本研究はLM2018126、チェコ科学アカデミーRVO 68378050、MEYS CR、OP RDE CZ.02.1.01/0.0/0.0/16_013/0001789(チェコ現象ゲノミクスセンターのアップグレード:MEYSとESIFによる翻訳研究に向けての開発)、OP RDE CZ.02.1.01/0.0/0.0/18_046/0015861(MEYSとESIFによるCCPインフラストラクチャアップグレードII)、およびOP RDI CZ.1.05/2.1.00/19.0395(MEYSとERDFによるトランスジェニックモデルの高品質と容量)の支援を受けました。さらに、この研究は、チェコ共和国の教育青年スポーツ省が提供するNGO「遺伝子治療協会(ASGENT)」、チェコ(https://asgent.org/)およびチェコLM2023036フェノゲノミクスセンターから資金提供を受けました。

行動試験を用いたアンジェルマン症候群のマウスモデルの検証

著者は、開示すべき利益相反はありません。

異なるマウス系統で作成されたASモデルは、ヒトの症状との比較を容易にするために、動物の感情状態、運動機能、および認知能力のテストで一般的に検証されます31,32。ASモデルの運動欠損は、実験室全体で最も一貫した所見であり、変異体の感情状態が変化しておらず、巣作りが困難であることが続いています31,32,33。<...

