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道德声明研究感官刺激是如何通过神经元的活动表示没有替代使用动物及使用体内的方法。沿着科研（意大利语生物伦理委员会，对动物的研究中治疗法法令，1992年1月27日，第116号）动物诊疗意大利和欧洲法律的所有动物所受到的待遇。国家研究理事会，这里的实验已经进行，坚持国际委员会实验动物科学（的ICLA）代表联合国教育，科学及文化组织（教科文组织），理事会的国际医学科学组织（CIOMS的）和国际生物科学联合会（生物科联）。因此，概无特定协议的批准是必要的。卫生部的批准，被列为“比耶拉1，3/2011”到的伦理委员会的文件米兰大学。 1。实验动物的制备<ol><li>选择雄性大鼠的规模约300-400克。 </li><li>麻醉大鼠的实验制备。 <ol><li>腹腔注射巴比妥类溶液（戊巴比妥钠，50毫克/千克的感应，10毫克/千克维护）。 </li><li>检查麻醉水平低阈值和高阈值机械刺激过一爪子保证不回缩反射的发生。 </li><li>通过准确，然后做一个切口，显露颅面妆颅顶准备颅皮肤。 </li><li>导管插入气管，通过气管套管（2毫米和2.5mm的外径内径）由围绕气管本身手术结扎固定它。 </li><li>横向的气管段露出颈静脉和导管插入它插入连接到注射器，容器周围手术结扎固定它的插管（PE10）。 </li></ol></li><li>安装到大鼠立体定位仪。 <ol><li>固定鼠体，并装在仰卧位置。 </li><li>由耳棒固定头和阻塞口鼻部。放入老鼠的耳朵一两滴麻药（利多卡因）配售耳酒吧前。 </li></ol></li><li>调节立体定向垫由一个电子控制的加热垫的温度，将温度保持在37.5℃。 </li><li>气管导管连接到呼吸麻醉设备。 <ol><li>在1冲程/ s的传送异氟醚（2.5％0.4-0.8升/分）和O 2（0.15-0.2升/分钟）设置呼吸麻醉设备。 </li></ol></li><li>由聚维酮碘局部消毒剂仔细清理颅顶。 </li><li>取无菌手术刀切纵向上从吻端的基础上中线枕骨的角度。 <ol><li>由拉钩极叉开的伤口边界和两个小镊子娇养修复它们应用于吨Ø伤口边界。 </li><li>通过指向一个灯的一个骷髅跳马观察半透明表面识别骨膜。小心划伤骨库取出骨膜和暴露的骨面。 </li><li>提供止血用棉签或海绵手术在骨面上，如果焦点出血出现在骨。 </li></ol></li><li>用细尖笔，找出立体定向点前囟门在mediosagittal和冠状suturas的交叉点。 <ol><li>在手术显微镜控制，识别立体定向区（S1HL）覆盖躯体感觉皮层对侧用于实验（前囟门，美联社-0.5至1.2毫米，LL -2.3至2.5毫米）的后爪。 </li></ol></li><li>用细尖钢笔绘制正方形周长划定要钻的孔。 <ol><li>钻在先前确定的蓝色正方形去除骨瓦9 平方毫米的孔。 </li><li>清洁和吸收潜在的骨出血。 </l我&gt; <li>小心取出硬脑膜并覆盖皮质表面与外科海绵浸泡在保持在37.5℃，人工脑脊液</li></ol></li><li>固定在电极基体，以连接到电子显微操作器的保持器。 <ol><li>由microconnector矩阵的头部连接到前置放大器。 </li><li> （在手术显微镜控制）手动驱动电极矩阵到皮质表面（不接触它）。 </li><li>开关与听觉信号的放大器。 </li><li>驱动，由电子控制，电极矩阵触摸皮质表面，直到接触达成，通过明确的噪声音变信号。 </li><li>直到350-400微米的深度达到（皮质层四）由电子控制的步骤（2微米/步）拉下电极矩阵。 </li><li>检查神经元的反应由光触摸对侧后方的爪子。 </li><li>通过几个步骤进一步调整的深度，直到一个明确的扣球反应观察。 </li><li>通过静脉加拉碘thriethiodide（20毫克/千克/小时）麻痹的大鼠。使用剂量折射整个实验过程中保持curarization水平（0.4毫升/ 1小时）。 </li></ol></li></ol> 2。信号处理<ol><li>设置软件带通滤波器[0.1 6000]赫兹。 <ol><li>记录采样32 kHz的8个通道微电极矩阵的神经信号。 </li></ol></li><li>采集结束后，输出所记录的信号转换成适合于后处理的二进制格式。 <ol><li>由Wave_clus工具箱11，履行秒杀分拣过程。 </li></ol></li></ol> 3。制造及刺激装置的配置<ol><li>挂载一个仙人掌茎（带钝尖）垂直于中档音箱胶柄的基础上向盖的表面。 </li><li>程序微控制器提供电压脉冲到一个驱动电路，用于将扬声器。 <ol><li>编程的微控制器，提供的电流在每个第二两个成对脉冲（参见图1C）。 </li></ol></li><li>由L293D H桥基本无源元件（ 见图1A）的方式组装的扬声器和微控制器。 </li><li>微控制器连接到12伏可充电电池。 </li></ol> 4。刺激方案<ol><li>胶水后肢足爪的背方面坚实的框架，露出掌面，避免关节扭力。 </li><li>将刺激装置的前端到所需肢/爪的位置。 </li><li>切换刺激装置上。 </li></ol>离线步骤： 5。评估穗的回应<ol><li>对于每个记录的神经元，计算刺激的香农互信息（MI）的诱发扣球活动（CA刺激tegory要么是1，刺激，或0，无刺激）。 <ol><li>估计的条件响应概率P（R | S），其中s表示刺激类别和r表示内一个固定的时间窗口发射尖峰的个数。 </li><li>通过使用洗牌过程中所述9纠正心肌梗死的估计。 </li></ol></li></ol> 6。评价LFP的回应<ol><li>过滤所记录的信号的频率范围[0.1 100]赫兹为了获得LFP信号</li><li>计算LFPSNR，度量LFP反应性，如在参照图10进行说明。 </li></ol> 7。模型估计<ol><li>生成的形状的模型 <img alt="" fo:content-width="2in" height="24" src="/files/ftp_upload/50941/50941eq1.jpg" width="155" /> 其中x参数代表三种不同的LFP功能（×1的磷酸锂铁本身，&lt;<html> em&gt;的×2及其衍生物以及x 3及其衍生物的希尔伯特相位） 和 F表示如下 <img alt="" fo:content-width="2in" height="57" src="/files/ftp_upload/50941/50941eq2.jpg" width="187" /> 的克系数是线性组合的权重，f是一个运算符，它取任一的绝对值，电源或各x i的原始值。 </li><li>使用NSGAII算法通过使用以下三个对象来估计模型参数和操作：ⅰ）一个地方，试验对试验的基础上，测量拟合<img alt="" fo:content-width="2in" height="40" src="/files/ftp_upload/50941/50941eq3.jpg" width="149" /> ，其中N 藻和N r个分别代表整体尖峰计数和响应向量的长度;ⅱ）基于平均响应全局拟合度量ad/50941/50941eq4.jpg“宽度=”205“/&gt;其中N RESP代表的平均响应的长度; iii）较模型复杂度的度量（另见）10 <img alt="" fo:content-width="2in" height="23" src="/files/ftp_upload/50941/50941eq5.jpg" width="161" /> 。 </li></ol> 8。组织学确认<ol><li>牺牲老鼠。 <ol><li>在实验记录的结尾，把动物在深麻醉气体（异氟醚2％，4升/分钟）和静脉注射巴比妥酸盐过量（&gt; 50毫克/千克，戊巴比妥）。 </li><li>等待心脏骤停。 </li><li>从立体定位仪卸载鼠</li></ol></li><li>灌注。 <ol><li>将大鼠网格铺设了到盆收集血液和体液从灌注。 </li><li>通过解剖从肋部插入分离胸骨，打开胸腔。 </li><li>与溺爱镊子和倾覆Cr块胸骨剑突过程anially胸骨和叉开将排骨过心脏。 </li><li>确定左心室和右心房，放置一个9轨距钝尖的针（连接到灌注套管）插入心室并与手术细剪刀打开右心房。 </li><li>开始灌注后的4％的福尔马林溶液（250毫升）中的灌注冷的（4℃）肝素化的生理溶液（250毫升）。 </li></ol></li><li>通过用合适的钳子口颅顶从头骨中提取大脑，并放置在大脑中有10％的福尔马林溶液中。 </li><li>一个星期后，用切片机（10微米厚）制备的组织学切片。 </li><li>染色用甲酚紫溶液大脑冠状面和矢状切面。 </li></ol>

