这项研究的部分原因是奈特校园本科生学者计划和菲尔和佩妮奈特基金会的慷慨资助。

此处详述的所有方法均按照IRB协议和批准（俄勒冈大学IRB协议编号10182017.017）进行。获得所有受试者的知情同意。
1. 伸展装置
<ol><li>将大型图形输入板平放在桌面上。</li>
	<li>使用可调节的 80-20 铝制框架将任务监视器平行放置在平板电脑上方 6-8 处，屏幕朝上（有关蓝图，请查看此处：https://github.com/greenhouselab/Reach_TMS 和补充图 1）。 注意：此设置允许参与者跨过平板电脑并获取任务监视器上显示的目标，同时遮挡其伸展手臂的视野。</li>
	<li>使用Kim等人3 中描述的设置作为参考。</li>
</ol>2. 机器接口
<ol><li>通过 USB 端口 将 平板电脑连接到计算机。通过HDMI端口 将 任务监视器连接到计算机。通过DB-9电缆 将 后面的TMS端口连接到计算机。</li>
	<li>通过PCI-6220卡DAQ 将 EMG系统连接到计算机。通过BNC电缆 将 光电二极管连接到EMG系统。</li>
</ol>3. 光电二极管传感器
<ol><li>将光电二极管传感器连接到 BNC 电缆。用胶带将光电二极管传感器固定在任务监视器的右上角，传感器面向屏幕，≤ 1 厘米远。 注意：这会将任务监视器上呈现的刺激的时序记录为独立输入通道中的模拟数据。</li>
</ol>4. 软件
<ol><li>下载适用于 MATLAB 2018 的 VETA 工具箱26 （https://github.com/greenhouselab/Veta），与用于数据收集的硬件接口。</li>
	<li>下载为控制实验参数和与平板电脑接口而开发的到达任务代码（https://github.com/greenhouselab/Reach_TMS）。</li>
</ol>5. 参与者筛选和知情同意
<ol><li>筛查受试者是否有 TMS 禁忌证。排除标准包括癫痫发作、头痛、脑外伤、昏厥、慢性应激或焦虑、睡眠问题和任何神经活性药物的个人或家族史。其他排除标准包括大脑或颅骨中的任何金属植入物，以及在测试前24小时内使用任何娱乐性药物或酒精。纳入标准包括右撇子和年龄在18至35岁之间。</li>
	<li>提供程序和相关风险的书面解释，澄清参与者可能提出的任何其他问题。</li>
	<li>获得参与者的知情同意。</li>
</ol>6. 科目设置
<ol><li>将拍摄对象放在面向平板电脑的舒适椅子上。确保膝盖弯曲至90°，腿在桌子下方。</li>
	<li>准备皮肤并放置肌电图电极。<ol><li>使用细粒砂纸轻轻擦拭右侧第一背骨间 （FDI）、桡腕伸肌和前三角肌部位的皮肤，以及颈部底部的 C4 突起，以检测 TMS 脉冲产生的电伪影。 注意：肌肉记录站点可以根据用户需求进行调整。</li>
		<li>每个电极部位用酒精制备垫擦拭每个磨损区域一次以进行清洁。</li>
		<li>在每个部位放置一个肌电图电极。确保电极垂直于肌肉纤维运行。将接地电极放在右肘的骨突出处。</li>
		<li>用医用胶带固定每个电极。</li>
	</ol></li>
	<li>检查肌电图记录的质量。使用 VETA 工具箱可视化所有肌电图迹线，并确保它们没有伪影。如果肌电图迹线有噪音，请确保接地正确放置，并且所有电极与皮肤正确接触。</li>
</ol>7. 经颅磁刺激
<ol><li>打开 TMS 机器。</li>
	<li>通过刺激左侧M1 找到 右侧FDI肌肉的TMS热点。<ol><li>将线圈横向放置~5厘米，在头部顶点前2厘米处放置，方向为中线~45°。</li>
		<li>每 4 秒施用一次 TMS 脉冲，同时在前-后和内侧侧以约 5 mm 的增量重新定位线圈。</li>
		<li>从 30% 的最大刺激器输出开始，以 2% 的增量逐渐增加 TMS 强度，直到观察到 MEP。</li>
		<li>一旦确定了最佳位置，在该位置上，MEP 可以以尽可能低的刺激器强度可靠地在大多数 （~75%） 脉冲上被引出，通过找到在 10 个脉冲中的 5 个脉冲上产生峰峰值幅度为 >50 μV 的 MEP 的强度水平来确定静息运动阈值 （RMT）。</li>
		<li>通过沿着线圈的周边在参与者的头上轻轻放置细条反光带来标记位置。通过手动握住线圈或使用支架支撑线圈来保持线圈定位。</li>
	</ol></li>
</ol>8. 到达任务设置
<ol><li>将魔术贴手套戴在参与者的右手上，以促进放松的力量握持姿势。</li>
	<li>将手写笔连接到手套上，并建议受试者在伸手运动之间保持手部放松。</li>
	<li>传达任务说明，如下所示： 将光标引导到屏幕底部的初始位置。您将在两个目标位置之一看到提示。当目标填充颜色时，尽可能快速准确地穿过目标。然后返回初始位置。指示初始位置、提示和目标的位置（图 1A）。</li>
	<li>指导参与者用手写笔尽可能快速准确地切开目标。关闭任务室中的灯，以遮挡参与者手臂运动的视野并提高任务监视器的可见性。</li>
</ol>9. 任务设计
<ol><li>在 Matlab 2018 中使用 Psychtoolbox 3.0 控制视觉刺激呈现（补充编码文件 1）。</li>
	<li>使用以下参数匹配当前数据：20 次练习试验;270次试验;4/5 的测试试验的 TMS;TMS 要么与预备提示开始（基线 TMS）重合，要么在命令性提示（延迟 TMS）之前 100 毫秒以相同的频率重合;总试验的1/10是捕获试验，其中没有出现命令性提示;起始位置是一个半径为 2 厘米的圆，位于工作区的底部中心;两个半径为 1 cm 的圆形目标位于距离起始位置 15 cm 处，距离中线 +45°和 -45°。</li>
	<li>按如下方式设置事件顺序和持续时间：准备提示为 900 毫秒，命令式提示为 900 毫秒。</li>
</ol>10. TMS管理
<ol><li>VETA工具箱同时管理TMS并记录肌电图 https://github.com/greenhouselab/Veta。</li>
	<li>使用VETA工具箱控制TMS脉冲的时间，以与所选的行为事件（即，准备提示的开始或目标开始前100毫秒）一致。</li>
	<li>以足够的频率提供 TMS，以确保有足够的 MEP 进行分析。 注意：如前所述，任务代码将在总试验的 4/5 上提供 TMS 脉冲，要么在准备提示开始时引出基线 MEP，要么在命令提示引发延迟 MEP 之前 100 毫秒。可以根据用户需要在代码中调整参数。没有TMS的试验可用于评估在没有TMS的情况下的行为表现。这对于确定 TMS 对性能的任何可能影响非常有用。</li>
</ol>

