in the brain, lung, heart and retina. The effect is known as remote ischemic preconditioning (RIP). It is a therapeutically promising way of protecting vital organs, and is already under clinical trials for heart and brain injuries. This publication demonstrates a controlled, minimally invasive method of making a limb &#8211; specifically the hind limb of a rat &#8211; ischemic. A blood pressure cuff developed for use in human neonates is connected to a manual sphygmomanometer and used to apply 160 mmHg pressure around the upper part of the hind limb. A probe designed to detect skin temperature is used to verify the ischemia, by recording the drop in skin temperature caused by pressure-induced occlusion of the leg arteries, and the rise in temperature which follows release of the cuff. This method of RIP affords protection to the rat retina against bright light-induced damage and degeneration.

Remote ischemic preconditioning (RIP) is a method of conditioning tissues against damaging stress. We have established a method of remote ischemia at the hind limb, by inflating a sphygmomanometer cuff for 5-10 min. The neuroprotective capabilities of RIP have been demonstrated in a model of retinal degeneration in rodents.

リモート四肢虚血プレコンディショニング：げっ歯類における神経保護技術

Sublethal ischemia protects tissues against subsequent, more severe ischemia through the upregulation of endogenous mechanisms in the affected tissue. ...

Performing Subretinal Injections in Rodents to Deliver Retinal Pigment Epithelium Cells in Suspension

The conversion of light into electrical impulses occurs in the outer retina and is accomplished largely by rod and cone photoreceptors and retinal pigment epithelium (RPE) cells. RPE provide critical support for photoreceptors and death or dysfunction of RPE cells is characteristic of age-related macular degeneration (AMD), the leading cause of permanent vision loss in people age 55 and older. While no cure for AMD has been identified, implantation of healthy RPE in diseased eyes may prove to be an effective treatment, and large numbers of RPE cells can be readily generated from pluripotent stem cells. Several interesting questions regarding the safety and efficacy of RPE cell delivery can still be examined in animal models, and well-accepted protocols used to inject RPE have been developed. The technique described here has been used by multiple groups in various studies and involves first creating a hole in the eye with a sharp needle. Then a syringe with a blunt needle loaded with cells is inserted through the hole and passed through the vitreous until it gently touches the RPE. Using this injection method, which is relatively simple and requires minimal equipment, we achieve consistent and efficient integration of stem cell-derived RPE cells in between the host RPE that prevents significant amount of photoreceptor degeneration in animal models. While not part of the actual protocol, we also describe how to determine the extent of the trauma induced by the injection, and how to verify that the cells were injected into the subretinal space using in vivo imaging modalities. Finally, the use of this protocol is not limited to RPE cells; it may be used to inject any compound or cell into the subretinal space.

Here we present a community accepted protocol in multimedia format for subretinally injecting a bolus of RPE cells in rats and mice. This approach can be used for determining rescue potentials, safety profiles, and survival capacities of grafted RPE cells upon implantation in animal models of retinal degeneration.

懸濁液中の網膜色素上皮細胞を送達するげっ歯類における網膜下注射の実行

The conversion of light into electrical impulses occurs in the outer retina and is accomplished largely by rod and cone photoreceptors and retinal pigment ...

Evaluation of the Efficacy of the H. pylori Protein HP-NAP as a Therapeutic Tool for Treatment of Bladder Cancer in an Orthotopic Murine Model

Bladder cancer is one of the most common malignancies of the urogenital tract. Intravesical injection of Bacillus Calmette-Gu&#233;rin (BCG) is the gold standard treatment for the high-grade non-muscle invasive bladder cancer (NMIBC). However, since the treatment-related side effects are relevant, newer biological response modifiers with a better benefit/side effects ratio are needed.
The tumour microenvironment can influence both tumour development and therapy efficacy. In order to obtain a good model, it is desirable to implant tumour cells in the organ from which the cancer originates.
In this protocol, we describe a method for establishing a tumour in the bladder cavity of female mice and subsequent delivery of therapeutic agents; the latter are exemplified by our use of Helicobacter pylori neutrophil activating protein (HP-NAP). A preliminary chemical burn of the mucosa, followed by the injection of mouse urothelial carcinoma cell line MB49 via urethral catheterization, enables the cells to attach to the bladder mucosa. After a period, required to allow an initial proliferation of the cells, mice are treated with HP-NAP, administrated again via catheterization. The anti-tumour activity of HP-NAP is evaluated comparing the tumour volume, the extent of necrosis and the degree of vascularization between vehicle- and HP-NAP-treated animals.

