Name: pupillometry to assess auditory sensation in guinea pigs
Uploaded: 2023-01-06T13:25:00
Description: Watch this Scientific Journal Video about Pupillometry to Assess Auditory Sensation in Guinea Pigs at JoVE.com

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。

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。
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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?...

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1，2，3。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4，5，6，7。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4，5，8，9。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10，11，12，13。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19，20和阿尔茨海默病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22，23，24，25。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27，28，29，30，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30，31，32，33，34，35。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29，36。在听力受损的听众中，例如与年龄相关的听力损失27，30，37，38，39，40，41和人工耳蜗使用者42，43，瞳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27，30，37，38，39，40，41，42，43。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44，45。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47，48。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50，51，52，53，54，55，56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。

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			<td>Analog output board</td>
			<td>Measurement Computing Corporation, Norton, MA</td>
			<td>PCI-DDA02/12</td>
			<td></td>
		</tr>
		<tr>
			<td>Anechoic foam</td>
			<td>Sonex One, Pinta Acoustic, Minneapolis, MN</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Condenser microphone</td>
			<td>Behringer, Willich, Germany</td>
			<td>C-2</td>
			<td></td>
		</tr>
		<tr>
			<td>Free-field microphone</td>
			<td>Bruel &#38; Kjaer, Denmark)</td>
			<td>Type 4940&#160;</td>
			<td></td>
		</tr>
		<tr>
			<td>Matlab</td>
			<td>Mathworks, Inc., Natick, MA</td>
			<td></td>
			<td>2018a version</td>
		</tr>
		<tr>
			<td>Monocular remote camera and illuminator system</td>
			<td>Arrington Research, Scottsdale, AZ</td>
			<td>MCU902</td>
			<td>Infrared LED array + camera with infrared filter</td>
		</tr>
		<tr>
			<td>Multifunction I/O Device</td>
			<td>National Instruments, Austin, TX</td>
			<td>PCI-6229</td>
			<td></td>
		</tr>
		<tr>
			<td>Neural interface processor</td>
			<td>Ripple Neuro, Salt Lake City, UT</td>
			<td>SCOUT</td>
			<td></td>
		</tr>
		<tr>
			<td>Piezoelectric motion sensor</td>
			<td>SparkFun Electronics, Niwot, CO</td>
			<td>SEN-10293</td>
			<td></td>
		</tr>
		<tr>
			<td>Pinch valve</td>
			<td>Cole-Palmer Instrument Co., Vernon Hills, IL</td>
			<td>EW98302-02</td>
			<td></td>
		</tr>
		<tr>
			<td>Programmable attenuator</td>
			<td>Tucker-Davis Technologies, Alachua, FL</td>
			<td>PA5</td>
			<td></td>
		</tr>
		<tr>
			<td>Silicon Tubing</td>
			<td>Cole-Parmer</td>
			<td></td>
			<td>~3 mm</td>
		</tr>
		<tr>
			<td>Sound attenuating chamber</td>
			<td>IAC Acoustics</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Speaker full-range driver</td>
			<td>Tang Band Speaker, Taipei, Taiwan</td>
			<td>W4-1879</td>
			<td></td>
		</tr>
		<tr>
			<td>Stereo Amplifier</td>
			<td>Tucker-Davis Technologies, Alachua, FL</td>
			<td>SA1</td>
			<td></td>
		</tr>
		<tr>
			<td>Tabletop - CleanTop Optical</td>
			<td>TMC vibration control / Ametek, Peabody, MA</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Viewpoint software</td>
			<td>ViewPoint, Arrington Research, Scottsdale, AZ</td>
			<td></td>
			<td></td>
		</tr>
	</tbody>
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pupillometry to assess auditory sensation in guinea pigs

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。

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...

Watch this Scientific Journal Video about Pupillometry to Assess Auditory Sensation in Guinea Pigs at JoVE.com

Pupillometry to Assess Auditory Sensation in Guinea Pigs

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。

评估豚鼠听觉的瞳孔测量法

Noise exposure is a leading cause of sensorineural hearing loss. Animal models of noise-induced hearing loss have generated mechanistic insight into the ...

