Summary

评估豚鼠听觉的瞳孔测量法

Published: January 06, 2023
doi:

Summary

瞳孔测量是一种简单且非侵入性的技术,被提出作为确定正常听力动物和各种听觉病理的动物模型中的噪声听觉阈值的方法。

Abstract

噪音暴露是感音神经性听力损失的主要原因。噪声引起的听力损失的动物模型已经对听力损失的潜在解剖学和生理病理学产生了机制见解。然而,将听力损失人类中观察到的行为缺陷与动物模型中的行为缺陷联系起来仍然具有挑战性。在这里,瞳孔测量法被提出作为一种能够直接比较动物和人类行为数据的方法。该方法基于一种改进的古怪范式 – 使受试者习惯于刺激的重复呈现,并间歇性地呈现与重复刺激以某种参数方式变化的偏差刺激。基本前提是,如果受试者检测到重复刺激和偏差刺激之间的变化,它将触发比重复刺激引起的更大的瞳孔扩张反应。这种方法在豚鼠中使用发声分类任务进行了演示,豚鼠是一种广泛用于听觉研究(包括听力损失研究)的动物模型。通过将一个发声类别的发声呈现为标准刺激,将另一个类别的发声呈现为嵌入不同信噪比的噪声中的古怪刺激,证明了响应古怪类别的瞳孔扩张幅度随信噪比单调变化。然后可以使用生长曲线分析来表征这些瞳孔扩张反应的时间过程和统计学意义。在该协议中,描述了使豚鼠适应设置,进行瞳孔测量和评估/分析数据的详细程序。尽管该技术在本协议中在正常听力的豚鼠中得到了证明,但该方法可用于评估每个受试者内各种形式的听力损失的感觉影响。然后,这些影响可能与同时进行的电生理测量和事后解剖观察相关联。

Introduction

瞳孔直径(PD)可能受到多种因素的影响,随时间变化的PD测量称为瞳孔测量法。PD由虹膜括约肌(参与收缩)和虹膜扩张肌(参与扩张)控制。收缩肌由副交感神经系统支配并涉及胆碱能投射,而虹膜扩张肌由涉及去甲肾上腺素能和胆碱能投射的交感神经系统支配123诱导PD变化的最着名的刺激是亮度收缩,瞳孔的扩张反应可以通过环境光强度的变化产生2。PD也随焦距2而变化。然而,几十年来,人们已经知道PD也显示出与亮度无关的波动4567例如,内部精神状态的变化可以引起短暂的PD变化。瞳孔因情绪刺激而扩张,或随着唤醒而增加4589瞳孔扩张也可能与其他认知机制有关,例如增加脑力劳动或注意力10111213由于瞳孔大小变化与精神状态之间的这种关系,PD变化已被探索为临床疾病的标志,例如精神分裂症14,15,焦虑症16,17,18,帕金森病1920和阿尔茨海默21等。在动物中,PD变化跟踪内部行为状态,并与皮质区域的神经元活动水平相关22232425。瞳孔直径也被证明是小鼠睡眠状态的可靠指标26。这些与唤醒和内部状态相关的PD变化通常发生在几十秒量级的长时间尺度上。

在听力研究领域,在正常听力和听力受损的受试者中,已经使用瞳孔测量法评估了听力努力和听觉感知。这些研究通常涉及训练有素的研究对象27282930,他们执行各种检测或识别任务。由于上述唤醒和PD之间的关系,增加的任务参与度和倾听努力已被证明与瞳孔扩张反应增加相关3031,3233,3435因此,瞳孔测量法已被用于证明增加的听力努力用于识别正常听力听众的频谱退化语音2936。在听力受损的听众中,例如与年龄相关的听力损失27,3037,38394041和人工耳蜗使用者42,43孔反应也随着语音清晰度的降低而增加;然而,与正常听力受试者相比,听力受损的听众在更容易的听力条件下表现出更大的瞳孔扩张27,3037383940414243但是,要求听众执行识别任务的实验并不总是可能的 – 例如,在婴儿或某些动物模型中。因此,在这些情况下,由声学刺激引起的非亮度相关瞳孔反应可能是评估听觉检测的可行替代方法4445。早期的研究表明,短暂和刺激相关的瞳孔扩张是定向反射的一部分46。后来的研究表明,使用刺激相关的瞳孔扩张来推导出猫头鹰的频率敏感性曲线4748。最近,这些方法已被应用于评估人类婴儿瞳孔扩张反应的敏感性48。瞳孔测量已被证明是一种可靠且非侵入性的方法,通过使用各种简单(音调)和复杂(GP发声)刺激来估计被动聆听豚鼠(GP)的听觉检测和辨别阈值49。这些与刺激相关的PD变化通常发生在几秒钟的较快时间尺度上,并与刺激时间有关。本文提出了刺激相关PD变化的瞳孔测量法,作为研究动物模型中各种听力障碍行为影响的方法。特别是,描述了用于GPs的瞳孔测量方案,一种各种类型的听觉病理学50,5152,53,54,5556的成熟动物模型(另见参考文献57以获得详尽的综述)。

