这项工作由奥地利科学院 (星期三) 的一个分开的津贴支持, 捷克共和国项目的技术代办处没有。TA04010877 (星期三、VH 和 JGS) 和捷克科学基金会 (GACR) 项目没有 GA16-01246S (JGS)。我们感谢 w 泰康的建议, 他使用假牙固定霜, 和 Ing。p. Liska 从捷克陆军森林服务处获得帮助, 获得了 larynges 的切除鹿。

本文分析的动物标本是按照捷克共和国奥洛穆茨 Palacky 大学的标准伦理要求进行处理的。它们来源于在森林中生活的红鹿, 在一个正常狩猎季节, 它们被捷克陆军森林局猎杀。
1. 半喉标本的制备
注意: 如4所示, 只应使用正确准备的标本。快速冷冻喉部25在切除和储存-80 ° c 后立即将组织退化和生物力学特性改变的可能性降到最低, 并允许在任何方便的时间进行实验。
<ol><li>
除霜喉
	<ol>
<li>将冷冻喉插入两个高压釜袋或任何其他带有防水密封的塑料袋中。密封袋, 并把它们放入水浴加热到30° c, 直到喉完全解冻。所需的时间范围从几个小时到超过一天, 取决于喉的大小和冻结温度。</li>
	</ol>
</li>
	<li>
清洁喉部
	<ol>
<li>喉解冻后, 将其从袋中取出, 用生理盐水 (0.9% 氯化钠) 彻底清洁。</li>
		<li>小心地去除多余的组织 (如外颈部肌肉, 舌骨等) 而不损害主喉结构, 并将气管缩短至足够的长度, 以将喉安装在供气管上 (通常约4-5 厘米)。</li>
		<li>检查喉组织的潜在组织异常, 如伤口, 有机变形, 或裂缝可能发生的冻结过程, 这可能使喉不适合的实验。</li>
	</ol>
</li>
	<li>
甲状腺和环状软骨软骨的暴露
	<ol>
<li>用手术刀除去甲状腺和环状软骨软骨周围的喉部肌肉组织的部分, 从而使软骨在准备 mid-sagittal 切口时产生半喉。此准备阶段在图 1A和1B中描述。</li>
	</ol>
</li>
	<li>
甲状腺软骨中矢状切口
	<ol>
<li>通过甲状腺软骨的前部进行初始垂直切口。</li>
		<li>小心地将切口稍微放在将要被移除的一侧, 以免破坏需要保持的声音褶皱。如果可能, 使用手术刀切割。如果软骨是僵化的, 使用一个小锯。</li>
	</ol>
</li>
	<li>
切割环状软骨软骨
	<ol>
<li>引导切口垂直 (下) 从在杓软骨之间, 然后通过环状软骨软骨到一个近似水平水平的下等甲状腺切口。</li>
	</ol>
</li>
	<li>
切除一声带, 在喉部形成 L 形切口
	<ol>
<li>从先前做的环状软骨软骨的垂直切口的下端开始进行水平切口, 并将新的切口引向下甲状腺切口。前折叠喉的一侧将被删除。</li>
		<li>通过在甲状腺软骨内侧的软组织上进行垂直切开--要小心, 同时将声带前部附着之间的切口引导到甲状腺软骨, 从而避免声带损伤。</li>
	</ol>
</li>
	<li>
切开通过甲状腺软骨的提炼
	<ol>
<li>使用手术刀、锯子或锉, 以便在甲状腺软骨中应用精确的直切口, 并尽可能接近先前检查过的声带的前部。</li>
		<li>也去除后甲状腺软骨的一个小部分, 为了创造空间为插入的刺为 adducting 杓软骨和因而声音折叠 (如下所示)。此准备阶段在图 1C和1D中描述。 注: 根据研究问题, 可能需要对整个声带进行充分的阐述, 以使其可见性。在这种情况下, 应该去掉 (真实的) 声带上方的结构 (即心室或前庭的褶皱, 如适用于标本的解剖)。在一些标本中, 声带上方的喉部软组织可能会失去与甲状腺软骨的连接, 并在振动时干扰声带, 有可能造成假 (大多不规则) 振荡模式。在这种情况下, 仔细清除该组织是不可避免的。</li>
	</ol>
</li>
</ol><img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/55303/55303fig1.jpg"> 图 1: 半喉准备和安装.(A) 和 (B) 清除喉标本、内侧和后视, 然后去除左声带;(C) 和 (D) 准备了 L 形切口 (左声带移除)、内侧和后视的半喉。<a href="//cloudflare.jove.com/files/ftp_upload/55303/55303fig1large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>