瞳孔测量法可用于估计复杂的声音识别阈值，并与其他方法结合使用，以确定各种形式的听力损失对这些阈值的影响。该技术可用于表征来自人类和未经训练的动物的相同模态的数据。它相对容易设置和微创。在听力损失的动物模型中很难获得行为和感知测量。这种方法是量化动物复杂声音识别行为的一种方法。正如我们做所有动物行为实验一样，耐心是关键。花点时间让动物适应实验设置。保持简短的实验过程并密切监视动物。首先，将校准的扬声器安装在声音衰减室壁上，高度与放置动物的位置相同。对于自由场刺激传递，将动物放在围栏内，确保不可能进行大的身体运动，并将动物的头部固定在刚性框架上。在外壳下方放置一个压电传感器，以检测和记录动物的运动。然后将瞳孔成像相机放置在距离受试者眼睛 25 厘米的地方。要设置气吹，请使用连接到桌面的支架将移液器吸头放置在动物鼻子前方约 15 厘米处。将直径约三毫米的硅胶管连接到移液器吸头，并将管连接到调节气缸。将气缸气压保持在 20 到 25 psi 之间。并使用计算机控制的继电器将管子通过夹管阀以控制气吹的时间和持续时间。使用放置在约 10 厘米距离处的红外 LED 阵列照亮眼睛。使用强度约为每平方米 2000 坎德拉的白色 LED 照明来照亮成像的眼睛，并使基线瞳孔直径达到约 3.5 毫米。打开瞳孔采集软件，使用带有16毫米镜头的相机获取瞳孔的视频，该镜头的空间分辨率为0.15度视角，红外滤光片放置在距成像眼睛25厘米的距离处。确保眼睛在成像区域中居中。调节相机的光圈和焦点以及红外电平，直到成像瞳孔的轮廓清晰对焦。在瞳孔采集软件中，通过使用鼠标选择一个矩形区域来定义包含瞳孔的感兴趣区域。然后使用控制面板调整采集视频的亮度和对比度。将扫描密度设置为 5 并调整阈值，使椭圆拟合与视频中瞳孔的轮廓紧密匹配。使用神经接口处理器软件，获取并保存来自瞳孔直径或PD迹线的模拟信号，来自记录运动的压电传感器的电压迹线，刺激传递时间和空气吹传递时间。从两种不同类别的发声中选择八种不同长度的豚鼠发声样本。例如，呜呜叫声和呜呜声。一类将作为标准刺激，另一类将作为古怪或异常刺激。要生成一秒钟长的标准和嵌入在不同信噪比或SNR水平的噪声中的偏差刺激，请在呼叫中添加等长的白噪声。本实验中采样的信噪比范围在负24到正40分贝之间。对于每个会话，准备一个伪随机刺激演示序列，其中包含大于 90% 时间的标准刺激。确保在偏差刺激之间至少有 20 到 40 次标准刺激试验。对所有刺激演示使用固定的刺激强度。以高时间规律呈现刺激。为了保持动物与刺激的接触并尽量减少习惯，可以选择在异常刺激后进行短暂的吹气。确保气吹的开始与刺激持续时间充分分离，以便刺激诱发瞳孔扩张反应。在吹气引起眨眼伪影之前达到峰值。运行代码pupil_avg_JOVE_m，并在弹出对话框中从单个会话中选择数据文件，以对每个会话执行运动检测和试用排除。然后运行相同的代码并在弹出对话框中选择要分析的所有数据文件。要消除眨眼伪影，请预处理数据，并获得跨会话中每个刺激的平均瞳孔散大。平均每个刺激条件在每个动物内和动物之间的会话中每个刺激条件引起的瞳孔直径变化，以生成对每个刺激条件的平均瞳孔扩张反应。垂直连接代码pupil_avg_JOVE的所有输出。m或所有会话动物，信噪比和衰减，以构建包含列，动物ID，信噪比，声级和瞳孔直径值的矩阵。对于每个SNR，绘制与截距相对应的权重以可视化结果。对线性和二次项执行相同的操作。将所有具有显着瞳孔变化的试验的会话百分比放入细胞阵列的每个细胞中，其中细胞从低到高SNR排列。使用代码pupil_threshold_estimate_JOVE_m，估计噪声分类阈值中的调用。该图显示了三只动物的平均瞳孔反应。平均瞳孔对标准呜呜声刺激的反应用蓝线表示，阴影对应于正负一个平均值的标准误差。灰线和阴影对应于由异常惠克刺激引起的瞳孔反应平均值的平均值和正负一个标准误差。灰色阴影强度对应于信噪比。绿线和阴影对应于阈值SNR下的平均瞳孔迹线。红色垂直线对应于刺激开始。橙色垂直线对应于气吹开始，蓝绿色虚线对应于 GCA 窗口。心理测量功能符合由偏差刺激引起的显着瞳孔直径变化的试验的百分比，作为SNR的函数。晶须对应于正负一个平均值标准误差。请注意，在大约负 20 分贝 SNR 时达到最大值的 50%。获得的高质量数据，重要的是使动物很好地适应实验设置并保持恒定的消除条件。保持实验时间短，以尽量减少动物对异常刺激的栖息。脑电图或电生理记录与瞳孔测量相结合，将产生对神经缺陷的额外见解，这是听力受损动物的潜在噪音分类缺陷。