虽然这种技术在正常听力全科医生中得到了证明,但这些方法可以很容易地适应其他动物模型和各种听觉病理的动物模型。重要的是,瞳孔测量法可以与其他非侵入性测量(如脑电图)以及侵入性电生理记录相结合,以研究可能的声音检测和感知缺陷的机制。最后,这种方法也可用于建立人类和动物模型之间的广泛相似性。

Protocol

对于所有实验程序,请获得机构动物护理和使用委员会(IACUC)的批准,并遵守NIH关于实验动物护理和使用指南。在美利坚合众国,全科医生还受美国农业部 (USDA) 法规的约束。该协议中的所有程序均已获得匹兹堡大学IACUC的批准,并遵守NIH关于实验动物护理和使用指南。在本实验中,使用了三个雄性野生型,色素沉着的GPs,年龄在4至10个月之间,体重~600-1,000克。 1. ?…

Representative Results

在三个男性色素GP中进行瞳孔测定,在实验过程中重量~600-1,000g。如该协议所述,为了估计呼叫噪声分类阈值,使用了一种奇怪的范式来表示刺激。在古怪的范式中,属于给定SNR下嵌入白噪声中的一类呼叫(呜呜声)被用作标准刺激(图2A),而来自同一SNR嵌入白噪声中的另一类(wheeks)的呼叫(图2A)作为偏差刺激。标准和偏差刺激从每个类别的八个…

Discussion

该协议展示了使用瞳孔测量法作为非侵入性和可靠的方法来估计被动聆听动物的听觉阈值。按照此处描述的协议,估计正常听力GP中的呼叫噪声分类阈值。发现使用瞳孔测量法估计的阈值与使用操作性训练获得的阈值一致62。然而,与操作性训练相比,瞳孔测量方案相对简单,可以快速设置和获取数据。每次数据采集会话(每个SNR水平)持续约12分钟,导致每只动物每天1-2小时的?…

Disclosures

The authors have nothing to disclose.

Acknowledgements

这项工作得到了NIH(R01DC017141),宾夕法尼亚狮子会听力研究基金会以及匹兹堡大学耳鼻喉科和神经生物学系的资助。

Materials

Analog output board Measurement Computing Corporation, Norton, MA PCI-DDA02/12
Anechoic foam Sonex One, Pinta Acoustic, Minneapolis, MN
Condenser microphone Behringer, Willich, Germany C-2
Free-field microphone Bruel & Kjaer, Denmark) Type 4940 
Matlab Mathworks, Inc., Natick, MA 2018a version
Monocular remote camera and illuminator system Arrington Research, Scottsdale, AZ MCU902 Infrared LED array + camera with infrared filter
Multifunction I/O Device National Instruments, Austin, TX PCI-6229
Neural interface processor Ripple Neuro, Salt Lake City, UT SCOUT
Piezoelectric motion sensor SparkFun Electronics, Niwot, CO SEN-10293
Pinch valve Cole-Palmer Instrument Co., Vernon Hills, IL EW98302-02
Programmable attenuator Tucker-Davis Technologies, Alachua, FL PA5
Silicon Tubing Cole-Parmer ~3 mm
Sound attenuating chamber IAC Acoustics
Speaker full-range driver Tang Band Speaker, Taipei, Taiwan W4-1879
Stereo Amplifier Tucker-Davis Technologies, Alachua, FL SA1
Tabletop – CleanTop Optical TMC vibration control / Ametek, Peabody, MA
Viewpoint software ViewPoint, Arrington Research, Scottsdale, AZ

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Cite This Article
Pernia, M., Kar, M., Montes-Lourido, P., Sadagopan, S. Pupillometry to Assess Auditory Sensation in Guinea Pigs. J. Vis. Exp. (191), e64581, doi:10.3791/64581 (2023).

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