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所有作者声明不存在利益冲突

上面概述的方法提供了一种在达到行为的背景下研究运动准备的新方法。尽管到达代表了运动控制和学习研究中的流行模型任务，但需要精确评估与到达行为相关的CS动力学。TMS提供了一种非侵入性的，时间精确的方法来捕获到达过程中离散时间点的CS活动。这里描述的方法结合了两个独立的子领域 - TMS和触及 - 涉及同时记录运动学和电生理指标的单一范式。 
虽?...

目标导向的到达是一种基本的运动行为，允许人类与外部环境互动并操纵外部环境。运动生理学、心理学和神经科学领域的研究产生了丰富而广泛的文献，其中包括各种方法论。早期的接触研究使用非人类灵长类动物的直接神经记录来研究单个神经元水平的神经活动1，2。最近的研究调查了使用行为范式来探索运动学习和控制的本质3，4，5。这种行为任务与功能性磁共振成像和脑电图相结合，可以测量人类到达时的全脑活动6，7。其他研究已应用在线TMS来调查到达准备和执行的各种特征8，9，10，11，12，13，14。然而，仍然需要一种开源和灵活的方法，将覆盖的行为评估与TMS相结合。虽然将TMS与行为协议相结合的效用已经非常成熟15，但在这里，我们专门研究了TMS在使用开源方法的背景下的应用。这是新颖的，因为使用这种方法组合发表文章的其他小组没有使他们的工具随时可用，因此禁止直接复制。这种开源方法促进了复制、数据共享和多中心研究的可能性。此外，如果其他人希望使用类似的工具追求新颖的研究问题，开源代码可以作为创新的发射台，因为它很容易适应。
TMS提供了一种在精确控制的时间点16探测电机系统的无创方法。当应用于初级运动皮层（M1）时，TMS可以在目标肌肉的肌电图中引起可测量的偏转。该电压波的振幅称为电机诱发电位（MEP），提供了皮质脊髓（CS）通路瞬时兴奋状态的指数 - 这是对CS通路17的所有兴奋性和抑制性影响的合成模拟。除了提供可靠的内在CS兴奋性受试者内测量外，TMS还可以与其他行为或运动学指标相结合，以时间精确的方式研究CS活动与行为之间的关系。许多研究已经利用TMS和肌电图（EMG）的组合来解决有关运动系统的各种问题，特别是因为这种方法的组合使得在各种行为条件下研究MEP成为可能15。事实证明，这特别有用的一个领域是动作准备的研究，最常见的是通过研究单关节运动18。然而，对自然主义多关节运动（如伸展）的TMS研究相对较少。
目前的目标是设计一种延迟反应到达任务，包括行为运动学，在线单脉冲TMS管理以及来自多个肌肉的同时肌电图记录。该任务包括一个二维点对点到达范式，使用水平方向的监视器进行在线视觉反馈，使得视觉反馈与到达轨迹相匹配（即，在真实反馈期间的 1：1 关系，视觉反馈和运动之间没有转换）。目前的设计还包括一组视觉运动扰动试验。在提供的示例中，这是光标反馈中的 20° 旋转偏移。以前的研究使用类似的到达范式来解决与感觉运动适应相关的机制和计算的问题19，20，21，22，23，24，25。此外，这种方法可以在在线运动学习期间的精确时间点评估运动系统的兴奋性动态。
由于接触已被证明是研究学习/适应的一种富有成效的行为，因此在这种行为的背景下评估CS兴奋性具有揭示这些行为所涉及的神经基质的巨大潜力。这些可能包括局部抑制影响，调谐特性的变化，神经事件的时间等，正如在非人类灵长类动物研究中已经建立的那样。然而，这些特征在人类和临床人群中更难量化。在人类没有明显运动的情况下，也可以使用TMS和EMG组合方法（即，在准备运动或休息时）研究神经动力学。
所提供的工具是开源的，代码很容易适应。这种新颖的范式将对伸展运动的准备、执行、终止和适应所涉及的机制产生重要的见解。此外，这种方法的组合有可能揭示电生理学与人类到达行为之间的关系。