2. 半喉实验
<ol><li>
半喉安装
	<ol>
<li>使用一个供气管, 提供温暖和湿润的空气进入喉。</li>
		<li>构造两个垂直排列的透明板作为替代切除喉部。</li>
		<li>使用尖头4提高喉部的稳定性, 并通过将剩余的声音折叠 adducting 到垂直玻璃板 (参见图 2A), 从而创建适当的 pre-phonatory 喉配置。 注: 理论上, 在滑轮-杠杆系统26中, 声带和重量也可能内。然而, 对这些作者最好的了解, 这种方法尚未被尝试进行 hemilarynx 的准备。</li>
	</ol>
</li>
</ol><img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/55303/55303fig2.jpg">
图 2: 半喉设置.(A) 支承结构: 供气管、L 形玻璃板排列、收叉。(B) 装入的半喉准备。(C) 和 (D) 半喉准备的特写镜头, 分别从侧面和顶部观看。<a href="//cloudflare.jove.com/files/ftp_upload/55303/55303fig2large.jpg" target="_blank">请单击此处查看此图的较大版本.</a>
<ol start="2"><li>
安装半喉
	<ol>
<li>用假牙固定膏盖住供气管, 用气管的剩余部分安装喉部。定影霜的作用是粘合剂, 并关闭潜在的间隙, 从而产生密封。</li>
		<li>用塑料收紧带或软管钳固定气管。</li>
		<li>还要用固定膏覆盖切口通过甲状腺软骨的边缘, 同时避免在声带或喉内软组织上涂上固定乳膏。</li>
		<li>附上透明的盘子。</li>
	</ol>
</li>
	<li>
甲状腺软骨的稳定, 声带的内收, 使用尖头
	<ol>
<li>用尖头将声带加到板上, 稳定甲状腺软骨。</li>
		<li>固定乳膏设置后, 应用气流以建立声带振荡, 并检查在半喉和玻璃板之间可能出现的渗漏。</li>
		<li>通过添加更多的固定乳膏来密封最终出现的间隙。</li>
	</ol>
</li>
</ol>

作者没有什么可透露的。

半喉预备具有 "常规" (完全) 切除喉设置的优点: 在这种实验方法中, 物理和生理边界条件和参数 (如门下压力或声带伸长) 可以控制得相当好。hemilarynx 的行为与一个完全的横向对称的全喉是同源的, 除了某些参数的大小 (e. g, 空气流速, 声压) 减少了大约 50%, 但仍然是在实际范围为16。完全切除喉的方法的主要缺点, i. e., 由于在半喉设置中克服了声带表面的不可见性 (ca...

