使用大鼠的耳镜可视化和行为拍片启动反应进行验证，对传导性听力损失进行简单手术诱导
1Instituto de Neurobiología,Universidad Nacional Autónoma de México, México, 2State Key Laboratory of Marine Pollution (SKLMP),City University of Hong Kong, Hong Kong, 3Department of Biomedical Sciences, College of Veterinary Medicine and Life Sciences,City University of Hong Kong, Hong Kong, 4Department of Physics,City University of Hong Kong, Hong Kong, 5Department of Clinical Oncology,Queen Elizabeth Hospital, Hong Kong, 6Department of Materials Science and Engineering,City University of Hong Kong, Hong Kong
Summary
在这里，我们提出了一个协议，通过手术性鼓膜穿刺建立可复制的传导性听力损失诱导，并通过耳镜可视化和行为评估通过拍拍惊吓进行验证。
Abstract
传导性听力损失 （CHL） 是人体常见的听力损伤。该协议的目的是描述一个简单的外科手术，诱导啮齿动物的CHL。该协议通过鼓膜穿刺演示CHL。CHL手术的验证是通过耳镜检查和行为评估，通过拍拍的启动反应，既可复制又可靠，是证明听力损失发生的简单方法。简单的CHL程序具有优越的优越性，因为其可重复性和灵活性对听力损失研究中的不同追求。通过手术方法诱导CHL的局限性与执行外科手术的学习曲线和对听力检查的信心有关。由CHL诱导听力损伤，使人们能够随时研究听力损失的神经表现和行为结果。
Introduction
儿童和成人听力损失的患病率分别为19.5%1和15.2%。然而，大约39.3%的新生儿有异常的听力筛查没有得到治疗，如疾病控制中心报告3。听力损失是一种被广泛研究的疾病，啮齿动物是研究正常听力和听力相关疾病4、5、6、7、8、9的有力模型 ，10，11，12，13，14，15。听力障碍，如传导性听力损失（CHL），导致听觉皮层4的短期突触抑郁症增加，导致与频率调制检测阈值相关的较浅的心理测量斜坡5.通过手术去除/置换的胸膜、鼓膜 （TM） 穿刺或耳塞的传导性听力损失模型易于使用，并允许快速诱导听力损失模型5，14 ，15，16，17，18.本协议和方法的目的是在啮齿类动物中演示一种简单且可重复的CHL模型。
本协议价格低廉（所有工具为300美元），可随时修改，适用于不同的研究领域。大鼠曾对中耳解剖进行了详细评估，其中19、20、21、22、23、手术方法24、中耳炎模型25、 26，27和 TM 穿刺再生16，17，18，28，29，30，使其成为理想的模型研究听力损失。在这里，一个简单的CHL诱导程序描述与耳镜和行为评估验证与拍拍启动反应在大鼠，然后可用于探索听力损失的其他后遗症。CHL 程序由 TM 的手术穿刺诱导。CHL 程序的验证通过耳镜可视化执行，以确定没有 TM。行为评估由高分贝 （dB） 声压水平 （SPL） 手拍执行。该方法以前已应用于各种啮齿动物。它很容易复制，产生强大的心理测量差异和神经生理反应的变化4，5，16，17，18。
Protocol
本研究及程序获香港城市大学、香港大学及香港特别行政区卫生署动物研究伦理委员会批准。 1. 动物 使用两个月的斯普拉格-道利 （SD） 大鼠（N = 90，200-250 g）。注：啮齿动物由香港大学认可化验动物组提供。 在实验室动物研究室保持大鼠在25°C恒定的温度和60-70%的湿度下。 房子啮齿动物在12/12-h光/暗周期与获得食物和饮用水，解放。 <li…
Representative Results
简单的CHL程序对90只大鼠进行，在组2有显性出血，2没有听力损失的第二天评估的行为拍手。这四只老鼠被丢弃了。由于并发症，在讨论中的原因说明，应该丢弃大鼠。诱导TM穿刺和/或马勒鲁斯位移/去除（图2B）导致CHL，导致行为表现（即CHL – 对响亮的dB SPL起动无反应）。大鼠在手术前应接受子宫内窥镜检查，以确保子宫膜正常，术后应确保鼓膜穿刺…
Discussion
我们描述了一个简单的手术诱导CHL与验证使用耳镜可视化和行为拍拍启动反应在大鼠。在这里，我们演示了方法在大鼠和以前这种方法已经应用于鼠和小鼠。该方法可以很容易地被采用到其他啮齿动物。CHL的诱导允许研究一种微妙的听力损失形式，它表现在听觉皮质改变和心理物理行为发现4，5，16，17，18<sup class="xref"…
Disclosures
The authors have nothing to disclose.
Acknowledgements
这项工作部分得到香港研究资助局、早期职业计划、#21201217项目、脑制图引导电生理学项目的支持，并应用于听觉和噪音污染研究。我们感谢墨西哥国立自治大学神经生物研究所、墨西哥国家技术委员会（CONACyT）向墨西哥国家大学提供578458年研究生奖学金。
Materials
| Latex, polyvinyl or nitrile gloves | AMMEX | Use unpowdered gloves 8-mil | |
| Micro spring scissors (see Fig. 1b) | RWD Life Science | S11035-08 | 8.0 cm total length, with 3.5mm cutting edge, or similar micro forceps. Standard tweezers with spring action will suffice |
| Otoscope mini 3000 | HEINE | D-008.70.120M | Standard LED otoscope will suffice |
| Rat or mouse | JAX labs | Any small rodent | |
| Small rodent cage | Tecniplast | 1284L | Need two cages to separate CHL rodent from hearing rodent. If rodent are in direct contact with one-another, they will startle each other. Cage dimensions 365 x 207 x 140 mm, floor area: 530 cm2/82.15 in2 |
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Cite This Article
Manno, F. A. M., Manno, S. H. C., Ma, V., Barrios, F. A., Cho, W. C., Cheng, S. H., Lau, C. Simple Surgical Induction of Conductive Hearing Loss with Verification Using Otoscope Visualization and Behavioral Clap Startle Response in Rat. J. Vis. Exp. (152), e57993, doi:10.3791/57993 (2019).