<table border="1" fo:keep-together.within-page="1" fo:keep-with-next.within-page="always">
	<tbody>
		<tr>
			<td>2-Port Native PCI Express&#160;</td>
			<td>StarTech.com</td>
			<td>RS232 Card with 16950 UART&#160;</td>
			<td>Must be compatible with desktop computer</td>
		</tr>
		<tr>
			<td>Adjustable 80-20 aluminum frame</td>
			<td>any</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Alcohol prep pads</td>
			<td>any</td>
			<td></td>
			<td>EMG preparation</td>
		</tr>
		<tr>
			<td>Bagnoli Bipolar Electrodes</td>
			<td>Delsys</td>
			<td>DE 2.1</td>
			<td></td>
		</tr>
		<tr>
			<td>Bagnoli Reference Electrode</td>
			<td>Delsys</td>
			<td>USX2000</td>
			<td>2&#8221; (5cm) Round</td>
		</tr>
		<tr>
			<td>Bagnoli-8 EMG System</td>
			<td>Delsys</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Chair</td>
			<td>any</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Computer monitor for EMG/TMS</td>
			<td>n/a</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Desk</td>
			<td>any</td>
			<td></td>
			<td></td>
		</tr>
		<tr>
			<td>Desktop Computer</td>
			<td>Dell</td>
			<td>xps 8930</td>
			<td>RAM: 16 GB, Storage: 1TB, Graphics: 1060 6GB&#160;</td>
		</tr>
		<tr>
			<td>EMG electrodes</td>
			<td>Delsys</td>
			<td>Sensor Adhesive Interface</td>
			<td></td>
		</tr>
		<tr>
			<td>Fine grain sandpaper</td>
			<td>any</td>
			<td></td>
			<td>EMG preparation</td>
		</tr>
		<tr>
			<td>Graphics tablet</td>
			<td>Wacom</td>
			<td>Intuos-4 XL</td>
			<td></td>
		</tr>
		<tr>
			<td>Handle of paint roller</td>
			<td>any</td>
			<td></td>
			<td>to be used as stylus handle, hollowed out center must be large enough for stylus to sit securely inside&#160;</td>
		</tr>
		<tr>
			<td>Medical tape</td>
			<td>any</td>
			<td></td>
			<td>To secure EMG electrodes</td>
		</tr>
		<tr>
			<td>PCI-6220 card DAQ</td>
			<td>National Instruments</td>
			<td></td>
			<td>To interface EMG system</td>
		</tr>
		<tr>
			<td>Photodiode Sensor</td>
			<td>Vishay</td>
			<td>BPW21R</td>
			<td>To record timing of task events into EMG trace.</td>
		</tr>
		<tr>
			<td>Rear TMS port</td>
			<td>Magstim</td>
			<td></td>
			<td>Included with TMS machine</td>
		</tr>
		<tr>
			<td>Right-handed polyethylene glove</td>
			<td>any</td>
			<td></td>
			<td>Cut out thumb and index finger of glove to expose FDI muscle</td>
		</tr>
		<tr>
			<td>Sensory Adhesive Interface, 2-slot</td>
			<td>Delsys</td>
			<td>SC-F01</td>
			<td></td>
		</tr>
		<tr>
			<td>Stylus</td>
			<td>Wacom</td>
			<td>Intuos-4 grip pen</td>
			<td></td>
		</tr>
		<tr>
			<td>Tablet-to-Computer USB cable&#160;</td>
			<td>any</td>
			<td></td>
			<td>Included in Tablet purchase</td>
		</tr>
		<tr>
			<td>Task Monitor</td>
			<td>Asus</td>
			<td>VG248</td>
			<td></td>
		</tr>
		<tr>
			<td>TMS coil</td>
			<td>Magstim</td>
			<td>D70 Remote Coil</td>
			<td>7cm diameter, figure-of-eight coil</td>
		</tr>
		<tr>
			<td>TMS machine</td>
			<td>Magstim</td>
			<td>200-2</td>
			<td></td>
		</tr>
		<tr>
			<td>TMS-to-Computer DB9 cable</td>
			<td>any</td>
			<td></td>
			<td>Connects to PCIe Serial Card</td>
		</tr>
		<tr>
			<td>Velcro</td>
			<td>any</td>
			<td></td>
			<td>To be placed on glove and stylus handle</td>
		</tr>
	</tbody>
</table>