声音通常是由振动喉组织 (主要是声带) 产生的, 它将由肺部提供的稳定气流转换成一系列气流脉冲。声波压力波形 (即, 主要声音) 出现从这个序列的流动脉冲声学兴奋的声带过滤他们, 并由此产生的声音从口和 (在一定程度上) 从鼻子辐射1.所产生的声音的光谱组成很大程度上受声带振动质量的影响, 受喉生物力学和与气管气流的相互作用2。无论是在临床和研究的背景下, 文献和评价声带振动因此是最重要的兴趣时, 研究语音生产。
在人类中, 对喉的直接内窥镜调查在酣然的生产在体内是富有挑战性的, 并且它几乎不可能在非人的哺乳动物, 鉴于当前技术手段。因此, 为了保证仔细控制物理和/或生理实验的边界条件, 使用切除 larynges3,4在许多情况下是充分替代研究的在体内语音生成机制。
声音褶皱振动是一个复杂的三维现象5。传统的调查方法, 如喉内窥镜 (在体内) 或切除喉的准备工作通常只提供一个优越的观点振动声乐褶皱6, 他们不允许完整的三维分析声音折叠运动。特别是在高级视图中, 在振动周期的主要部分中, 声带的下部 (尾缘) 边缘是不可见的。这是由于下 (尾) 与声带的上级 (颅) 边缘之间的相位延迟, 这一现象通常在声带振荡5时出现。直接经验主义的证据为备份发现从数学和物理模型是缺乏的, 知识的几何和运动的更低的声带边缘7, 并且因而几何门下渠道8,9,10对于更好地理解喉气流、声带组织和产生的力和压力之间的相互作用至关重要,1112。声带振动的另一个方面是隐藏在传统的优越观点是垂直 (caudo 颅) 的接触之间的两个声音褶皱的深度。垂直接触深度与声带的垂向厚度有关, 这是歌唱中用于演唱 ("胸部" 与 "假声" 寄存器)1314的潜在指示器。
为了克服常规 (充分) 切除喉准备的缺点, 可以使用 so-called 半喉设置, 其中一半喉被删除, 从而促进评估的振动特性的剩余声带在三维度。令人惊讶的是, 自从在二十世纪六十年代15中引入此设置并在 1993年16中对概念进行了初步验证之后, 没有多少实验室用这种有希望的实验方法进行了实验17,18 ,19,20,21,22,23。对此的解释可能存在于建立一个可行的半喉准备的困难。虽然常规切除 (完整) 喉准备是很好的文件4, 没有这样的 in-depth 指令是目前尚未可用于创建一个半喉设置。因此, 本文的目的是提供一个教程, 建立一个可靠的可再生的半喉设置, 并辅以红鹿标本的实验结果。
一个半喉安装与 "常规" 切除喉设置共享许多功能, 如测量设备, 高速或其他成像技术, 以充分记录在声音产生的喉结构的振动, 或适当供应加热, 湿润的空气。这些一般的设置注意事项将在书的4和国家语音和语音中心的技术报告24中详细介绍。重申这些指示将超出本手稿的范围。在这里, 只有专门的指令, 生成一个半喉设置提出。