pupillometry-to-assess-auditory-sensation-in-guinea-pigs

Research

JoVE Journal

Neuroscience

Operant Sensation Seeking in the Mouse

Operant methods are powerful behavioral tools for the study of motivated behavior. These 'self-administration' methods have been used extensively in drug addiction research due to their high construct validity. Operant studies provide researchers a tool for preclinical investigation of several aspects of the addiction process. For example, mechanisms of acute reinforcement (both drug and non-drug) can be tested using pharmacological or genetic tools to determine the ability of a molecular target to influence self-administration behavior1-6. Additionally, drug or food seeking behaviors can be studied in the absence of the primary reinforcer, and the ability of pharmacological compounds to disrupt this process is a preclinical model for discovery of molecular targets and compounds that may be useful for the treatment of addiction3,7-9. One problem with performing intravenous drug self-administration studies in the mouse is the technical difficulty of maintaining catheter patency. Attrition rates in these experiments are high and can reach 40% or higher10-15. Another general problem with drug self-administration is discerning which pharmacologically-induced effects of the reinforcer produce specific behaviors. For example, measurement of the reinforcing and neurological effects of psychostimulants can be confounded by their psychomotor effects. Operant methods using food reinforcement can avoid these pitfalls, although their utility in studying drug addiction is limited by the fact that some manipulations that alter drug self-administration have a minimal impact on food self-administration. For example, mesolimbic dopamine lesion or knockout of the D1 dopamine receptor reduce cocaine self-administration without having a significant impact on food self-administration 12,16. 
Sensory stimuli have been described for their ability to support operant responding as primary reinforcers (i.e. not conditioned reinforcers)17-22. Auditory and visual stimuli are self-administered by several species18,21,23, although surprisingly little is known about the neural mechanisms underlying this reinforcement. The operant sensation seeking (OSS) model is a robust model for obtaining sensory self-administration in the mouse, allowing the study of neural mechanisms important in sensory reinforcement24. An additional advantage of OSS is the ability to screen mutant mice for differences in operant behavior that may be relevant to addiction. We have reported that dopamine D1 receptor knockout mice, previously shown to be deficient in psychostimulant self-administration, also fail to acquire OSS24. This is a unique finding in that these mice are capable of learning an operant task when food is used as a reinforcer. While operant studies using food reinforcement can be useful in the study of general motivated behavior and the mechanisms underlying food reinforcement, as mentioned above, these studies are limited in their application to studying molecular mechanisms of drug addiction. Thus, there may be similar neural substrates mediating sensory and psychostimulant reinforcement that are distinct from food reinforcement, which would make OSS a particularly attractive model for the study of drug addiction processes. The degree of overlap between other molecular targets of OSS and drug reinforcers is unclear, but is a topic that we are currently pursuing. While some aspects of addiction such as resistance to extinction may be observed with OSS, we have found that escalation 25 is not observed in this model24. Interestingly, escalation of intake and some other aspects of addiction are observed with self-administration of sucrose26. Thus, when non-drug operant procedures are desired to study addiction-related processes, food or sensory reinforcers can be chosen to best fit the particular question being asked. 
In conclusion, both food self-administration and OSS in the mouse have the advantage of not requiring an intravenous catheter, which allows a higher throughput means to study the effects of pharmacological or genetic manipulation of neural targets involved in motivation. While operant testing using food as a reinforcer is particularly useful in the study of the regulation of food intake, OSS is particularly apt for studying reinforcement mechanisms of sensory stimuli and may have broad applicability to novelty seeking and addiction.

