Name: preparation of the rat vocal fold for neuromuscular analyses
Uploaded: 2020-05-15T14:45:00.000+00:00
Duration: 7 min 17 s
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この研究は、国立国立衛生研究所の難聴やその他のコミュニケーション障害研究所のF31DC017053-01A1(LENELL、PI)およびK23DC014517(ジョンソン、PI)の助成金によって支えられた。

この研究は、ニューヨーク大学医学部の施設動物ケア・使用委員会に従って行われました。
1. ラット喉頭を解剖する
<ol><li>制度的に承認されたプロトコルに従ってラットを安楽死させる。組織標本の毛皮の汚染を防ぐために、下顎からマニュリウムに腹側の首を剃り、アルコールで綿棒を剃ります。</li>
	<li>10倍の拡大率を持つ解剖スコープの下で、気管が露出するまでメスで中線の首の切開を作成することによって喉頭全体を切除する。</li>
	<li>中線で腹側外線喉頭筋を分離し、鉗子と解剖ハサミまたはメスを使用して喉頭を露出させます。</li>
	<li>気管尾骨を第3気管リングに切断し、舌骨に切開ロストラルを作り、切除ハサミを使って喉頭全体を切除する。</li>
	<li>拡大縮小の下で、マイクロディスセクションツール(ピンセット、ピン、ピン、マイクロシザー)を使用して喉頭から外因性喉頭組織(食道、甲状腺、外回喉頭筋)を取り除きます。</li>
	<li>マイクロシザーでは、後部コラリテノイド筋肉の間の正中線をランドマークとして使用して、喉頭をarytenoidの間にドーセリーします。喉頭の横壁をピンで留め、声帯を露出させ、次いでマイクロシザーで声の前部のコミュシャの間の甲状腺軟骨の正中線を通って腹孔を二分する(図1)。 注: この手順はオプションです。喉頭全体を保つためにスキップすることができます。喉頭の二角切除は、同じ喉頭の左右の側面を別々に使用することにより、複数の免疫染色技術を可能にする。</li>
	<li>各ヘミ喉頭をリン酸緩衝溶液(PBS)で~10秒にリンスし、凍結中の氷晶形成を低減する作業ワイパーで繊細に乾燥させます。</li>
</ol>2. 喉頭組織の固定および/またはフラッシュフリーズ
注: 固定は、すべての免疫染色プロトコルに最適とはいえない場合があります。多くの場合、喉頭組織は、解剖直後に新鮮なフラッシュ冷凍である。ステップ 2.1 をスキップして、固定せずに喉頭組織をフラッシュフリーズします。
<ol><li>ヘミ喉頭を固定するために、PBSで4%ホルムアルデヒドを充填した遠心管に組織を配置し、70rpmで軌道シェーカー上の室温で1時間置く。ティッシュをきれいな遠心管に移し、PBSで20分間3倍洗いします。その後、クリーン遠心管に移し、4°Cで20%スクロース/5%グリセロール溶液(約18時間または組織が沈むまで)に沈水する。 注意:ホルムアルデヒドは危険であり、適切な個人用保護具と一緒にヒュームフードで使用する必要があります。</li>
	<li>すべてのヘミ喉頭を均一な位置に配置し、最適な切削温度(OCT)化合物で満たされたクライオモールドに入れます。ヘミラNXの場合は、声帯の内側の表面がクリオマールの底に面し、声楽の縦方向の側面がクリオマール開口部の下端と平行に配置されます。喉頭全体の場合、クリオマールの底に向かって後部クリコアリテノイドと、声楽の縦方向の側面を、クリオマール開口部の下端と平行に配置します。 注:OCTコンパウンド内の一貫した喉頭配は、ラットの声帯の凍結切断に重要です。ヘミヤNXが埋め込まれ凍結されると、その向きを変更するために解凍する必要があり、それによって、複数の融解凍結サイクルによる組織損傷のリスクを導入する。</li>
	<li>液体窒素に囲まれたスチールビーカーで冷やされたイセペンタン(2-メチルブタン)を用いたフラッシュフリーズ組織。 注:白い沈殿物がビーカー13の側面および底部に形成され始めると、isopentaneは組織凍結のための最適な温度に達する。イセペタンは液体窒素よりも熱伝導率が高いため使用され、急速凍結時の組織ブロックの割れを防ぐのに役立ちます。OTCにおける凍結組織のより詳細な説明については、クマールら13を参照してください。</li>