<table><tbody><tr><td>Surgical blades</td><td>Surgeon</td><td>Jai Surgical Ltd., New Delhi, India</td><td></td></tr><tr><td>Saw</td><td>Hand saw (Lux, 150 mm length)</td><td>Lux, Wermelskirchen, Germany</td><td></td></tr><tr><td>Thermometer</td><td>Testo 922</td><td>Testo Ltd., Hampshire, UK</td><td>K-type Probe, Operating temperature -20 to +50 &amp;deg;C</td></tr><tr><td>Autoclave bags</td><td>Autoclave bags</td><td>vwr.com, VWR International s.r.o., Stribrna Skalice, Czech republic</td><td></td></tr><tr><td>Conductive glass plates</td><td>Custom made</td><td>UPOL - Joint laboratory of Optics&lt;br/&gt; Trida 17. listopadu 50A, 772 07 Olomouc, the Czech Rep.</td><td></td></tr><tr><td>Fixative cream</td><td>Denture fixative cream</td><td>Blend-a-dent Natural</td><td></td></tr><tr><td>Prongs and fastening system</td><td>Customized Kanya Al eloxed profiles</td><td>Distributor: VISIMPEX a.s.. Seifertova 33, 750 02 Prerov, the Czech Rep.;</td><td>&amp;nbsp;Combination of Kanya RVS and PVS fastening systems (http://www.kanya.cz/) + custom made prongs</td></tr><tr><td>Mounting tube</td><td>Custom made</td><td>UPOL - Joint laboratory of Optics,&lt;br/&gt; Trida 17. listopadu 50A, 772 07 Olomouc, the Czech Rep.</td><td></td></tr><tr><td>LED Light</td><td>Verbatim 52204 LED Lamp</td><td>Mitsubishi Chemical Holdings Corporation, Tokyo, Japan</td><td></td></tr><tr><td>Camera</td><td>Canon EOS1100D</td><td>Canon Inc.</td><td>18-55 mm lens</td></tr><tr><td>Airpump</td><td>Resun LP100</td><td>Resun</td><td></td></tr><tr><td>Strobe light</td><td>ELMED Helio-Strob micro2</td><td>ELMED Dr. Ing. Mense GmbH, Heiligenhaus, Germany</td><td></td></tr><tr><td>Humidifier</td><td>Custom made</td><td>Voice Research Lab, Dept. Biophysics, Faculty of Sciences, Palacky University Olomouc, Czech republic</td><td></td></tr><tr><td>Subglottic tract</td><td>Custom made adjustable subglottic tract</td><td>Voice Research Lab, Dept. Biophysics, Faculty of Sciences, Palacky University Olomouc, Czech republic</td><td>Hampala, V., Svec, Jan, Schovanek, P., and Mandat, D. Uzitny vzor c. 25585: Model subglotickeho traktu. [Utility model no. 25585: Model of subglottal tract] (In Czech) Soukup, P. 2013-27834(CZ 25505 U1), 1-7. 24-6-2013. Praha, Urad prumysloveho vlastnictvi</td></tr></tbody></table>

hemi-laryngeal setup for studying vocal fold vibration in three dimensions

人类和大多数非人类哺乳动物的声音是通过声带的自我维持振荡在喉部产生的。直接视觉文献的声带振动是挑战, 特别是在非人类的哺乳动物。作为替代, 切除喉实验提供了机会, 研究声带振动在受控生理和物理条件。然而, 使用一个完整的喉只是提供了一个顶部视图的声音褶皱, 不包括关键部分的摆动结构的观察期间, 他们与空气动力学的力量的互动。这一限制可以克服, 利用半喉设置, 其中一半喉是 mid-sagittally 移除, 提供了一个优越的和横向的意见, 其余的声带在自我维持振荡。
在此, 给出了半喉结构的解剖学准备和在实验室工作台上安装的 step-by 步骤指南。发声的半喉准备的模范性的记录与高速视频数据捕获的两个同步相机 (优越和横向的意见), 显示三维声带运动和相应的时变接触面积。该出版物的半喉设置文件将有助于在实验研究中的应用和可靠的重复性, 为语音科学家提供了更好地理解声音生产的生物力学的潜力。

如前一节所提到的, 半喉准备的插图及其在供气管上的安装, 分别在图 1和图 2中提供。
从两个摄像机角度来记录声带振动
气流诱导的半喉声带自我维持振荡从顶部和侧面记录了两个同步高速视频 (HSV) 摄像头在6000帧/秒运行, ...

Watch this Scientific Journal Video about Hemi-laryngeal Setup for Studying Vocal Fold Vibration in Three Dimensions at JoVE.com

Hemi-laryngeal Setup for Studying Vocal Fold Vibration in Three Dimensions

本文介绍了一种用于制备用于促进声音褶皱振动多维观的半喉标本的协议, 目的是研究人类和非人类哺乳动物发声的各种生物物理方面。

三维声带振动的半喉设置研究

The voice of humans and most non-human mammals is generated in the larynx through self-sustaining oscillation of the vocal folds. Direct visual ...

hemi-laryngeal-setup-for-studying-vocal-fold-vibration-three

Research

JoVE Journal

Biology

10.9K 次观看.  Palacky University Olomouc. 本文介绍了一种用于制备用于促进声音褶皱振动多维观的半喉标本的协议, 目的是研究人类和非人类哺乳动物发声的各种生物物理方面。