アンジェルマン症候群(AS)はまれな神経発達疾患です。ASの最も一般的な遺伝的起源は、母体由来の染色体の15q11-q13領域の大きな欠失であり、これは患者のほぼ74%に見られます1。この領域が欠失すると、E3ユビキチンリガーゼをコードするASの主な原因遺伝子であるUBE3Aが失われます。ニューロンにおけるUBE3A遺伝子の父方対立遺伝子は、刷り込みとして知られるプロセスでサイレンシングされます。その結果、遺伝子の父方の刷り込みは、中枢神経系(CNS)2での母方の発現のみを可能にします。したがって、母体由来の染色体からのUBE3A遺伝子欠失は、AS症状の発症につながる。ヒトでは、ASは生後約6か月で発症し、発達遅滞はすべての発達段階を通じて持続し、影響を受けた個人に重度の衰弱症状をもたらします3,4。この障害の主な症状には、ぎくしゃくした運動失調歩行、重度の言語障害、知的障害など、細かい運動能力と粗大運動能力の欠如が含まれます。AS患者の約80%も睡眠障害やてんかんに苦しんでいます。今日まで、利用可能な唯一の治療法は、てんかん発作を軽減し、睡眠の質を改善する対症療法薬です1。したがって、再現可能な行動表現型を有する堅牢な動物モデルの開発と洗練された表現型解析は、疾患の病態生理学的メカニズムを解明し、効果的な薬物療法と治療法を発見するために不可欠です。
中枢神経系に影響を与えるヒトの疾患の複雑さは、モデル生物が同等のゲノム、生理学、および行動を有することを要求します。マウスは、生殖周期が短く、サイズが小さく、DNA修飾が比較的容易であるため、モデル生物として人気があります。1984年、Paul Willnerは、モデルの値を決定するために使用される3つの基本的な疾患モデル検証基準、つまり、構成、顔、および予測妥当性を提案しました5。簡単に言えば、構築物の妥当性は障害の発症に関与する生物学的メカニズムを反映し、顔の妥当性はその症状を要約し、予測妥当性は治療薬に対するモデル反応を記述します。
上記の原則を遵守するために、最も一般的な遺伝的病因であるUBE3A遺伝子を含む母体の15q11.2-13q遺伝子座の大量欠失を選択し、ASモデルマウスを作成しました。CRISPR/Cas9技術を用いて、C57BL/6Nバックグラウンドからマウスにおいて、UBE3A遺伝子全体にまたがる76,225 bpの長さの領域を欠失させ、遺伝子のコード要素と非コード要素の両方を網羅しました6。その後、これらの動物を繁殖させ、UBE3A+/-ヘテロ接合型マウスを得た。モデルの顔面検証のために、UBE3A+/-雌と野生型の雄の交配動物を用いてUBE3A+/-子孫(C57BL/6NCrl-UBE3A<em1(IMPC)Ccpcz>/Phと命名され、後にUBE3A mGenedel/+として割り当てられた株)を獲得し、同腹仔を対照した。コアASの症状を再現するために、彼らの細かい運動能力と肉眼的な運動能力、感情、および影響をテストしました。前回の記事では、AS患者も知的障害に苦しんでいるため、動物の認知機能も評価しました6。しかし、UBE3AmGenedel/+マウスでは、おそらく試験7の時点で動物の年齢が若かったため、認知障害は見られませんでした。その後、約18週齢の高齢動物を調べたところ、場所選好パラダイムでの逆転学習中の行動の柔軟性の欠如が明らかになりました。ただし、この分析に使用される機器の複雑さには、別の方法論モジュールが必要であり、ここでは含まれていません。
ここで提示された行動テストは、その高い予測値と十分な構成概念の有効性のおかげで、遺伝子研究における一般的な表現型ツールに属します8,9,10。これらのテストを使用して、ヒト疾患の中核症状を再現可能な年齢に依存しない方法で再現することにより、ASのマウスモデルを検証しました。動物の感情性は、高架プラス迷路およびオープンフィールドテストで評価されました。これらのテストは両方とも、動物が食物、避難所、または交尾の機会を求めて新しい環境を探索すると同時に、不安原性コンパートメントを回避するアプローチ回避の対立に基づいています11。さらに、オープンフィールドテストは、マウスの自発運動をテストするために使用されます8。尾懸垂試験は、マウスノックアウトモデル12における新しい抗うつ薬またはうつ病様表現型をスクリーニングするために、うつ病研究で広く使用されています。このテストでは、避けられない状況で動物が時間の経過とともに発達する絶望を評価します。運動学習と詳細な歩行特性は、それぞれロタロッドとデジゲイトで決定されました。加速棒での動物の持久力は、そのバランスと運動協調能力を特徴づけますが、マウスのステップパターンの詳細な分析は、多くの神経発生運動障害に関連する神経筋障害の敏感な評価です13、14、15。ネストレットシュレッダーテストは、げっ歯類の衝動的な行動を検出するための標準的な方法論の一部であり、自然なげっ歯類の建物の行動を利用しているため、動物の健康状態を示します16,17。
実験群の規模は、3Rルールの要求を満たすための妥協とコロニー育種性能の効率的な使用の結果でした。しかし、統計的検出力を得るために、十分な量の繁殖ペアが確立されているため、グループには10人以上の個体がいました。残念ながら、繁殖性能は必ずしも十分な数の動物をもたらすわけではありませんでした。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>Cages, individually ventilated</td>
			<td>Techniplast</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>DigiGait</td>
			<td>Mouse Specifics, Inc., 2 Central Street Level 
			Unit 110 
			Framingham, MA 01701, USA</td>
			<td>Equipment was tendered, no catalogue&#160; number was provided, nor could be find on company&#39;s web site</td>
			<td>Detailed analysis of mouse gait, hardware and software provided.&#160;</td>
		</tr>
		<tr>
			<td>FDA Nestlet squares</td>
			<td>Datesand Ltd., 7 Horsfield Way, Bredbury, Stockport SK6, UK</td>
			<td>Material was bought from Velaz vendor via direct email request. Velaz do not provide any catalogue no.</td>
			<td>Cotton nestlets for nest building test. Nestlet discription: 2-3 g each, with diameter around 5 x 5 x 0.5cm.</td>
		</tr>
		<tr>
			<td>Mouse chow</td>
			<td>Altramion</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Rotarod</td>
			<td>TSE Systems GmbH, Barbara-McClintock-Str.4 
			12489 Berlin, Germany</td>
			<td>Equipment was tendered, no catalogue&#160; number was provided, nor could be find on company&#39;s web site</td>
			<td>Rotarod for 5 mice, hardware and software provided. Drum dimensions: Diameter: 30 mm, width per lane: 50 mm, falling distance 147 mm.</td>
		</tr>
		<tr>
			<td>Tail Suspension Test</td>
			<td>Bioseb, In Vivo Research Instruments, 13845 Vitrolles 
			FRANCE</td>
			<td>Reference: BIO-TST5</td>
			<td>Fully automated equipment for immobility time evaluation of 3 mice hanged by tail, hardware and software provided</td>
		</tr>
		<tr>
			<td>Transpore medical tape</td>
			<td>Medical M, Ltd.</td>
			<td>P-AIRO1291</td>
			<td>The tape used to attach an animal to the hook by its tail.</td>
		</tr>
		<tr>
			<td>Viewer - Video Tracking System</td>
			<td>Biobserve GmbH, Wilhelmstr. 23 A 
			53111 Bonn, Germany</td>
			<td>Equipment with software were tendered, no catalogue&#160; number was provided, nor could be find on company&#39;s web site</td>
			<td>Software with custom made hardware: maze, IR base, IR sensitive cameras. Custom-made OF dimensions: 42 x 42 cm area, 49 cm high wall, central zone area: 39 cm2. A custom-made EPM was elevated 50 cm above the floor, with an open arm 79 cm long,&#160; 9 cm wide, and closed arm 77 cm long, 7.6 cm wide.&#160;</td>
		</tr>
	</tbody>
</table>