assessing corticospinal excitability during goal-directed reaching behavior

伸展是运动生理学和神经科学研究中广泛研究的行为。虽然已经使用各种行为操作检查了到达，但在理解到达计划、执行和控制所涉及的神经过程方面仍然存在重大差距。这里描述的新方法将二维到达任务与经颅磁刺激（TMS）和来自多个肌肉的同步肌电图（EMG）记录相结合。该方法允许在伸展运动展开期间的精确时间点无创检测皮质脊髓活动。示例任务代码包括一个延迟响应到达任务，其中两个可能的目标显示在偏离中线 45° ±。单脉冲TMS在大多数任务试验中提供，无论是在准备提示（基线）开始时还是在命令性提示（延迟）之前100毫秒。该样品设计适用于研究 REACH 制备过程中皮质脊髓兴奋性的变化。示例代码还包括视觉运动扰动（即光标旋转 ± 20°），以研究适应对 REACH 准备过程中皮质脊髓兴奋性的影响。可以调整任务参数和TMS交付，以解决有关到达行为期间运动系统状态的特定假设。在最初的实施中，83%的TMS试验成功诱发了运动诱发电位（MEP），并在所有试验中记录了到达轨迹。

所描述方法的成功执行包括记录片剂数据、肌电图痕迹和可靠地引出 MEP。完成了一项实验，其中包括270项测试试验，其中4/5试验（216项试验）提供了TMS。
数据来自16名年龄在25岁±10岁的参与者（8名女性8名男性），他们都自我报告为右撇子。我们通过推导一位代表性参与者的学习函数来评估视觉扰动对行为表现的有效性。 这些数据如图1B 所示，并显示?...

Watch this Scientific Journal Video about Assessing Corticospinal Excitability During Goal-Directed Reaching Behavior at JoVE.com

Assessing Corticospinal Excitability During Goal-Directed Reaching Behavior

伸手是允许人类与环境互动的基本技能。一些研究旨在使用各种方法表征到达行为。本文提供了经颅磁刺激的开源应用，以评估人类在达到任务执行期间的皮质脊髓兴奋状态。

评估目标导向到达行为期间的皮质脊髓兴奋性

Reaching is a widely studied behavior in motor physiology and neuroscience research. While reaching has been examined using a variety of behavioral ...

This method combines a flexible behavioral reaching paradigm with transcranial magnetic stimulation electromyography all from a single computer.This is significant because it streamlines existing approaches.The main advantages of this technique are that it is openly available and offers flexibility in terms of behavioral task and TMS parameters.This setup can easily be adapted for use with clinical populations, as it is versatile and mobile.The method offers unique insights into corticospinal excitability and muscle activity during behavioral reaching and motor adaptation.Place a large graphics tablet flat on a desktop.Use an adjustable 80-20 aluminum frame to position the task monitor six to eight inches above the tablet in parallel with the screen facing upward.Next, position the subject in a comfortable chair facing the tablet and ensure that the knees are flexed to 90

assessing-corticospinal-excitability-during-goal-directed-reaching

Research

JoVE Journal

Neuroscience

1.6K 次观看.  University of Oregon. This method combines a flexible behavioral reaching paradigm with transcranial magnetic stimulation electromyography all from a single computer.This is significant because it streamlines existing approaches.The main advantages of this technique are that it is openly available and offers flexibility in terms of behavioral task and TMS parameters.This setup can easily be adapted for use with clinical populations, as it is versatile and mobile.The method offers unique insights into corticospinal...