视频: Hemi-laryngeal Setup for Studying Vocal Fold Vibration in Three Dimensions

Synthetic, Multi-Layer, Self-Oscillating Vocal Fold Model Fabrication

Sound for the human voice is produced via flow-induced vocal fold vibration. The vocal folds consist of several layers of tissue, each with differing material properties 1. Normal voice production relies on healthy tissue and vocal folds, and occurs as a result of complex coupling between aerodynamic, structural dynamic, and acoustic physical phenomena. Voice disorders affect up to 7.5 million annually in the United States alone 2 and often result in significant financial, social, and other quality-of-life difficulties. Understanding the physics of voice production has the potential to significantly benefit voice care, including clinical prevention, diagnosis, and treatment of voice disorders.
Existing methods for studying voice production include in vivo experimentation using human and animal subjects, in vitro experimentation using excised larynges and synthetic models, and computational modeling. Owing to hazardous and difficult instrument access, in vivo experiments are severely limited in scope. Excised larynx experiments have the benefit of anatomical and some physiological realism, but parametric studies involving geometric and material property variables are limited. Further, they are typically only able to be vibrated for relatively short periods of time (typically on the order of minutes).
Overcoming some of the limitations of excised larynx experiments, synthetic vocal fold models are emerging as a complementary tool for studying voice production. Synthetic models can be fabricated with systematic changes to geometry and material properties, allowing for the study of healthy and unhealthy human phonatory aerodynamics, structural dynamics, and acoustics. For example, they have been used to study left-right vocal fold asymmetry 3,4, clinical instrument development 5, laryngeal aerodynamics 6-9, vocal fold contact pressure 10, and subglottal acoustics 11 (a more comprehensive list can be found in Kniesburges et al. 12)
Existing synthetic vocal fold models, however, have either been homogenous (one-layer models) or have been fabricated using two materials of differing stiffness (two-layer models). This approach does not allow for representation of the actual multi-layer structure of the human vocal folds 1 that plays a central role in governing vocal fold flow-induced vibratory response. Consequently, one- and two-layer synthetic vocal fold models have exhibited disadvantages 3,6,8 such as higher onset pressures than what are typical for human phonation (onset pressure is the minimum lung pressure required to initiate vibration), unnaturally large inferior-superior motion, and lack of a "mucosal wave" (a vertically-traveling wave that is characteristic of healthy human vocal fold vibration).
In this paper, fabrication of a model with multiple layers of differing material properties is described. The model layers simulate the multi-layer structure of the human vocal folds, including epithelium, superficial lamina propria (SLP), intermediate and deep lamina propria (i.e., ligament; a fiber is included for anterior-posterior stiffness), and muscle (i.e., body) layers 1. Results are included that show that the model exhibits improved vibratory characteristics over prior one- and two-layer synthetic models, including onset pressure closer to human onset pressure, reduced inferior-superior motion, and evidence of a mucosal wave.

The methodology for fabricating synthetic vocal fold models is described. The models are life-sized and mimic the multi-layer structure of the human vocal folds. Results show the models to self-oscillate at pressures comparable to lung pressure and demonstrate flow-induced vibratory responses that are similar to those of human vocal folds.

合成，多层，自激振荡的声带模型制作

Sound for the human voice is produced via flow-induced vocal fold vibration. The vocal folds consist of several layers of tissue, each with differing ...