In this protocol we describe a method of operant learning using sensory stimuli as a reinforcer in the mouse. It requires no prior training or food restriction, and it allows the study of motivated behavior without the use of a pharmacological or natural reinforcer such as food.

在鼠标的操作性感觉寻求

Operant methods are powerful behavioral tools for the study of motivated behavior.  These 'self-administration' methods have been used extensively in drug ...

科研

神经科学

Preparation and Culture of Chicken Auditory Brainstem Slices

The chicken auditory brainstem is a well-established model system that has been widely used to study the anatomy and physiology of auditory processing at discreet periods of development 1-4 as well as mechanisms for temporal coding in the central nervous system 5-7.
Here we present a method to prepare chicken auditory brainstem slices that can be used for acute experimental procedures or to culture organotypic slices for long-term experimental manipulations.
The chicken auditory brainstem is composed of nucleus angularis, magnocellularis, laminaris and superior olive. These nuclei are responsible for binaural sound processing and single coronal slice preparations preserve the entire circuitry. Ultimately, organotypic slice cultures can provide the opportunity to manipulate several developmental parameters such as synaptic activity, expression of pre and postsynaptic components, expression of aspects controlling excitability and differential gene expression
This approach can be used to broaden general knowledge about neural circuit development, refinement and maturation.

The chicken auditory brainstem is comprised of nuclei responsible for binaural sound processing. A single coronal slice preparation maintains the entire circuitry while the cultured approach provides a unique preparation to study the development of neuronal structure and auditory function at the molecular, cellular and network levels.

鸡脑干切片的制备及文化

The chicken auditory brainstem is a well-established model system that has been widely used to study the anatomy and physiology of auditory processing at ...

A Lightweight, Headphones-based System for Manipulating Auditory Feedback in Songbirds

Experimental manipulations of sensory feedback during complex behavior have provided valuable insights into the computations underlying motor control and sensorimotor plasticity1. Consistent sensory perturbations result in compensatory changes in motor output, reflecting changes in feedforward motor control that reduce the experienced feedback error. By quantifying how different sensory feedback errors affect human behavior, prior studies have explored how visual signals are used to recalibrate arm movements2,3 and auditory feedback is used to modify speech production4-7. The strength of this approach rests on the ability to mimic naturalistic errors in behavior, allowing the experimenter to observe how experienced errors in production are used to recalibrate motor output.
Songbirds provide an excellent animal model for investigating the neural basis of sensorimotor control and plasticity8,9. The songbird brain provides a well-defined circuit in which the areas necessary for song learning are spatially separated from those required for song production, and neural recording and lesion studies have made significant advances in understanding how different brain areas contribute to vocal behavior9-12. However, the lack of a naturalistic error-correction paradigm - in which a known acoustic parameter is perturbed by the experimenter and then corrected by the songbird - has made it difficult to understand the computations underlying vocal learning or how different elements of the neural circuit contribute to the correction of vocal errors13.
The technique described here gives the experimenter precise control over auditory feedback errors in singing birds, allowing the introduction of arbitrary sensory errors that can be used to drive vocal learning. Online sound-processing equipment is used to introduce a known perturbation to the acoustics of song, and a miniaturized headphones apparatus is used to replace a songbird's natural auditory feedback with the perturbed signal in real time. We have used this paradigm to perturb the fundamental frequency (pitch) of auditory feedback in adult songbirds, providing the first demonstration that adult birds maintain vocal performance using error correction14. The present protocol can be used to implement a wide range of sensory feedback perturbations (including but not limited to pitch shifts) to investigate the computational and neurophysiological basis of vocal learning.