	<li>各金型を事前にラベル付けされたホイルで包み、脱水を防ぐために個々の冷凍袋に入れ、-80°C冷凍庫に保管するために移されるまですぐにドライアイスに保管します。</li>
</ol>3. 断面面におけるクライオセクションヘミヤクス
<ol><li>クライオスタットのチャンバー温度を-20 °Cに設定し、メーカーのマニュアルで筋肉組織の切断に推奨される温度範囲(15-25 °C)の真ん中にあります。</li>
	<li>クライオスタットセクションの厚さを10μmの厚いセクションに設定します。 注:筋線維断字解析では、10 μmの厚い断面が最適で、繊維タイピング解析用のラベル付き筋繊維の完全な染色と堅牢なイメージング強度を可能にします14,15,16。いくつかのプロトコルは、神経筋の標的に応じて異なるセクション厚さを必要とするかもしれない。</li>
	<li>凍結スタットチャンバーに組織を移し、凍結スタット標本ディスク(チャック)にOCT化合物の均一な層を追加し、組み込まれた組織ブロックをOCT化合物の上に置きます。チロイルテノイド(TA)筋線維解析のために声帯の断面を得るために、腹側甲状腺軟骨がクライオスタットブレードに面し、アリテノイド軟骨が検体円盤に面するように、試料をチャックに貼り付ける。 注: これらのランドマークは、OCT コンパウンドが凍結すると白色になり不透明になるため、この段階では表示されないことに注意してください。この視界の欠如は、フラッシュ凍結段階でヘミラリクスの向きに注意することが重要である理由です。</li>
	<li>甲状腺軟骨の腹側部分が現れるまで、検体頭部を100μm進めることによりOCT化合物をトリムする。</li>
	<li>次に、甲状腺軟骨の発症から、層状のプロプリア、内側TA筋肉、および横TA筋肉が露出するまでの30μmの切片をトリムして追跡します。 注:喉頭のランドマークは、断面の角度が斜めではないことを確認するために、100 μmごとに甲状腺軟骨の発症から追跡および注意する必要があります。 図2 は、10倍の倍率で断面面の喉頭ランドマークの2つのセットを表しています。</li>
	<li>ターゲットTA筋肉に到達したら、10μmで正に帯電したスライドのセクションを収集します。</li>
	<li>4°CでPBSにセクションを保管し、汚れる準備ができるまで水分を保持します。 注:固定組織はIHCターゲットに応じて1週間までPBSに保存できますが、固定されていない組織は直ちに処理する必要があります。</li>
</ol>4. 縦方向の平面におけるクライオセクションヘミナクス
<ol><li>クライオスタットチャンバーを再び-20°Cに設定して、断面の厚さを30μmに変更します。 注:NMJ分析では、30-60 μmの間の組織厚を使用して、神経末端またはモータエンドプレート11,12,17の断片化を伴わずに喉頭筋内の複数の完全なNmJをキャプチャすることができます。</li>
	<li>TA筋肉のNMJ分析のための縦方向の声帯セクションを得るために、喉頭蓋がクライオスタットブレードに向き、気管腔が検体ディスクに向かって下向きになるように、試料をチャックに貼り付ける。</li>
	<li>甲状腺軟骨が現れるまで、検体頭部を100μm進めてOCT化合物をトリミングします。</li>
	<li>TA筋の層および内側の部分および内側の分裂が露出するまで、甲状腺の発症から30μmのトリムおよびトラックセクション。 注:縦方向の平面の喉頭のランドマークの5つのセットは、ターゲットTA筋肉に向かって組織の深さの進行を追跡することをお勧めします。 図3 は、10倍の倍率で縦方向の面における喉頭のランドマークを表しています。</li>
	<li>ターゲットTA筋肉に到達したら、30μmで正に帯電したスライドのセクションを収集します。</li>
	<li>4°CでPBSにセクションを保管し、汚れる準備ができるまで水分を保持します。</li>
</ol>