视频: Assessing Corticospinal Excitability During Goal-Directed Reaching Behavior

Cryopreservation of Cortical Tissue Blocks for the Generation of Highly Enriched Neuronal Cultures

In this study, we outline a standardized protocol for the successful cryopreservation and thawing of cortical brain tissue blocks to generate highly enriched neuronal cultures. For this protocol the freezing medium used is 10% dimethyl sulfoxide (DMSO) diluted in Hank's Buffered Salt Solution (HBSS). Blocks of cortical tissue are transferred to cryovials containing the freezing medium and slowly frozen at -1&deg;C/min in a rate-controlled freezing container. Post-thaw processing and dissociation of frozen tissue blocks consistently produced neuronal-enriched cultures which exhibited rapid neuritic growth during the first 5 days in culture and significant expansion of the neuronal network within 10 days. Immunocytochemical staining with the astrocytic marker glial fibrillary acidic protein (GFAP) and the neuronal marker beta-tubulin class III, revealed high numbers of neurons and astrocytes in the cultures. Generation of neural precursor cell cultures after tissue block dissociation resulted in rapidly expanding neurospheres, which produced large numbers of neurons and astrocytes under differentiating conditions. This simple cryopreservation protocol allows for the rapid, efficient, and inexpensive preservation of cortical brain tissue blocks, which grants increased flexibility for later generation of neuronal, astrocyte, and neuronal precursor cell cultures.

Here, we describe a method for efficient cryopreservation and thawing of cortical brain tissue blocks to generate highly enriched neuronal cultures. This simple protocol provides flexibility for later generation of neuronal, astrocyte, and neuronal precursor cell cultures.

生成高度丰富的神经文化皮层组织块冷冻保存

In this study, we outline a standardized protocol for the successful cryopreservation and thawing of cortical brain tissue blocks to generate highly ...

科研

神经科学

T-maze Forced Alternation and Left-right Discrimination Tasks for Assessing Working and Reference Memory in Mice

Forced alternation and left-right discrimination tasks using the T-maze have been widely used to assess working and reference memory, respectively, in rodents. In our laboratory, we evaluated the two types of memory in more than 30 strains of genetically engineered mice using the automated version of this apparatus. Here, we present the modified T-maze apparatus operated by a computer with a video-tracking system and our protocols in a movie format. The T-maze apparatus consists of runways partitioned off by sliding doors that can automatically open downward, each with a start box, a T-shaped alley, two boxes with automatic pellet dispensers at one side of the box, and two L-shaped alleys. Each L-shaped alley is connected to the start box so that mice can return to the start box, which excludes the effects of experimenter handling on mouse behavior. This apparatus also has an advantage that in vivo microdialysis, in vivo electrophysiology, and optogenetics techniques can be performed during T-maze performance because the doors are designed to go down into the floor. In this movie article, we describe T-maze tasks using the automated apparatus and the T-maze performance of &alpha;-CaMKII+/- mice, which are reported to show working memory deficits in the eight-arm radial maze task. Our data indicated that &alpha;-CaMKII+/- mice showed a working memory deficit, but no impairment of reference memory, and are consistent with previous findings using the eight-arm radial maze task, which supports the validity of our protocol. In addition, our data indicate that mutants tended to exhibit reversal learning deficits, suggesting that &alpha;-CaMKII deficiency causes reduced behavioral flexibility. Thus, the T-maze test using the modified automatic apparatus is useful for assessing working and reference memory and behavioral flexibility in mice.

This article presents the protocol of T-maze tests using a modified automated apparatus for assessing the learning and memory functions in mice.

T-迷宫强迫交替和评估工作，并参考在小鼠记忆的左，右的歧视任务

Forced alternation and left-right discrimination tasks using the T-maze have been widely used to assess working and reference memory, respectively, in ...