We describe the design and assembly of miniaturized headphones suitable for replacing a songbird&#x2019;s natural auditory feedback with a manipulated acoustic signal. Online sound processing hardware is used to manipulate song output, introduce real-time errors in auditory feedback via the headphones, and drive vocal motor learning.

一个轻量级的，耳机为基础的系统操作鸣禽听觉反馈

Experimental manipulations of sensory feedback during complex behavior have provided valuable insights into the computations underlying motor control and ...

Selective Tracing of Auditory Fibers in the Avian Embryonic Vestibulocochlear Nerve

The embryonic chick is a widely used model for the study of peripheral and central ganglion cell projections. In the auditory system, selective labeling of auditory axons within the VIIIth cranial nerve would enhance the study of central auditory circuit development. This approach is challenging because multiple sensory organs of the inner ear contribute to the VIIIth nerve 1. Moreover, markers that reliably distinguish auditory versus vestibular groups of axons within the avian VIIIth nerve have yet to be identified. Auditory and vestibular pathways cannot be distinguished functionally in early embryos, as sensory-evoked responses are not present before the circuits are formed. Centrally projecting VIIIth nerve axons have been traced in some studies, but auditory axon labeling was accompanied by labeling from other VIIIth nerve components 2,3. Here, we describe a method for anterograde tracing from the acoustic ganglion to selectively label auditory axons within the developing VIIIth nerve. First, after partial dissection of the anterior cephalic region of an 8-day chick embryo immersed in oxygenated artificial cerebrospinal fluid, the cochlear duct is identified by anatomical landmarks. Next, a fine pulled glass micropipette is positioned to inject a small amount of rhodamine dextran amine into the duct and adjacent deep region where the acoustic ganglion cells are located. Within thirty minutes following the injection, auditory axons are traced centrally into the hindbrain and can later be visualized following histologic preparation. This method provides a useful tool for developmental studies of peripheral to central auditory circuit formation.

Here we describe a microdissection technique followed by fluorescent dye injection into the acoustic ganglion of early chick embryos for selective tracing of auditory axon fibers in the nerve and hindbrain.

禽流感胚胎听神经听觉神经纤维的选择性跟踪

The embryonic chick is a widely used model for the study of peripheral and central ganglion cell projections. In the auditory system, selective labeling ...

Functional Imaging of Auditory Cortex in Adult Cats using High-field fMRI

Current knowledge of sensory processing in the mammalian auditory system is mainly derived from electrophysiological studies in a variety of animal models, including monkeys, ferrets, bats, rodents, and cats. In order to draw suitable parallels between human and animal models of auditory function, it is important to establish a bridge between human functional imaging studies and animal electrophysiological studies. Functional magnetic resonance imaging (fMRI) is an established, minimally invasive method of measuring broad patterns of hemodynamic activity across different regions of the cerebral cortex. This technique is widely used to probe sensory function in the human brain, is a useful tool in linking studies of auditory processing in both humans and animals and has been successfully used to investigate auditory function in monkeys and rodents. The following protocol describes an experimental procedure for investigating auditory function in anesthetized adult cats by measuring stimulus-evoked hemodynamic changes in auditory cortex using fMRI. This method facilitates comparison of the hemodynamic responses across different models of auditory function thus leading to a better understanding of species-independent features of the mammalian auditory cortex.

Functional studies of the auditory system in mammals have traditionally been conducted using spatially-focused techniques such as electrophysiological recordings. The following protocol describes a method of visualizing large-scale patterns of evoked hemodynamic activity in the cat auditory cortex using functional magnetic resonance imaging.

听觉皮层在成年猫用高场磁共振成像功能成像

Current knowledge of sensory processing in the mammalian auditory system is mainly derived from electrophysiological studies in a variety of animal ...