behavioral characterization of an angelman syndrome mouse model

この原稿は、ASの確立されたマウスモデルにおけるアンジェルマン症候群(AS)様表現型を特徴付けるために利用可能な一連の行動試験について説明しています。ロタロッド学習パラダイム、詳細な歩行分析、巣作りテストを使用して、動物の運動障害を検出および特徴付けます。オープンフィールドでの動物の感情と高架プラス迷路テスト、およびテールサスペンションテストでの影響をテストします。ASマウスをオープンフィールドテストでテストする場合、運動機能障害は迷路でのマウスの行動に影響を与え、活動スコアを変化させるため、結果を慎重に解釈する必要があります。
提示された行動テストの再現性と有効性は、異なるノックアウトバリアントを持ついくつかの独立したUba3aマウス系統ですでに検証されており、この一連のテストをAS研究における優れた検証ツールとして確立しています。関連する構成と顔の妥当性を備えたモデルは、疾患の病態生理学を解明し、原因となる治療法の開発を可能にするために、さらなる調査を必要とします。

Angelman syndrome (AS) is a rare neurodevelopmental disease. The most common genetic origin of AS is a large deletion of the 15q11-q13 region of the maternally-derived chromosome, which is found in nearly 74% of patients1. Deletion of this region causes the loss of UBE3A, the main causative gene of AS that encodes an E3 ubiquitin ligase. The paternal allele of the UBE3A gene in neurons is silenced in a process known as imprinting. As a consequence, paternal imprinting of the gene allows only maternal expression in the central nervous system (CNS)2. Therefore, UBE3A gene deletion from the maternally-derived chromosome leads to the development of AS symptoms. In humans, AS manifests at around 6 months of age, with developmental retardation that persists throughout all developmental stages and results in severe debilitating symptoms in affected individuals3,4. The core symptoms of the disorder include the deficit of fine and gross motor skills, including jerky ataxic gait, serious speech impairment, and intellectual disability. Approximately 80% of AS patients also suffer from sleep disturbances and epilepsy. To date, the only available treatment are symptomatic drugs, which reduce epileptic seizures and improve sleep quality1. Therefore, the development of robust animal models with reproducible behavioral phenotypes alongside refined phenotyping analysis will be essential to elucidate the pathophysiological mechanisms of the disorder and discover effective medications and treatments.
The complexity of the human disorder affecting the CNS demands model organisms to possess a comparable genome, physiology, and behavior. Mice are popular as a model organism due to their short reproductive cycle, small size, and relative ease of DNA modification. In 1984, Paul Willner proposed three basic disease model validation criteria: the construct, face, and predictive validity, which are used to determine the model&#39;s value5. Simply, construct validity reflects the biological mechanisms responsible for the disorder development, face validity recapitulates its symptoms, and predictive validity describes the model response to therapeutic drugs.
To adhere to the above principles, we have chosen the most common genetic etiology, a large deletion of the maternal 15q11.2-13q locus including the UBE3A gene, to create AS model mice. We used the CRISPR/Cas9 technique to delete a 76,225 bp long region spanning the entire UBE3A gene, encompassing both the coding and non-coding elements of the gene, in mice from a C57BL/6N background6. We then bred the animals to obtain UBE3A+/&#8722; heterozygous mice. For face validation of the model, we used animals from crosses of UBE3A+/&#8722; females and wild-type males to gain UBE3A+/- progeny (strain named C57BL/6NCrl-UBE3A&#60;em1(IMPC)Ccpcz&#62;/Ph and later assigned as UBE3AmGenedel/+) and control littermates. We tested their fine and gross motor skills, emotionality, and affect to recapitulate core AS symptoms. In a previous article, we also evaluated the animals&#39; cognitive functions, as AS patients also suffer from intellectual disability6. However, we found no cognitive impairments in UBE3AmGenedel/+ mice, perhaps due to the young age of the animals at the time of testing7. Later examination of the older animals, around 18 weeks old, revealed a deficit in behavioral flexibility during reversal learning in the place preference paradigm. However, the complexity of the employed equipment for this analysis requires a separate methodological module and it is not included here.
The behavioral tests presented here belong to the common phenotyping tools in genetic research, thanks to their high predictive value and sufficient construct validity8,9,10. We used these tests to validate a mouse model of AS by recapitulating core symptoms of the human disease in a reproducible, age-independent manner. The emotionality of the animal was evaluated in the elevated plus maze and open field tests. Both of these tests are based on the approach-avoidance conflict, where animals explore a new environment in search of food, shelter, or mating opportunities while simultaneously avoiding anxiogenic compartments11. Additionally, the open field test is used to test a mouse&#39;s locomotor activity8. The tail suspension test is widely used in depression research to screen for new antidepressant drugs or depressive-like phenotypes in mouse knockout models12. This test evaluates the despair that animals develop over time in an inescapable situation. Motor learning and detailed gait characteristics were determined on the rotarod and in DigiGait, respectively. Animal endurance on the accelerating rod characterizes its balance and movement coordination skills, while detailed analysis of a mouse&#39;s step patterns is a sensitive evaluation of neuromuscular impairments connected to many neurogenerative movement disorders13,14,15. The nestlet shredding test is part of the standard methodology for detecting impulsive behavior in rodents, and since it utilizes natural rodent building behavior, it indicates the animal&#39;s well-being16,17.
The size of the experimental groups was a result of a compromise to meet the 3R rule demands and efficient usage of colony breeding performance. However, to obtain statistical power, the groups had no fewer than 10 individuals, due to the establishment of a sufficient amount of breeding pairs. Unfortunately, breeding performance did not always result in a sufficient number of animals.