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作者宣称没有竞争的财务权益。

这项工作首先提出了一种新的，简单的，低成本的装置，可帮助提供快速和空间点状的感官刺激。然后随机配对脉冲刺激方案和一套计算分析进行了验证。总体目标是建立在期间触觉刺激电生理记录LFP-秒杀关系的估计的框架。 该设备的协议和分析方法已共同促成的结果，即确定性方法能够描述和预测LFP到秒杀转变的第一个示范，一个神经过程仍然了解1很差。 <p cl...

在信号的情况下处理所述脉冲响应提供了一种动力系统的行为的一个基本特征。 虽然理想的脉冲刺激几乎是无法实现的，它可以通过使用产生高频率的位移的致动器元件，以取得它的合理近似。这种类型的轻触觉振动刺激是众所周知的同时针对皮肤深层（ 如快速反应，快速适应环层小体）2和表面受体（ 如低门槛慢慢适应默克尔盘状结构）2。 电流刺激装置，主要是压电致动器，被装入许多缺点，尤其共振和小的位移。为了克服这些缺陷，另一种实现的类似脉冲刺激，提出了用钝尖（在我们的情况下，仙人掌平滑尖）垂直安装在一个中档扬声器纸盆膜中心。这提供了较大的位移和更广泛的频谱的优点。 这种装置的有效应用是局部区域电位的相关神经生理问题钉鞋依赖关系的研究。由于这些电活动之间的微妙时间关联的是需要为外设提供一个刺激的精细调节装置。刺激必须是尽可能快和空间选择性越好，以减少“背景噪音”和锐化感兴趣的信号。到这个目的，刺激装置及刺激传递协议的任务一同被优化。在本文中，我们描述了该技术，并提出了一些有代表性的结果。 基于随机配对脉冲刺激一个协议已被设计和优化，以避免习惯。这个协议提供了对经典的优点编脉冲和降低刺激和神经元活动的自发周期性脉冲串之间的杂散锁定的可能性。 通过使用这个随机配对脉冲有可能获得快速和可靠的LFP和穗反应和捕捉这些反应相关的两个局部区域电位和尖峰对刺激的历史依赖性的特殊功能。的确，从原始的LFP反应，一组3 LFP的特点（对LFP本身，LFP一阶导数的一阶导数和相位）与平均峰值响应强烈关联，也被提取。 几个方法被提出，以适应该预测从局部区域电位3,4尖峰模型。在一般的模型拟合​​过程的一个临界点，也常见于从刺激信号尖峰事件的预测，是通过对目标函数最大化/最小化的适当选择构成。虽然一系列的目标函数已经提出D（ 例如，相关性和连贯性）5没有这些共同捕捉尖峰响应的整体复杂性。因此，基于多目标优化的一种新的框架被引入。我们发现，通过使用建议的制定和这个计算框架，可以估算一个基于强大的LFP秒杀关系集合预测模型。

<table border="0" cellpadding="0" cellspacing="0" width="1204">
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Microstepper</td>
			<td style="width:348px;">AB Transvertex (Stockholm, Sweden)</td>
			<td style="width:119px;"> </td>
			<td style="width:424px;">The microstepper used to pull down the electrode matrix</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">32 channels Cheetah System</td>
			<td style="width:348px;">Neuralynx (MT, USA)</td>
			<td style="width:119px;"> </td>
			<td>The electrophysiological recording system</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">L293D h-bridge</td>
			<td style="width:348px;">RS Components (Cinisello Balsamo, Italy)</td>
			<td style="width:119px;"> </td>
			<td>The bridge used to connect the microcontroller to the speaker</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;">H21A3 Optical Interrupter Switch</td>
			<td style="width:348px;">Fairchild Semiconductor Corporation (San Jose, California)</td>
			<td style="width:119px;"> </td>
			<td>The phototransistor used to estabilish the tip displacement</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Arduino Uno</td>
			<td style="width:348px;">Arduino (Duemilanove, Italy)</td>
			<td style="width:119px;"> </td>
			<td>The microcontroller used to deliver current pulse to the speaker</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Microelectrode Matrices GB1</td>
			<td style="width:348px;">FHC</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Isoflurane</td>
			<td style="width:348px;">Rhodia Organique Fine Ltd.</td>
			<td style="width:119px;"> </td>
			<td>The anaestetic used to prepare animals</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Stereotaxic apparatus</td>
			<td style="width:348px;">Narishighe (Tokyo, Japan)</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Sprague-Dawley male rats</td>
			<td style="width:348px;">Charles River (Calco, LC, Italy)</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Gallamine thriethiodide</td>
			<td style="width:348px;">Sigma-Aldrich</td>
			<td style="width:119px;"> </td>
			<td>The compound used to curarize the animals</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Cresyl violet</td>
			<td style="width:348px;">Sigma-Aldrich</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Topical antiseptics (Betadine 10%)</td>
			<td style="width:348px;">Meda Pharma (Milanm Italy)</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:313px;">Heparine</td>
			<td style="width:348px;">Sigma-Aldrich</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:313px;">Formaldehyde</td>
			<td style="width:348px;">Carlo Erba Reagents (Pomigliano Milanese, Milan, Italy)</td>
			<td style="width:119px;"> </td>
			<td></td>
		</tr>
	</tbody>
</table>