<ol>
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著者らは開示するものは何もない。

神経筋解析のためのラットの声帯を準備すると、様々な課題を提示することができます。喉頭筋は軟骨に囲まれ小さいだけでなく、標的筋を直接抽出することが困難となり、喉頭解剖学的ランドマークの深部に動物間に高いばらつきも見られた。筋肉の断面平面プロトコルの場合、完全な声の折り目のセクションは、腹側甲状腺軟骨の最初の出現後に21-85セクション(1セクションあたり10 μm)?...

ラット喉頭は、発達、老化、疾患および薬理学的薬剤1,2,3,4,5に対する構造的および機能的な神経筋喉頭適応を調査するための確立されたモデルである。組織学的方法の一貫性は、筋肉の準備と分析に関与する複数の複雑さだけでなく、喉頭軟骨内にカプセル化された筋肉の喉頭の大きさ、形状、および地形に関連する課題があるので、この作業ラインに不可欠です1,6,7,8,9,10,11 .ラットの本来の喉頭筋のサイズが小さいため、系統的埋め込み、凍結、および凍結切断は、一貫した正確な結果を達成するために重要です。例えば、ラットの声帯をコロナ平面で切り離すとき、4つの内在性喉頭筋の神経筋接合(NJ)は、組織深度11の1.8mm未満以内に位置する。したがって、凍結切断中の喉頭筋解剖学の正確なモニタリングは、対象のセクションを正確に特定し、組織の過剰切除を防ぐことが不可欠です。ターゲット筋肉の過大な切断は、NMJs11の数と地形の不正確な識別をもたらすか、または目標筋肉がランドマークの向きの混乱のために廃棄された場合、サンプルサイズの全体的な減少をもたらす可能性があります12。喉頭筋の研究とそれぞれの適応の新しいモデルが開発される中で、結果が正確で信頼性が高く、研究全体で再現可能であることを保証するためには、標準的な手術手順が不可欠です。
本稿の目的は、最適な縦断および断面分析のためのラットの声の折り目の調製を詳述することです。当研究室で定期的に使用される詳細な方法は、凍結切断中に標的筋のランドマークを同定するために記述されています。同様の方法は複数の研究室で使用されていますが、初心者の研究者によって実装された場合に信頼性と正確なレプリケーションを確実にするために、文献よりも詳細な情報が提供されています。このチュートリアルの目的は、検査室と調査の一貫性を向上させるために、ラットの声帯の免疫組織化学(IHC)評価のための標準的な方法論を提供することです。

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preparation of the rat vocal fold for neuromuscular analyses

このチュートリアルの目的は、神経筋の研究のためのラットの声の折り目の準備を説明することです。このプロトコルは、ラット喉頭解剖、フラッシュ凍結、および声帯の凍結切断の手順を概説する。本研究では、縦方向および断面面の両方で声を折るクライオセクションの方法を説明する。このプロトコルの目新しさは、本質的な喉頭筋の正確な同定を保証し、組織喪失の可能性を低減する、凍結切断中の喉頭追跡である。図は、両方の平面の進行性の凍結切断を示しています。29匹のラットヘミ喉頭は、甲状腺軟骨の出現から、完全な声の折り目を含む最初のセクションの出現まで、凍結切断され、追跡された。完全なボーカルフォールドは、両方の平面のすべての動物のために視覚化されました。甲状腺軟骨の出現から両方の平面におけるフルボーカルフォールドの出現までの距離には、高いばらつきがあった。体重は喉頭のランドマークの深さと相関しておらず、個々の変動性および組織製剤に関連する他の要因が、断面化中のランドマークの出現における高い変動性を引き起たす可能性があることを示唆した。本研究は、方法論を詳述し、組織化学的神経筋の調査のためにラットの声の折り目を準備するための形態学的データを提示する。個人の変動性が高いため、喉頭のランドマークは、組織および組織の損失を切除することを防ぐために、凍結切断中に密接に追跡する必要があります。ラット喉頭内のランドマークの適切な組織準備と意識を含む一貫した方法論の使用は、研究全体で一貫した結果を支援し、喉頭神経筋メカニズムを調査するためのモデルとしてラットボーカルフォールドを使用することに興味を持つ新しい研究者を支援します。

代表的な結果は、喉頭神経筋系における声の運動の影響に関する継続的な調査の一環であった。29匹の雄フィッシャー344/褐色ノルウェーラット(生後12匹、24ヶ月齢12匹)を計量し、CO2 吸入で安楽死させ、続いて両側性乳房離脱手術を行った。
手順は、NmJと側面および内側TA筋肉の繊維サイズにラベルを付けるために概説されたプロトコルに従った。喉頭のランドマ?...