著者スポットライト:アンジェルマン症候群マウスモデルを検証するための再現性の高い一連の行動試験 

アンジェルマン症候群モデルマウスの行動特性

Our goal is to elucidate the physiological roles of genes using mice as an experimental model. We also focus on developing mouse models of specific diseases that scientists can use to investigate disease causation and develop effective treatments. An example of such activity is the mouse model of Angelman syndrome.Arguably, the most promising new technology with potential is integrating artificial intelligence based on deep learning techniques in animal research. This approach can be used to monitor animal behavior in the home cage directly without need for the experimenter to disturb the animals. The biggest challenge in behavioral studies currently is reproducibility.Therefore, we introduce a battery of tests that validate the Angelman syndrome of model in a way that does not depend on the model's age or species. We use established behavioral tests to validate our AS model. However, with the novel approach for the model development closely resembling the human condition with a large deletion.Using CRISPR technology, we deleted the entire UBE3A gene from the mouse genome. In contrast to other models where the deletion is limited to 3KB. We're working on developing a genetic therapy that could restore a symptoms in a mouse model.

高架プラス迷路とオープンフィールドテスト EPMおよびOFテストでは、げっ歯類が新しい環境を探索する自然な傾向を使用します18,19。探査は、げっ歯類が新しい環境の探査と起こりうる危険の回避のどちらかを選択するアプローチ回避紛争によって支配されています。動物は、避難所、社会的接触、または採餌を求めて未知の場所を?...

Watch this Scientific Journal Video about A Battery of Highly Reproducible Behavioral Tests to Validate an Angelman Syndrome Murine Model at JoVE.com

この原稿は、アンジェルマン症候群マウスモデルを検証するための再現性の高い行動テストのセットを提示します。

This manuscript describes a battery of behavioral tests available to characterize Angelman syndrome (AS)-like phenotypes in an established murine model of ...