Evaluation of the Efficacy of the H. pylori Protein HP-NAP as a Therapeutic Tool for Treatment of Bladder Cancer in an Orthotopic Murine Model

Here the method to establish a syngeneic mouse model of orthotopic bladder tumour to evaluate the anti-tumour efficacy of the bacterial protein HP-NAP is described.

の有効性の評価<em&gt; H。ピロリ</em同所性マウスモデルにおける膀胱癌の治療のための治療ツールとして&gt;タンパク質HP-NAP

Bladder cancer is one of the most common malignancies of the urogenital tract. Intravesical injection of Bacillus Calmette-Gu&#233;rin (BCG) is the gold ...

Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

Several in vivo pre-clinical studies in Polycystic Kidney Disease (PKD) utilize orthologous rodent models to identify and study the genetic and molecular mechanisms responsible for the disease, and are very convenient for rapid drug screening and testing of promising therapies. A limiting factor in these studies is often the lack of efficient non-invasive methods for sequentially analyzing the anatomical and functional changes in the kidney. Magnetic resonance imaging (MRI) is the current gold standard imaging technique to follow autosomal dominant polycystic kidney disease (ADPKD) patients, providing excellent soft tissue contrast and anatomic detail and allowing Total Kidney Volume (TKV) measurements.A major advantage of MRI in rodent models of PKD is the possibility for in vivo imaging allowing for longitudinal studies that use the same animal and therefore reducing the total number of animals required. In this manuscript, we will focus on using Ultra-high field (UHF) MRI to non-invasively acquire in vivo images of rodent models for PKD. The main goal of this work is to introduce the use of MRI as a tool for in vivo phenotypical characterization and drug monitoring in rodent models for PKD.

Use of Ultra-high Field MRI in Small Rodent Models of Polycystic Kidney Disease for In Vivo Phenotyping and Drug Monitoring

The use of ultra-high field MRI as a non-invasive way to obtain phenotypic information of rodent models for polycystic kidney disease and to monitor interventions is described. Compared with the traditional histological approach, MRI images can be acquired in vivo, allowing for longitudinal follow-up.

のための多発性嚢胞腎疾患の小齧歯類モデルにおける超高磁場MRIの使用<em&gt;インビボ</em&gt;表現型と薬物モニタリング

Several in vivo pre-clinical studies in Polycystic Kidney Disease (PKD) utilize orthologous rodent models to identify and study the genetic and molecular ...

Use of a Piglet Model for the Study of Anesthetic-induced Developmental Neurotoxicity (AIDN): A Translational Neuroscience Approach

Anesthesia cannot be avoided in many cases when surgery is required, particularly in children. Recent investigations in animals have raised concerns that anesthesia exposure may lead to neuronal apoptosis, known as anesthesia-induced developmental neurotoxicity (AIDN). Furthermore, some clinical studies in children have suggested that anesthesia exposure may lead to neurodevelopmental deficits later in life. Nonetheless, an ideal animal model for preclinical study has yet to be developed. The neonatal piglet represents a valuable model for preclinical study, as they share a striking number of developmental similarities with humans.
The anatomy and physiology of piglets allow for implementation of rigorous human perioperative conditions in both survival and non-survival procedures. Femoral artery catheterization allows for close monitoring, thus enabling prompt correction of any deviation of the piglet's vital signs and chemistries. In addition, there are multiple developmental similarities between piglets and human neonates. The techniques required to use piglets for experimentation will require experience to master. A pediatric anesthesiologist is a critical member of the investigative team. We describe, in a general sense, the appropriate use of a piglet model for neurodevelopmental study.

Use of a Piglet Model for the Study of Anesthetic-induced Developmental Neurotoxicity AIDN: A Translational Neuroscience Approach

Anesthesia-induced developmental neurotoxicity (AIDN) research has focused on rodents, which are not broadly applicable to humans. Non-human primate models are more relevant, but are cost-prohibitive and difficult to use for experimentation. The piglet, in contrast, is a clinically relevant, practical animal model ideal for the study of anesthetic neurotoxicity.

麻酔誘発性発達神経毒（AIDN）研究のための子豚モデルの使用：翻訳神経科学的アプローチ

Anesthesia cannot be avoided in many cases when surgery is required, particularly in children.  Recent investigations in animals have raised concerns that ...