Nerve Excitability Assessment in Chemotherapy-induced Neurotoxicity

Chemotherapy-induced neurotoxicity is a serious consequence of cancer treatment, which occurs with some of the most commonly used chemotherapies1,2. Chemotherapy-induced peripheral neuropathy produces symptoms of numbness and paraesthesia in the limbs and may progress to difficulties with fine motor skills and walking, leading to functional impairment. In addition to producing troubling symptoms, chemotherapy-induced neuropathy may limit treatment success leading to dose reduction or early cessation of treatment. Neuropathic symptoms may persist long-term, leaving permanent nerve damage in patients with an otherwise good prognosis3. As chemotherapy is utilised more often as a preventative measure, and survival rates increase, the importance of long-lasting and significant neurotoxicity will increase.
There are no established neuroprotective or treatment options and a lack of sensitive assessment methods. Appropriate assessment of neurotoxicity will be critical as a prognostic factor and as suitable endpoints for future trials of neuroprotective agents. Current methods to assess the severity of chemotherapy-induced neuropathy utilise clinician-based grading scales which have been demonstrated to lack sensitivity to change and inter-observer objectivity4. Conventional nerve conduction studies provide information about compound action potential amplitude and conduction velocity, which are relatively non-specific measures and do not provide insight into ion channel function or resting membrane potential. Accordingly, prior studies have demonstrated that conventional nerve conduction studies are not sensitive to early change in chemotherapy-induced neurotoxicity4-6. In comparison, nerve excitability studies utilize threshold tracking techniques which have been developed to enable assessment of ion channels, pumps and exchangers in vivo in large myelinated human axons7-9.
Nerve excitability techniques have been established as a tool to examine the development and severity of chemotherapy-induced neurotoxicity10-13. Comprising a number of excitability parameters, nerve excitability studies can be used to assess acute neurotoxicity arising immediately following infusion and the development of chronic, cumulative neurotoxicity. Nerve excitability techniques are feasible in the clinical setting, with each test requiring only 5 -10 minutes to complete. Nerve excitability equipment is readily commercially available, and a portable system has been devised so that patients can be tested in situ in the infusion centre setting. In addition, these techniques can be adapted for use in multiple chemotherapies.
In patients treated with the chemotherapy oxaliplatin, primarily utilised for colorectal cancer, nerve excitability techniques provide a method to identify patients at-risk for neurotoxicity prior to the onset of chronic neuropathy. Nerve excitability studies have revealed the development of an acute Na+ channelopathy in motor and sensory axons10-13. Importantly, patients who demonstrated changes in excitability in early treatment were subsequently more likely to develop moderate to severe neurotoxicity11. However, across treatment, striking longitudinal changes were identified only in sensory axons which were able to predict clinical neurological outcome in 80% of patients10. These changes demonstrated a different pattern to those seen acutely following oxaliplatin infusion, and most likely reflect the development of significant axonal damage and membrane potential change in sensory nerves which develops longitudinally during oxaliplatin treatment10. Significant abnormalities developed during early treatment, prior to any reduction in conventional measures of nerve function, suggesting that excitability parameters may provide a sensitive biomarker.

This abstract describes a novel method to assess the development of neurotoxicity in patients receiving chemotherapy treatment. While conventional assessment methods are limited in their ability to detect early changes in nerve function, nerve excitability techniques provide early identification of patients at risk of severe neurotoxicity and insight into pathophysiology.

化疗引起的毒性神经的兴奋性评估

Chemotherapy-induced neurotoxicity is a serious consequence of cancer treatment, which occurs with some of the most commonly used chemotherapies1,2. ...

A Procedure for Implanting a Spinal Chamber for Longitudinal In Vivo Imaging of the Mouse Spinal Cord

Studies in the mammalian neocortex have enabled unprecedented resolution of cortical structure, activity, and response to neurodegenerative insults by repeated, time-lapse in vivo imaging in live rodents. These studies were made possible by straightforward surgical procedures, which enabled optical access for a prolonged period of time without repeat surgical procedures. In contrast, analogous studies of the spinal cord have been previously limited to only a few imaging sessions, each of which required an invasive surgery. As previously described, we have developed a spinal chamber that enables continuous optical access for upwards of 8 weeks, preserves mechanical stability of the spinal column, is easily stabilized externally during imaging, and requires only a single surgery. Here, the design of the spinal chamber with its associated surgical implements is reviewed and the surgical procedure is demonstrated in detail. Briefly, this video will demonstrate the preparation of the surgical area and mouse for surgery, exposure of the spinal vertebra and appropriate tissue debridement, the delivery of the implant and vertebral clamping, the completion of the chamber, the removal of the delivery system, sealing of the skin, and finally, post-operative care. The procedure for chronic in vivo imaging using nonlinear microscopy will also be demonstrated. Finally, outcomes, limitations, typical variability, and a guide for troubleshooting are discussed.

A Procedure for Implanting a Spinal Chamber for Longitudinal In Vivo Imaging of the Mouse Spinal Cord

In this video, we describe a procedure for implanting a chronic optical imaging chamber over the dorsal spinal cord of a live mouse. The chamber, surgical procedure, and chronic imaging are reviewed and demonstrated.

一个程序，植入脊柱商会纵<em&gt;在体内</em&gt;鼠标脊髓成像

Studies in the mammalian neocortex have enabled unprecedented resolution of cortical structure, activity, and response to neurodegenerative insults by ...

Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

Many organisms use cues and signals beyond human sensitivity during social interactions. It is important to take into account how organisms perceive their worlds when trying to understand their behavior and ecology. Sensitivity to the ultraviolet spectrum (UV; 300 - 400 nm) is found across multiple genera of birds, fish, reptiles, amphibians, and even mammals. This protocol describes a technique for examining organisms for the presence of UV-reflecting structures and a method for testing whether these cues are used as social signals in the context of mate choice. A spectrophotometer is used to detect the presence of UV reflectance and variation in reflective intensity between individuals and sexes. An example of this technique is presented in which a dichotomous mate choice test exposes sexually receptive individuals to opposite sex individuals whose visual appearance can be manipulated by filters that either transmit full spectrum or block UV wavelengths. This system allowed for the determination that female, but not male, sailfin mollies (Poecilia latipinna) were using UV markings as part of their mating decisions. These types of studies serve to expand our knowledge of the range of organisms that utilize UV and provide insight into how UV plays a role in their lives.

Assays to Detect UV-reflecting Structures and Determine their Importance in Mate Preference using the Sailfin Molly Poecilia latipinna

This protocol outlines the use of spectrophotometry to detect ultraviolet-reflecting structures on organisms (in this example, the sailfin molly Poecilia latipinna) and describes dichotomous choice tests for fish that allow inferences to be made on the role of ultraviolet cues during mate selection.

试验以检测紫外线反射结构，并使用茉莉花鳉确定其重要性伴侣偏好<em&gt;孔雀鱼latipinna</em

Many organisms use cues and signals beyond human sensitivity during social interactions. It is important to take into account how organisms perceive their ...

In Utero Electroporation Approaches to Study the Excitability of Neuronal Subpopulations and Single-cell Connectivity

The nervous system is composed of an enormous range of distinct neuronal types. These neuronal subpopulations are characterized by, among other features, their distinct dendritic morphologies, their specific patterns of axonal connectivity, and their selective firing responses. The molecular and cellular mechanisms responsible for these aspects of differentiation during development are still poorly understood.
Here, we describe combined protocols for labeling and characterizing the structural connectivity and excitability of cortical neurons. Modification of the in utero electroporation (IUE) protocol allows the labeling of a sparse population of neurons. This, in turn, enables the identification and tracking of the dendrites and axons of individual neurons, the precise characterization of the laminar location of axonal projections, and morphometric analysis. IUE can also be used to investigate changes in the excitability of wild-type (WT) or genetically modified neurons by combining it with whole-cell recording from acute slices of electroporated brains. These two techniques contribute to a better understanding of the coupling of structural and functional connectivity and of the molecular mechanisms controlling neuronal diversity during development. These developmental processes have important implications on axonal wiring, the functional diversity of neurons, and the biology of cognitive disorders.

In Utero Electroporation Approaches to Study the Excitability of Neuronal Subpopulations and Single-cell Connectivity

This manuscript provides protocols that use in utero electroporation (IUE) to describe the structural connectivity of neurons at the single-cell level and the excitability of fluorescently labeled neurons. Histology is used to characterize dendritic and axonal projections. Whole-cell recording in acute slices is used to investigate excitability.

在子宫内电途径研究神经元亚群的兴奋性和单细胞连接

The nervous system is composed of an enormous range of distinct neuronal types. These neuronal subpopulations are characterized by, among other features, ...

Operant Protocols for Assessing the Cost-benefit Analysis During Reinforced Decision Making by Rodents

Reinforcement-guided decision making is the ability to choose between competing courses of action based on the relative value of the benefits and their consequences. This process is integral to the normal human behavior and has been shown to be disrupted by neurological and psychiatric disorders such as addiction, schizophrenia, and depression. Rodents have long been used to uncover the neurobiology of human cognition. To this end, several behavioral tasks have been developed; however, most are non-automated and are labor-intensive. The recent development of the open-source microcontroller has enabled researchers to automate operant-based tasks for assessing a variety of cognitive tasks, standardizing the stimulus presentation, improving the data recording and consequently, improving the research output. Here, we describe an automated delay-based reinforcement-guided decision-making task, using an operant T-maze controlled by custom-written software programs. Using these decision-making tasks, we show the changes in the local field potential activities in the anterior cingulate cortex of a rat whilst it performs a delay-based cost-and-benefit decision-making task.

A cost-benefit analysis is a weighing-scale approach that the brain performs during the course of decision making. Here, we propose a protocol to train rats on an operant-based decision-making paradigm where rats choose higher rewards at the expense of waiting for 15 s to receive them.

啮齿动物强化决策中成本效益分析的操作协议

Reinforcement-guided decision making is the ability to choose between competing courses of action based on the relative value of the benefits and their ...