私たちの目標は、マウスを実験モデルとして、遺伝子の生理的役割を解明することです。また、科学者が疾患の原因を調査し、効果的な治療法を開発するために使用できる特定の疾患のマウスモデルの開発にも焦点を当てています。そのような活性の例は、アンジェルマン症候群のマウスモデルである。間違いなく、可能性を秘めた最も有望な新技術は、動物研究における深層学習技術に基づく人工知能の統合です。このアプローチは、実験者が動物を邪魔することなく、ホームケージ内の動物の行動を直接監視するために使用できます。現在の行動研究における最大の課題は再現性です。そこで、モデルの年齢や種に依存しない方法でモデルのアンジェルマン症候群を検証する一連のテストを紹介します。確立された動作テストを使用して、ASモデルを検証します。しかし、モデル開発のための新しいアプローチでは、大きな欠失を伴う人間の状態によく似ています。CRISPR技術を用いて、マウスゲノムからUBE3A遺伝子全体を欠失させました。削除が3KBに制限されている他のモデルとは対照的です。私たちは、マウスモデルの症状を回復できる遺伝子治療の開発に取り組んでいます。

behavioral-characterization-of-an-angelman-syndrome-mouse-model

Research

JoVE Journal

Behavior

Behavioral Characterization of an Angelman Syndrome Mouse Model

1.6K ビュー.  Institute of Molecular Genetics of the Czech Academy of Sciences. 私たちの目標は、マウスを実験モデルとして、遺伝子の生理的役割を解明することです。また、科学者が疾患の原因を調査し、効果的な治療法を開発するために使用できる特定の疾患のマウスモデルの開発にも焦点を当てています。そのような活性の例は、アンジェルマン症候群のマウスモデルである。間違いなく、可能性を秘めた最も有望な新技術は、動物研究?...

ビデオ: 著者スポットライト:アンジェルマン症候群マウスモデルを検証するための再現性の高い一連の行動試験 

This manuscript describes a battery of behavioral tests available to characterize Angelman syndrome (AS)-like phenotypes in an established murine model of AS. We use the rotarod learning paradigm, detailed gait analysis, and nest building test to detect and characterize animal motor impairments. We test animal emotionality in the open field and elevated plus maze tests, as well as the affect in the tail suspension test. When AS mice are tested in the open field test, the results should be interpreted with care, since motor dysfunctions influence mouse behavior in the maze and alter activity scores.
The reproducibility and effectiveness of the presented behavioral tests has already been validated in several independent Uba3a mouse lines with different knockout variants, establishing this set of tests as an excellent validation tool in AS research. Models with the relevant construct and face validity will warrant further investigations to elucidate the pathophysiology of the disease and grant the development of causal treatments.

This manuscript presents a set of highly reproducible behavioral tests to validate an Angelman syndrome mouse model.

行動学

Elevated Plus Maze Test in an Angelman Syndrome Mouse Model

To begin, place the plus shaped maze on the testing platform just below the camera. Using the potentiometer on the wall, adjust the light intensity to 70 luxe at its center with the help of a luxometer with its sensor placed at the center of the maze during the adjustment. Open the software by double clicking on the viewer software icon and load the configuration for Elevated Plus Maze or EPM testing by clicking on the icon on the upper left side of the configuration tab.Enter the animal information into the corresponding fields of the experiment tab including animal id, genotype, gender, and experiment information. Verify that the zone's position, open arms, and closed arms are correctly configured. Using a visual control and computer mouse, ensure that the virtual outline zones match the corresponding EPM zones.Place the mouse cursor on the arrow icon on the upper left side of the acquisition tab. Remove the animal from the home cage and place it gently in the center of the EPM. Start the experiment by left clicking on the computer mouse.After five minutes, save the recorded data by clicking okay. Name the file and click save. Export the results to a csv file for offline analysis by clicking on the icon on the left vertical panel of the data analysis tab.In the EPM test, none of the parameters related to anxiety-like behavior were altered in maternal heterozygous Ube3A mutants.

アンジェルマン症候群マウスモデルにおける上昇プラス迷路テスト

Open Field Test in an Angelman Syndrome Mouse Model

Place the four open field, or OF, test boxes on the testing platform below the camera. Using the potentiometer on the wall, adjust the light intensity to 200 lux at the center of each OF test with the help of a luxometer with its sensor placed at the center of each box during the adjustment. Open the software by double clicking on the viewer software icon and load the configuration for EF testing by clicking on the icon on the upper left side of the configuration tab.Enter the animal information into the corresponding fields of the experiment tab. Verify that the zone's position matches the OF test boxes and adjust them if needed. Using a visual control and computer mouse, ensure that the virtual outline center and periphery zones match the corresponding OF test zones.Place the mouse cursor on the arrow icon on the upper left side of the acquisition tab. Remove four animals from the home cage and place them in the corner of each test box. Stop the experiment by left clicking on the computer mouse.In the OF test, the maternal heterozygous Ube3a mutant animals were significantly hypoactive as reflected by a shorter traversed distance, lower average speed, and longer resting time. However, none of the parameters related to anxiety-like behavior were altered in Ube3a mutants.