Watch this Scientific Journal Video about Preparation of the Rat Vocal Fold for Neuromuscular Analyses at JoVE.com

Preparation of the Rat Vocal Fold for Neuromuscular Analyses

このプロトコルは、検査用神経筋研究のためのラットの声の折り目を準備するために使用される方法を記述します。

神経筋解析のためのラット声帯の調製

The purpose of this tutorial is to describe the preparation of the rat vocal fold for histochemical neuromuscular study. This protocol outlines procedures ...

preparation-of-the-rat-vocal-fold-for-neuromuscular-analyses

Research

JoVE Journal

Neuroscience

2.7K ビュー.  New York University School of Medicine. このプロトコルは、検査用神経筋研究のためのラットの声の折り目を準備するために使用される方法を記述します。

ビデオ: Preparation of the Rat Vocal Fold for Neuromuscular Analyses

Physiological, Morphological and Neurochemical Characterization of Neurons Modulated by Movement

The role of individual neurons and their function in neuronal circuits is fundamental to understanding the neuronal mechanisms of sensory and motor functions. Most investigations of sensorimotor mechanisms rely on either examination of neurons while an animal is static1,2 or record extracellular neuronal activity during a movement.3,4 While these studies have provided the fundamental background for sensorimotor function, they either do not evaluate functional information which occurs during a movement or are limited in their ability to fully characterize the anatomy, physiology and neurochemical phenotype of the neuron. A technique is shown here which allows extensive characterization of individual neurons during an in vivo movement. This technique can be used not only to study primary afferent neurons but also to characterize motoneurons and sensorimotor interneurons. Initially the response of a single neuron is recorded using electrophysiological methods during various movements of the mandible followed by determination of the receptive field for the neuron. A neuronal tracer is then intracellularly injected into the neuron and the brain is processed so that the neuron can be visualized with light, electron or confocal microscopy (Fig. 1). The detailed morphology of the characterized neuron is then reconstructed so that neuronal morphology can be correlated with the physiological response of the neuron (Figs. 2,3). In this communication important key details and tips for successful implementation of this technique are provided. Valuable additional information can be determined for the neuron under study by combining this method with other techniques. Retrograde neuronal labeling can be used to determine neurons with which the labeled neuron synapses; thus allowing detailed determination of neuronal circuitry. Immunocytochemistry can be combined with this method to examine neurotransmitters within the labeled neuron and to determine the chemical phenotypes of neurons with which the labeled neuron synapses. The labeled neuron can also be processed for electron microscopy to determine the ultrastructural features and microcircuitry of the labeled neuron. Overall this technique is a powerful method to thoroughly characterize neurons during in vivo movement thus allowing substantial insight into the role of the neuron in sensorimotor function.

A technique is described to quantify the in vivo physiological response of mammalian neurons during movement and correlate the physiology of the neuron with neuronal morphology, neurochemical phenotype and synaptic microcircuitry.

運動によって変調された神経細胞の生理的、形態学的および神経化学的キャラクタリゼーション

The role of individual neurons and their function in neuronal circuits is fundamental to understanding the neuronal mechanisms of sensory and motor ...

神経科学

Targeted Training of Ultrasonic Vocalizations in Aged and Parkinsonian Rats

Voice deficits are a common complication of both Parkinson disease (PD) and aging; they can significantly diminish quality of life by impacting communication abilities. 1, 2 Targeted training (speech/voice therapy) can improve specific voice deficits,3, 4 although the underlying mechanisms of behavioral interventions are not well understood. Systematic investigation of voice deficits and therapy should consider many factors that are difficult to control in humans, such as age, home environment, age post-onset of disease, severity of disease, and medications. The method presented here uses an animal model of vocalization that allows for systematic study of how underlying sensorimotor mechanisms change with targeted voice training. The ultrasonic recording and analysis procedures outlined in this protocol are applicable to any investigation of rodent ultrasonic vocalizations.
The ultrasonic vocalizations of rodents are emerging as a valuable model to investigate the neural substrates of behavior.5-8 Both rodent and human vocalizations carry semiotic value and are produced by modifying an egressive airflow with a laryngeal constriction.9, 10 Thus, rodent vocalizations may be a useful model to study voice deficits in a sensorimotor context. Further, rat models allow us to study the neurobiological underpinnings of recovery from deficits with targeted training.
To model PD we use Long-Evans rats (Charles River Laboratories International, Inc.) and induce parkinsonism by a unilateral infusion of 7 &mu;g of 6-hydroxydopamine (6-OHDA) into the medial forebrain bundle which causes moderate to severe degeneration of presynaptic striatal neurons (for details see Ciucci, 2010).11, 12 For our aging model we use the Fischer 344/Brown Norway F1 (National Institute on Aging).
Our primary method for eliciting vocalizations is to expose sexually-experienced male rats to sexually receptive female rats. When the male becomes interested in the female, the female is removed and the male continues to vocalize. By rewarding complex vocalizations with food or water, both the number of complex vocalizations and the rate of vocalizations can be increased (Figure 1).
An ultrasonic microphone mounted above the male's home cage records the vocalizations. Recording begins after the female rat is removed to isolate the male calls. Vocalizations can be viewed in real time for training or recorded and analyzed offline. By recording and acoustically analyzing vocalizations before and after vocal training, the effects of disease and restoration of normal function with training can be assessed. This model also allows us to relate the observed behavioral (vocal) improvements to changes in the brain and neuromuscular system.