Generation of 3-D Collagen-based Hydrogels to Analyze Axonal Growth and Behavior During Nervous System Development

This protocol uses natural type I collagen to generate three-dimensional (3-D) hydrogel for monitoring and analyzing the axonal growth. The protocol is centered on culturing small pieces of embryonic or early postnatal rodent brains inside a 3-D hydrogel formed by the rat tail tendon-derived type I collagen with specific porosity. Tissue pieces are cultured inside the hydrogel and confronted to specific brain fragments or genetically-modified cell aggregates to produce and secrete molecules suitable for creating a gradient inside the porous matrix. The steps of this protocol are simple and reproducible but include critical steps to be considered carefully during its development. Moreover, the behavior of growing axons can be monitored and analyzed directly using a phase-contrast microscope or mono/multiphoton fluorescence microscope after fixation by immunocytochemical methods.

Here, we provide a method for analyzing the behavior of growing axons in 3D matrices, mimicking their natural development.

生成基于3-D胶原蛋白的水凝胶,分析神经系统发育过程中的Axonal生长和行为

This protocol uses natural type I collagen to generate three-dimensional (3-D) hydrogel for monitoring and analyzing the axonal growth. The protocol is ...

Measuring Neural Mechanisms Underlying Sleep-Dependent Memory Consolidation During Naps in Early Childhood

Sleep is critical for daily functioning. One important function of sleep is the consolidation of memories, a process that makes them stronger and less vulnerable to interference. The neural mechanisms underlying the benefit of sleep for memory can be investigated using polysomnography (PSG). PSG is a combination of physiological recordings including signals from the brain (EEG), eyes (EOG), and muscles (EMG) that are used to classify sleep stages. In this protocol, we describe how PSG can be used in conjunction with behavioral memory assessments, actigraphy, and parent-report to examine sleep-dependent memory consolidation. The focus of this protocol is on early childhood, a period of significance as children transition from biphasic sleep (consisting of a nap and overnight sleep) to monophasic sleep (overnight sleep only). The effects of sleep on memory performance are measured using a visuospatial memory assessment across periods of sleep and wakeful-rest. A combination of actigraphy and parent report is used to assess sleep rhythms (i.e., characterizing children as habitual or non-habitual nappers). Finally, PSG is used to characterize sleep stages and qualities of those stages (such as frequencies and the presence of spindles) during naps. The advantage of using PSG is that it is the only tool currently available to assess sleep quality and sleep architecture, pointing to the relevant brain state that supports memory consolidation. The main limitations of PSG are the length of time it takes to prepare the recording montage and that recordings are typically taken over one sleep bought. These limitations can be overcome by engaging young participants in distracting tasks during application and combining PSG with actigraphy and self/parent-report measures to characterize sleep cycles. Together, this unique combination of methods allows for investigations into how naps support learning in preschool children.

This protocol describes methods used to examine neural mechanisms underlying sleep-dependent memory consolidation during naps in early childhood. It includes procedures for examining the effect of sleep on behavioral memory performance, as well as the application and recording of both polysomnography and actigraphy.

测量幼儿期午睡期间睡眠依赖记忆巩固背后的神经机制

Sleep is critical for daily functioning. One important function of sleep is the consolidation of memories, a process that makes them stronger and less ...

Simultaneous Application of Transcranial Direct Current Stimulation during Virtual Reality Exposure

Transcranial direct current stimulation (tDCS) is a form of non-invasive brain stimulation that changes the likelihood of neuronal firing through modulation of neural resting membranes. Compared to other techniques, tDCS is relatively safe, cost-effective, and can be administered while individuals are engaged in controlled, specific cognitive processes. This latter point is important as tDCS may predominantly affect intrinsically active neural regions. In an effort to test tDCS as a potential treatment for psychiatric illness, the protocol described here outlines a novel procedure that allows the simultaneous application of tDCS during exposure to trauma-related cues using virtual reality (tDCS+VR) for veterans with posttraumatic stress disorder (NCT03372460). In this double-blind protocol, participants are assigned to either receive 2 mA tDCS, or sham stimulation, for 25 minutes while passively watching three 8-minute standardized virtual reality drives through Iraq or Afghanistan, with virtual reality events increasing in intensity during each drive. Participants undergo six sessions of tDCS+VR over the course of 2-3 weeks, and psychophysiology (skin conductance reactivity) is measured throughout each session. This allows testing for within and between session changes in hyperarousal to virtual reality events and adjunctive effects of tDCS. Stimulation is delivered through a built-in rechargeable battery-driven tDCS device using a 1 (anode) x 1 (cathode) unilateral electrode set-up. Each electrode is placed in a 3 x 3 cm (current density 2.22 A/m2) reusable sponge pocket saturated with 0.9% normal saline. Sponges with electrodes are attached to the participant&#8217;s skull using a rubber headband with the electrodes placed such that they target regions within the ventromedial prefrontal cortex. The virtual reality headset is placed over the tDCS montage in such a way as to avoid electrode interference.

This manuscript outlines a novel protocol to allow the simultaneous application of transcranial direct current stimulation during exposure to warzone trauma-related cues using virtual reality for veterans with posttraumatic stress disorder.