アンジェルマン症候群マウスモデルにおけるオープンフィールド試験

Tail Suspension Test in an Angelman Syndrome Mouse Model

Connect the tail suspension cage to the computer using a USB cable. Insert the USB dongle into the computer and start the software by double clicking on the BioTSD software icon. In the settings tab on the global, ensure the acquisition duration is set to 360 seconds.In the experiment tab, select the new list of subjects and create a new experiment file and a new list of tested subjects by following the instructions in the opened tab. Prepare the animals for the test by wrapping single-sided adhesive tape around three-fourths of the animal's tail starting from the base. Start the run by clicking start run then continue in the acquisition tab.Immediately after hanging the animal on the hook, click the start icon under the visualized position to acquire data. After testing all the animals, in the analysis tab, select the last four minutes of the acquisition for analysis. Select all valid runs in the analysis period and click analyze selected subjects.Choose the desired data format and click export selected data to export the collected data. In the tail suspension test, Ube3a mutant animals were immobile for significantly longer than their control littermates, indicating their depression-like behavior.

アンジェルマン症候群マウスモデルにおける尾懸垂試験

Gait Analysis in an Angelman Syndrome Mouse Model

Turn on the treadmill and manually set the belt speed to 20 centimeters per second on the equipment panel before switching on the apparatus light. Launch the DigiGait Imager software and set the shutter speed to 100 for albino mice or 130 for black or dark mice in the field for shutter speed. Place the mouse onto the treadmill belt and confirm that the mouse can perform the test by setting it to a slow walking speed for around three seconds and then stopping it while observing the mouse continuously.Start the belt by pressing the start button and record for 10 seconds. Ensure that a clear and fluent locomotion of at least 10 to 15 steps is observable. Stop the belt by pressing the stop button and return the mouse to the temporary holding cage.Screen the recording for a sequence of images with fluent steps by clicking play and reviewing the recording with the visual control in edit mode. Select 10 to 15 fluent movements by manually writing their starting and ending frame numbers into the relevant fields. Enter the animal information, including animal ID, date of birth, sex, weight, belt speed, and belt angle.Save the file by clicking save. Next, open the DigiGait Analysis software and perform gait analysis based on a fully automated analysis of video recordings of animal footprints. The gait indices of UBE3A mutant animals were altered.UBE3A mutant animals had a longer swing and stance that resulted in prolonged stride duration and length. Mice's hind limbs, propulsion, duration, and deceleration were also increased. These findings also revealed a larger pore area at the peak stance.

アンジェルマン症候群マウスモデルにおける歩行解析

Rotarod Test in an Angelman Syndrome Mouse Model

Turn on the rotarod equipment and launch the software by double-clicking the rod software icon. Create a new file in the file tab and save it with an appropriate name. In the setup window, fill out the experiment details, such as the date, user's name, and any eventual comments.Set the speed profile to 300 seconds, initial speed to 4 RPM, and terminal speed to 40 RPM. Open the measuring panel by clicking Measure. Start the initial rotation of the rod at 4 RPM by clicking Start and place the first five animals in their assigned positions.Start the testing protocol by clicking start profile, and the rod will gradually accelerate to 40 RPM over five minutes. After all the animals have fallen from the rod, close the Measure window by clicking Close and click Show to display the collected data. Export the acquired data in csv format by clicking Export CSV.Rotarod performance showed a reduced latency to fall in UBE3A mutant animals.

アンジェルマン症候群マウスモデルにおけるロータロッド試験

Nestlet Shredding: Nest Building Test in an Angelman Syndrome Mouse Model

Take a cotton nestlet with forceps and record its weight manually using scales. Place the nest lit randomly in a cage but on the opposite side of the water supply. Weigh each nestlet for the next four days manually using scales.Ensure that the nestlet is dry when weighed. If not dry on a heating pad. If the nestlet is torn into several parts, weigh the largest part.Record the weights on paper or in a pre-made spreadsheet. The UBE3A mutant animals used significantly less material to build their nests and this difference was particularly prominent between transgenic females and their control counterparts.