科研

生物工程

Chip-based Three-dimensional Cell Culture in Perfused Micro-bioreactors

We have developed a chip-based cell culture system for the three-dimensional cultivation of cells. The chip is typically manufactured from non-biodegradable polymers, e.g., polycarbonate or polymethyl methacrylate by micro injection molding, micro hot embossing or micro thermoforming. But, it can also be manufactured from bio-degradable polymers. Its overall dimensions are 0.7 1 x 20 x 20 x 0.7 1 mm (h x w x l). The main features of the chips used are either a grid of up to 1156 cubic micro-containers (cf-chip) each the size of 120-300 x 300 x 300 &mu; (h x w x l) or round recesses with diameters of 300 &mu; and a depth of 300 &mu; (r-chip). The scaffold can house 10 Mio. cells in a three-dimensional configuration. For an optimal nutrient and gas supply, the chip is inserted in a bioreactor housing. The bioreactor is part of a closed steril circulation loop that, in the simplest configuration, is additionaly comprised of a roller pump and a medium reservoir with a gas supply. The bioreactor can be run in perfusion, superfusion, or even a mixed operation mode. We have successfully cultivated cell lines as well as primary cells over periods of several weeks. For rat primary liver cells we could show a preservation of organotypic functions for more than 2 weeks. For hepatocellular carcinoma cell lines we could show the induction of liver specific genes not or only slightly expressed in standard monolayer culture. The system might also be useful as a stem cell cultivation system since first differentiation experiments with stem cell lines were promising.

We describe a chip-based platform for the three-dimensional cultivation of cells in micro-bioreactors. One chip can house up to 10 Mio. cells that can be cultivated under precisely defined conditions with regard to fluid flow, oxygen tension etc. in a sterile, closed circulation loop.

基于芯片的三维细胞培养，在灌注微生物反应器

We have developed a chip-based cell culture system for the three-dimensional cultivation of cells. The chip is typically manufactured from ...

生物学

Investigating the Three-dimensional Flow Separation Induced by a Model Vocal Fold Polyp

The fluid-structure energy exchange process for normal speech has been studied extensively, but it is not well understood for pathological conditions. Polyps and nodules, which are geometric abnormalities that form on the medial surface of the vocal folds, can disrupt vocal fold dynamics and thus can have devastating consequences on a patient&#39;s ability to communicate. Our laboratory has reported particle image velocimetry (PIV) measurements, within an investigation of a model polyp located on the medial surface of an in&#160;vitro driven vocal fold model, which show that such a geometric abnormality considerably disrupts the glottal jet behavior. This flow field adjustment is a likely reason for the severe degradation of the vocal quality in patients with polyps. A more complete understanding of the formation and propagation of vortical structures from a geometric protuberance, such as a vocal fold polyp, and the resulting influence on the aerodynamic loadings that drive the vocal fold dynamics, is necessary for advancing the treatment of this pathological condition. The present investigation concerns the three-dimensional flow separation induced by a wall-mounted prolate hemispheroid with a 2:1 aspect ratio in cross flow, i.e. a model vocal fold polyp, using an oil-film visualization technique. Unsteady, three-dimensional flow separation and its impact of the wall pressure loading are examined using skin friction line visualization and wall pressure measurements.

Vocal fold polyps can disrupt vocal fold dynamics and thus can have devastating consequences on a patient&#39;s ability to communicate. Three-dimensional flow separation induced by a wall-mounted model polyp and its impact on the wall pressure loading are examined using particle image velocimetry, skin friction line visualization, and wall pressure measurements.

研究模型声乐折叠聚合物引起的三维流量分离

The fluid-structure energy exchange process for normal speech has been studied extensively, but it is not well understood for pathological conditions. ...