Voice disorders are debilitating in aging and Parkinson disease. The ultrasonic vocalizations of rats, also affected by these conditions, can be used to study these voice disorders, their neural substrates, and the nature of functional recovery with behavioral intervention.

高齢者とパーキンソンラットにおける超音波発声の対象とトレーニング

Voice deficits are a common complication of both Parkinson disease (PD) and aging; they can significantly diminish quality of life by impacting ...

Subcutaneous Neurotrophin 4 Infusion Using Osmotic Pumps or Direct Muscular Injection Enhances Aging Rat Laryngeal Muscles

Laryngeal dysfunction in the elderly is a major cause of disability, from voice disorders to dysphagia and loss of airway protective reflexes. Few, if any, therapies exist that target age-related laryngeal muscle dysfunction. Neurotrophins are involved in muscle innervation and differentiation of neuromuscular junctions (NMJs). It is thought that neurotrophins enhance neuromuscular transmission by increasing neurotransmitter release. The neuromuscular junctions (NMJs) become smaller and less abundant in aging rat laryngeal muscles, with evidence of functional denervation. We explored the effects of NTF4 for future clinical use as a therapeutic to improve function in aging human laryngeal muscles. Here, we provide the detailed protocol for systemic application and direct injection of NTF4 to investigate the ability of aging rat laryngeal muscle to remodel in response to NTF4 application. In this method, rats either received NTF4 either systemically via osmotic pump or by direct injection through the vocal folds. Laryngeal muscles were then dissected and used for histological examination of morphology and age-related denervation.

Here, we present a protocol to describe the use of neurotrophin 4 (NTF4) systemically and directly to remodel rat aging laryngeal muscles.

浸透圧ポンプまたは直接的筋肉注射を用いる皮下ニューロトロフィン4注入は、老齢ラット喉頭筋を増強する

Laryngeal dysfunction in the elderly is a major cause of disability, from voice disorders to dysphagia and loss of airway protective reflexes. Few, if ...

発生生物学

Transplantation of Olfactory Ensheathing Cells to Evaluate Functional Recovery after Peripheral Nerve Injury

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth and regrowth of the primary olfactory neurons. Indeed, the primary olfactory system is characterized by its ability to give rise to new neurons even in adult animals. This particular ability is partly due to the presence of OECs which create a favorable microenvironment for neurogenesis. This property of OECs has been used for cellular transplantation such as in spinal cord injury models. Although the peripheral nervous system has a greater capacity to regenerate after nerve injury than the central nervous system, complete sections induce misrouting during axonal regrowth in particular after facial of laryngeal nerve transection. Specifically, full sectioning of the recurrent laryngeal nerve (RLN) induces aberrant axonal regrowth resulting in synkinesis of the vocal cords. In this specific model, we showed that OECs transplantation efficiently increases axonal regrowth.
OECs are constituted of several subpopulations present in both the olfactory mucosa (OM-OECs) and the olfactory bulbs (OB-OECs). We present here a model of cellular transplantation based on the use of these different subpopulations of OECs in a RLN injury model. Using this paradigm, primary cultures of OB-OECs and OM-OECs were transplanted in Matrigel after section and anastomosis of the RLN. Two months after surgery, we evaluated transplanted animals by complementary analyses based on videolaryngoscopy, electromyography (EMG), and histological studies. First, videolaryngoscopy allowed us to evaluate laryngeal functions, in particular muscular cocontractions phenomena. Then, EMG analyses demonstrated richness and synchronization of muscular activities. Finally, histological studies based on toluidine blue staining allowed the quantification of the number and profile of myelinated fibers.
All together, we describe here how to isolate, culture, identify and transplant OECs from OM and OB after RLN section-anastomosis and how to evaluate and analyze the efficiency of these transplanted cells on axonal regrowth and laryngeal functions.