Optogenetic Stimulation of the Auditory Nerve

Direct electrical stimulation of spiral ganglion neurons (SGNs) by cochlear implants (CIs) enables open speech comprehension in the majority of implanted deaf subjects1-6. Nonetheless, sound coding with current CIs has poor frequency and intensity resolution due to broad current spread from each electrode contact activating a large number of SGNs along the tonotopic axis of the cochlea7-9. Optical stimulation is proposed as an alternative to electrical stimulation that promises spatially more confined activation of SGNs and, hence, higher frequency resolution of coding. In recent years, direct infrared illumination of&#160;the cochlea has been used to evoke responses in the auditory nerve10. Nevertheless it requires higher energies than electrical stimulation10,11 and uncertainty remains as to the underlying mechanism12. Here we describe a method based on optogenetics to stimulate SGNs with low intensity blue light, using transgenic mice with neuronal expression of channelrhodopsin 2 (ChR2)13 or virus-mediated expression of the ChR2-variant CatCh14. We used micro-light emitting diodes (&#181;LEDs) and fiber-coupled lasers to stimulate ChR2-expressing SGNs through a small artificial opening (cochleostomy) or the round window. We assayed the responses by scalp recordings of light-evoked potentials (optogenetic auditory brainstem response: oABR) or by microelectrode recordings from the auditory pathway and compared them with acoustic and electrical stimulation.

Cochlear implants (CIs) enable hearing by direct electrical stimulation of the auditory nerve. However, poor frequency and intensity resolution limits the quality of hearing with CIs. Here we describe optogenetic stimulation of the auditory nerve in mice as an alternative strategy for auditory research and developing future CIs.

听觉神经的光遗传学刺激

Direct electrical stimulation of spiral ganglion neurons (SGNs) by cochlear implants (CIs) enables open speech comprehension in the majority of implanted ...

The Use of Trace Eyeblink Classical Conditioning to Assess Hippocampal Dysfunction in a Rat Model of Fetal Alcohol Spectrum Disorders

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain undergoes rapid organizational change and is similar to accelerated brain changes that occur during the third trimester in humans. This model of fetal alcohol spectrum disorders (FASDs) produces severe brain damage, mimicking the amount and pattern of binge-drinking that occurs in some pregnant alcoholic mothers. We describe the use of trace eyeblink classical conditioning (ECC), a higher-order variant of associative learning, to assess long-term hippocampal dysfunction that is typically seen in alcohol-exposed adult offspring. At 90 days of age, rodents were surgically prepared with recording and stimulating electrodes, which measured electromyographic (EMG) blink activity from the left eyelid muscle and delivered mild shock posterior to the left eye, respectively. After a 5 day recovery period, they underwent 6 sessions of trace ECC to determine associative learning differences between alcohol-exposed and control rats. Trace ECC is one of many possible ECC procedures that can be easily modified using the same equipment and software, so that different neural systems can be assessed. ECC procedures in general, can be used as diagnostic tools for detecting neural pathology in different brain systems and different conditions that insult the brain.

Trace eyeblink classical conditioning (ECC) was used to assess hippocampal-dependent associative learning in adult rats that were administered a high concentration (11.9% v/v) of alcohol during early neonatal brain development. In general, ECC procedures are sound diagnostic tools for detecting brain dysfunction across many psychological and biomedical settings.

使用的跟踪经典条件反射，评估胎儿酒精谱系障碍大鼠模型海马功能障碍

Neonatal rats were administered a relatively high concentration of ethyl alcohol (11.9% v/v) during postnatal days 4-9, a time when the fetal brain ...