Construction and Characterization of a Novel Vocal Fold Bioreactor

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To emulate the dynamic environment of vocal folds, a novel vocal fold bioreactor capable of producing vibratory stimulations at fundamental phonation frequencies is constructed and characterized. The device is composed of a function generator, a power amplifier, a speaker selector and parallel vibration chambers. Individual vibration chambers are created by sandwiching a custom-made silicone membrane between a pair of acrylic blocks. The silicone membrane not only serves as the bottom of the chamber but also provides a mechanism for securing the cell-laden scaffold. Vibration signals, generated by a speaker mounted underneath the bottom acrylic block, are transmitted to the membrane aerodynamically by the oscillating air. Eight identical vibration modules, fixed on two stationary metal bars, are housed in an anti-humidity chamber for long-term operation in a cell culture incubator. The vibration characteristics of the vocal fold bioreactor are analyzed non-destructively using a Laser Doppler Vibrometer (LDV). The utility of the dynamic culture device is demonstrated by culturing cellular constructs in the presence of 200-Hz sinusoidal vibrations with a mid-membrane displacement of 40 &#181;m. Mesenchymal stem cells cultured in the bioreactor respond to the vibratory signals by altering the synthesis and degradation of vocal fold-relevant, extracellular matrix components. The novel bioreactor system presented herein offers an excellent in vitro platform for studying vibration-induced mechanotransduction and for the engineering of functional vocal fold tissues.

A novel vocal fold bioreactor capable of delivering physiologically relevant, vibratory stimulation to cultured cells is constructed and characterized. This dynamic culture device, when combined with a fibrous poly(&#949;-caprolactone) scaffold, creates a vocal fold-mimetic environment that modulates the behaviors of mesenchymal stem cells.

一种新型声带生物反应器的构建与鉴定

In vitro engineering of mechanically active tissues requires the presentation of physiologically relevant mechanical conditions to cultured cells. To ...

Microinjection Techniques for Studying Mitosis in the Drosophila melanogaster Syncytial Embryo

This protocol describes the use of the Drosophila melanogaster syncytial embryo for studying mitosis1. Drosophila has useful genetics with a sequenced genome, and it can be easily maintained and manipulated. Many mitotic mutants exist, and transgenic flies expressing functional fluorescently (e.g. GFP) - tagged mitotic proteins have been and are being generated. Targeted gene expression is possible using the GAL4/UAS system2. 

The Drosophila early embryo carries out multiple mitoses very rapidly (cell cycle duration, &asymp;10 min). It is well suited for imaging mitosis, because during cycles 10-13, nuclei divide rapidly and synchronously without intervening cytokinesis at the surface of the embryo in a single monolayer just underneath the cortex. These rapidly dividing nuclei probably use the same mitotic machinery as other cells, but they are optimized for speed; the checkpoint is generally believed to not be stringent, allowing the study of mitotic proteins whose absence would cause cell cycle arrest in cells with a strong checkpoint. Embryos expressing GFP labeled proteins or microinjected with fluorescently labeled proteins can be easily imaged to follow live dynamics (Fig. 1). In addition, embryos can be microinjected with function-blocking antibodies or inhibitors of specific proteins to study the effect of the loss or perturbation of their function3. These reagents can diffuse throughout the embryo, reaching many spindles to produce a gradient of concentration of inhibitor, which in turn results in a gradient of defects comparable to an allelic series of mutants. Ideally, if the target protein is fluorescently labeled, the gradient of inhibition can be directly visualized4. It is assumed that the strongest phenotype is comparable to the null phenotype, although it is hard to formally exclude the possibility that the antibodies may have dominant effects in rare instances, so rigorous controls and cautious interpretation must be applied. Further away from the injection site, protein function is only partially lost allowing other functions of the target protein to become evident.

Microinjection Techniques for Studying Mitosis in the Drosophila melanogaster Syncytial Embryo

This protocol describes the use of microinjection and high resolution imaging in the Drosophila melanogaster syncytial embryo to study mitosis.

研究有丝分裂的显微注射技术果蝇合胞体胚胎

This protocol describes the use of the Drosophila melanogaster syncytial embryo  for studying mitosis1.  Drosophila has useful genetics with a sequenced ...

多层硅胶声带模型的 3D 打印

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.

作者聚焦：用于语音和机器人应用的合成声带模型的制造进展

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.