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth of the primary olfactory neurons. This specific property can be used for cellular transplantation. We present here a model of cellular transplantation based on the use of OECs in a laryngeal nerve injury model.

末梢神経損傷後の機能回復を評価する嗅神経鞘細胞の移植

Olfactory ensheathing cells (OECs) are neural crest cells which allow growth and regrowth of the primary olfactory neurons. Indeed, the primary olfactory ...

A Murine Model of Cervical Spinal Cord Injury to Study Post-lesional Respiratory Neuroplasticity

A cervical spinal cord injury induces permanent paralysis, and often leads to respiratory distress. To date, no efficient therapeutics have been developed to improve/ameliorate the respiratory failure following high cervical spinal cord injury (SCI). Here we propose a murine pre-clinical model of high SCI at the cervical 2 (C2) metameric level to study diverse post-lesional respiratory neuroplasticity. The technique consists of a surgical partial injury at the C2 level, which will induce a hemiparalysis of the diaphragm due to a deafferentation of the phrenic motoneurons from the respiratory centers located in the brainstem. The contralateral side of the injury remains intact and allows the animal recovery. Unlike other SCIs which affect the locomotor function (at the thoracic and lumbar level), the respiratory function does not require animal motivation and the quantification of the deficit/recovery can be easily performed (diaphragm and phrenic nerve recordings, whole body ventilation). This pre-clinical C2 SCI model is a powerful, useful, and reliable pre-clinical model to study various respiratory and non-respiratory neuroplasticity events at different levels (molecular to physiology) and to test diverse putative therapeutic strategies which might improve the respiration in SCI patients.

Respiratory failure is the leading cause of death following a cervical spinal cord injury. Having a reproducible, quantifiable, and reliable pre-clinical animal model of respiratory failure induced by a partial cervical injury will help to understand the subsequent respiratory and non-respiratory neuroplasticity and allow testing putative repair strategies.

ポスト病変呼吸神経可塑性を研究するための子宮頸脊髄損傷のマウスモデル

A cervical spinal cord injury induces permanent paralysis, and often leads to respiratory distress. To date, no efficient therapeutics have been developed ...

医学

多層シリコーン声帯モデルの3Dプリンティング

Manufacturing Process for Non-Adhesive Super-Soft Vocal Fold Models

This study aims to develop super-soft, non-sticky vocal fold models for voice research. The conventional manufacturing process of silicone-based vocal fold models results in models with undesirable properties, such as stickiness and reproducibility issues. Those vocal fold models are prone to rapid aging, leading to poor comparability across different measurements. In this study, we propose a modification to the manufacturing process by changing the order of layering the silicone material, which leads to the production of non-sticky and highly consistent vocal fold models. We also compare a model produced using this method with a conventionally manufactured vocal fold model that is adversely affected by its sticky surface. We detail the manufacturing process and characterize the properties of the models for potential applications. The outcomes of the study demonstrate the efficacy of the modified fabrication method, highlighting the superior qualities of our non-sticky vocal fold models.&#160;The findings contribute to the development of realistic and reliable vocal fold models for research and clinical applications.

著者スポットライト:音声およびロボットアプリケーションのための合成声帯モデルの製造における進歩

This study demonstrates the manufacturing of non-sticky and super-soft vocal fold models by introducing a specific way to create the vocal fold layers, providing a detailed description of the manufacturing procedure, and characterizing the properties of the models.

非粘着性超軟質ボーカルフォールドモデルの製造工程

This study aims to develop super-soft, non-sticky vocal fold models for voice research. The conventional manufacturing process of silicone-based vocal ...