A Preclinical Model to Assess Brain Recovery After Acute Stroke in Rats

The purpose of this study was to establish and validate an animal brain ischemia model in the recovery and sequela stages. A middle cerebral artery occlusion/reperfusion (MCAO/R) model in male Sprague-Dawley rats was chosen. By changing the rat&#39;s weight (260&#8722;330 g), the thread bolt type (2636/2838/3040/3043) and the brain infarct time (2-3 h), a higher Longa&#39;s score, a larger infarct volume and a greater model success ratio were screened using the Longa&#39;s score and TTC staining. The optimum model condition (300 g, 3040 thread bolt, 3 h brain infarct time) was acquired and used in a 1-90 day observation period after reperfusion via assessment of sensorimotor functions and infarct volume. At these conditions, the bilateral asymmetry test had a significant difference from 1 to 90 days, and the grid-walking test had a significant difference from 1 to 60 days; both differences could be a suitable sensorimotor functional test. Thus, the most appropriate condition of a novel rat model in the recovery and sequela stages of brain ischemia was found: 300 g rats that underwent MCAO with a 3040 thread bolt for a 3 h brain infarct and then reperfused. The appropriate sensorimotor functional tests were a bilateral asymmetry test and a grid-walking test.

The&#160;purpose of this study is to establish and validate an animal model for research in the recovery and sequela stages of brain ischemia by testing brain infarction and sensorimotor function after middle cerebral artery occlusion/reperfusion (MCAO/R) after 1-90 days in rats.

评估大鼠急性中风后脑恢复的临床前模型

The purpose of this study was to establish and validate an animal brain ischemia model in the recovery and sequela stages. A middle cerebral artery ...

A Revised Surgical Approach to Induce Endolymphatic Hydrops in the Guinea Pig

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) remain unclear. In order to adequately study the attributes of endolymphatic hydrops, such as the origins of low-frequency hearing loss, a reliable model is needed. The guinea pig is a&#160;good model because it hears in the low-frequency regions that are putatively affected by endolymphatic hydrops. Previous research has demonstrated that endolymphatic hydrops can be induced surgically via intradural or extradural approaches that involve drilling on the endolymphatic duct and sac. However, whether it was possible to create an endolymphatic hydrops model using an extradural approach that avoided dangerous drilling on the endolymphatic duct and sac was unknown. The objective of this study was to demonstrate a revised extradural approach to induce experimental endolymphatic hydrops at 30 days post-operatively by obliterating the endolymphatic sac and injuring the endolymphatic duct with a fine pick. The sample size consisted of seven guinea pigs. Functional measurements of hearing were made and temporal bones were subsequently harvested for histologic analysis. The approach had a success rate of 86% in achieving endolymphatic hydrops. The risk of cerebrospinal fluid leak was minimal. No perioperative deaths or injuries to the posterior semicircular canal occurred in the sample. The presented method demonstrates a safe and reliable way to induce endolymphatic hydrops at a relatively quick time point of 30 days. The clinical implications are that the presented method provides a reliable model to further explore the origins of low-frequency hearing loss that can be associated endolymphatic hydrops.

This article demonstrates an extradural approach to obliterate the guinea pig endolymphatic sac and injure the endolymphatic duct with a fine pick in order to induce experimental endolymphatic hydrops.

几内亚猪诱导内部水解的修正手术方法

Endolymphatic hydrops is an enlargement of scala media that is most often associated with Meniere&#39;s disease, though the pathophysiologic mechanism(s) ...

Direct Cannula Implantation in the Cisterna Magna of Pigs

The glymphatic system is a waste clearance system in the brain that relies on the flow of cerebrospinal fluid (CSF) in astrocyte-bound perivascular spaces and has been implicated in the clearance of neurotoxic peptides such as amyloid-beta. Impaired glymphatic function exacerbates disease pathology in animal models of neurodegenerative diseases, such as Alzheimer's, which highlights the importance of understanding this clearance system. The glymphatic system is often studied by cisterna magna cannulations (CMc), where tracers are delivered directly into the cerebrospinal fluid (CSF). Most studies, however, have been carried out in rodents. Here, we demonstrate an adaptation of the CMc technique in pigs. Using CMc in pigs, the glymphatic system can be studied at a high optical resolution in gyrencephalic brains and in doing so bridges the knowledge gap between rodent and human glymphatics.

This article presents a step-by-step protocol for the direct cannula implantation in the cisterna magna of pigs.

直接插管植入猪的蓄水池

The glymphatic system is a waste clearance system in the brain that relies on the flow of cerebrospinal fluid (CSF) in astrocyte-bound perivascular spaces ...