非粘性超软人声折叠模型的制造工艺

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

工程学

Patch Clamp and Perfusion Techniques for Studying Ion Channels Expressed in Xenopus oocytes

The protocol presented here is designed to study the activation of the large conductance, voltage- and Ca2+-activated K+ (BK) channels. The protocol may also be used to study the structure-function relationship for other ion channels and neurotransmitter receptors1. BK channels are widely expressed in different tissues and have been implicated in many physiological functions, including regulation of smooth muscle contraction, frequency tuning of inner hair cells and regulation of neurotransmitter release2-6. BK channels are activated by membrane depolarization and by intracellular Ca2+ and Mg2+ 6-9. Therefore, the protocol is designed to control both the membrane voltage and the intracellular solution. In this protocol, messenger RNA of BK channels is injected into Xenopus laevis oocytes (stage V-VI) followed by 2-5 days of incubation at 18&deg;C10-13. Membrane patches that contain single or multiple BK channels are excised with the inside-out configuration using patch clamp techniques10-13. The intracellular side of the patch is perfused with desired solutions during recording so that the channel activation under different conditions can be examined. To summarize, the mRNA of BK channels is injected into Xenopus laevis oocytes to express channel proteins on the oocyte membrane; patch clamp techniques are used to record currents flowing through the channels under controlled voltage and intracellular solutions.

Patch Clamp and Perfusion Techniques for Studying Ion Channels Expressed in Xenopus oocytes

Ionic current of BK channels is recorded using patch clamp techniques. BK channels are expressed in Xenopus oocytes by injecting messenger RNA. The intracellular solution during patch clamp recordings is controlled by a perfusion system.

膜片钳和灌注技术研究离子通道的表达爪蟾卵母细胞

The protocol presented here is designed to study the activation of the large conductance, voltage- and Ca2+-activated K+ (BK) channels. The protocol may ...

Preparation of the Rat Vocal Fold for Neuromuscular Analyses

The purpose of this tutorial is to describe the preparation of the rat vocal fold for histochemical neuromuscular study. This protocol outlines procedures for rat laryngeal dissection, flash-freezing, and cryosectioning of the vocal folds. This study describes how to cryosection vocal folds in both longitudinal and cross-sectional planes. A novelty of this protocol is the laryngeal tracking during cryosectioning that ensures accurate identification of the intrinsic laryngeal muscles and reduces the chance of tissue loss. Figures demonstrate the progressive cryosectioning in both planes. Twenty-nine rat hemi-larynges were cryosectioned and tracked from the emergence of the thyroid cartilage to the appearance of the first section that included the full vocal fold. The full vocal fold was visualized for all animals in both planes. There was high variability in the distance from the appearance of the thyroid cartilage to the appearance of the full vocal fold in both planes. Weight was not correlated to depth of laryngeal landmarks, suggesting individual variability and other factors related to tissue preparation may be responsible for the high variability in the appearance of landmarks during sectioning. This study details a methodology and presents morphological data for preparing the rat vocal fold for histochemical neuromuscular investigation. Due to high individual variability, laryngeal landmarks should be closely tracked during cryosectioning to prevent oversectioning tissue and tissue loss. The use of a consistent methodology, including adequate tissue preparation and awareness of landmarks within the rat larynx, will assist with consistent results across studies and aid new researchers interested in using the rat vocal fold as a model to investigate laryngeal neuromuscular mechanisms.

This protocol describes methods used to prepare rat vocal folds for histochemical neuromuscular study.

用于神经肌肉分析的大鼠声带的制备

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

三维声带振动的半喉设置研究

摘要

探索更多视频

合成，多层，自激振荡的声带模型制作

基于芯片的三维细胞培养，在灌注微生物反应器

研究模型声乐折叠聚合物引起的三维流量分离

一种新型声带生物反应器的构建与鉴定

研究有丝分裂的显微注射技术果蝇合胞体胚胎

非粘性超软人声折叠模型的制造工艺

非粘性超软人声折叠模型的制造工艺

非粘性超软人声折叠模型的制造工艺

膜片钳和灌注技术研究离子通道的表达爪蟾卵母细胞

用于神经肌肉分析的大鼠声带的制备