工学

In Vivo Evaluation of the Mechanical and Viscoelastic Properties of the Rat Tongue

The tongue is a highly innervated and vascularized muscle hydrostat on the floor of the mouth of most vertebrates. Its primary functions include supporting mastication and deglutition, as well as taste-sensing and phonetics. Accordingly, the strength and volume of the tongue can impact the ability of vertebrates to accomplish basic activities such as feeding, communicating, and breathing. Human patients with sleep apnea have enlarged tongues, characterized by reduced muscle tone and increased intramuscular fat that can be visualized and quantified by magnetic resonance imaging (MRI). The abilities to measure force generation and viscoelastic properties of the tongue constitute important tools for obtaining functional information to correlate with imaging data. Here, we present techniques for measuring tongue force production in anesthetized Zucker rats via electrical stimulation of the hypoglossal nerves and for determining the viscoelastic properties of the tongue by applying passive Lissajous force/deformation curves.

In Vivo Evaluation of the Mechanical and Viscoelastic Properties of the Rat Tongue

We describe a surgical procedure in an anesthetized rat model for determining the muscle tone and viscoelastic properties of the tongue. The procedure involves specific stimulation of the hypoglossal nerves and application of passive Lissajous force/deformation curves to the muscle.

ラット舌の機械的および粘弾性的特性のインビボ評価

The tongue is a highly innervated and vascularized muscle hydrostat on the floor of the mouth of most vertebrates. Its primary functions include ...

Endoscopic Approach for Studying Laryngeal Dysfunction in Rodents

Minimally Invasive Murine Laryngoscopy for Close&#45;Up Imaging of Laryngeal Motion During Breathing and Swallowing

The larynx is an essential organ in mammals with three primary functions - breathing, swallowing, and vocalizing. A wide range of disorders are known to impair laryngeal function, which results in difficulty&#160;breathing (dyspnea), swallowing impairment (dysphagia), and/or voice impairment (dysphonia). Dysphagia, in particular, can lead to aspiration pneumonia and associated morbidity, recurrent hospitalization, and early mortality. Despite these serious consequences, existing treatments for laryngeal dysfunction are largely aimed at surgical and behavioral interventions that unfortunately do not typically restore normal laryngeal function, thus highlighting the urgent need for innovative solutions.
To bridge this gap, we have been developing an experimental endoscopic approach to investigate laryngeal dysfunction in murine (i.e., mouse and rat) models. However, endoscopy in rodents is quite challenging due to their small size relative to current endoscope technology, anatomical differences in the upper airway, and the necessity for anesthesia to optimally access the larynx. Here, we describe a novel transoral laryngoscopy approach that permits close-up, unobstructed video imaging of laryngeal motion in mice and rats. Critical steps in the protocol include precise anesthesia management (to prevent overdosing that abolishes swallowing and/or risks respiratory distress-related mortality) and micromanipulator control of the endoscope (for stable video recording of laryngeal motion by a single researcher for subsequent quantification).
Importantly, the protocol can be performed over time in the same animals to study the impact of various pathological conditions specifically on laryngeal function. A novel advantage of this protocol is the ability to visualize airway protection during swallowing, which is not possible in humans due to epiglottic inversion over the laryngeal inlet that obstructs the glottis from view. Rodents therefore provide a unique opportunity to specifically investigate the mechanisms of normal versus pathological laryngeal airway protection for the ultimate purpose of discovering treatments to effectively restore normal laryngeal function.

This protocol describes a serial transoral laryngoscopy approach for mice and rats that permits close-up, unobstructed video imaging of the larynx during breathing and swallowing using an optimized anesthetic regimen and finely tuned endoscopic manipulation techniques.

The larynx is an essential organ in mammals with three primary functions - breathing, swallowing, and vocalizing. A wide range of disorders are known to ...

神経筋解析のためのラット声帯の調製

要約

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運動によって変調された神経細胞の生理的、形態学的および神経化学的キャラクタリゼーション

高齢者とパーキンソンラットにおける超音波発声の対象とトレーニング

浸透圧ポンプまたは直接的筋肉注射を用いる皮下ニューロトロフィン4注入は、老齢ラット喉頭筋を増強する

末梢神経損傷後の機能回復を評価する嗅神経鞘細胞の移植

ポスト病変呼吸神経可塑性を研究するための子宮頸脊髄損傷のマウスモデル

非粘着性超軟質ボーカルフォールドモデルの製造工程

非粘着性超軟質ボーカルフォールドモデルの製造工程

非粘着性超軟質ボーカルフォールドモデルの製造工程

ラット舌の機械的および粘弾性的特性のインビボ評価

Minimally Invasive Murine Laryngoscopy for Close-Up Imaging of Laryngeal Motion During Breathing and Swallowing