a simple stimulatory device for evoking point-like tactile stimuli: a searchlight for lfp to spike transitions

目前神经生理学的研究具有的宗旨，以制定方法，调查从神经元的信号途径神经元，即从峰值到局部场电位（的LFP）的转换和从局部区域电位尖峰。 局部区域电位有对秒杀活动的复杂的依赖，它们之间的关系仍然是了解1很差。这些信号关系的阐明将有助于双方为临床诊断（ 如刺激范式的深部脑刺激）和神经编码策略在正常和病理条件下（ 如癫痫，帕金森病，慢性疼痛）有更深的理解。到这个目的，就必须解决与刺激装置，刺激模式和计算分析的技术问题。因此，一个定制的刺激装置，以提供刺激以及调控的空间和时间，这并不在机械共振产生的开发。随后，作为一个示例，一组可靠LFP-穗关系萃取。 该设备的性能是由细胞外记录调查，共同尖峰和LFP反应施加的刺激，从大鼠初级躯体感觉皮层。然后，通过一个多目标优化策略的手段，估计基于局部区域电位的预测模型为尖峰发生。 这种模式的应用表明，该装置有足够的适合提供高频率的触觉刺激，表现优于常见的压电致动器。作为该装置的效能的证明，以下结果表示：1）磷酸锂铁的反应的时间和可靠性良好匹配穗反应，2）的LFP是对刺激敏感的历史和捕获不仅平均响应也是试验对试验的波动峰电活动，最后3）通过使用LFP信号是可能的估算范围Øf预测模型，捕捉了秒杀活动的不同方面。

提示偏移功能为了表征所提出刺激装置的动力学性质，一系列的实验设置。它由一个砷化镓红外发光二极管加上一个硅光电晶体管的一个具体装置是用来评估尖端位移，位移持续时间和可能的位移延迟。通过这种光断续器开关的手段，我们放在刺激尖上的发光二极管孔（高度= 1毫米）的边缘，无论是微控制器和光电晶体管输出的记录。配售过程提供了便利一microstepper装置为1毫?...

Watch this Scientific Journal Video about A Simple Stimulatory Device for Evoking Point-like Tactile Stimuli: A Searchlight for LFP to Spike Transitions at JoVE.com

A Simple Stimulatory Device for Evoking Point-like Tactile Stimuli: A Searchlight for LFP to Spike Transitions

为了阐明从局部场电位（的LFP）的复​​杂过渡到尖峰合适的刺激轻机械刺激外周建。作为一个应用，从躯体感觉皮层记录扣球活动由一个多目标优化策略进行了分析。结果表明，所提出的刺激是能够提供触觉刺激与毫秒和毫米精度。

一个简单的刺激装置唤起点状触觉刺激：探照灯为LFP到穗转换

Current neurophysiological research has the aim to develop methodologies to investigate the signal route from neuron to neuron, namely in the transitions ...

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Linear Time- Invariant Systems

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9.8K 次观看.  National Research Council. 为了阐明从局部场电位（的LFP）的复​​杂过渡到尖峰合适的刺激轻机械刺激外周建。作为一个应用，从躯体感觉皮层记录扣球活动由一个多目标优化策略进行了分析。结果表明，所提出的刺激是能够提供触觉刺激与毫秒和毫米精度。 

视频: A Simple Stimulatory Device for Evoking Point-like Tactile Stimuli: A Searchlight for LFP to Spike Transitions

A Simple Composite Phenotype Scoring System for Evaluating Mouse Models of Cerebellar Ataxia

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebella ataxia. It is derived from previously published phenotype assessments in several disease models, including spinocerebellar ataxias, Huntington s disease and spinobulbar muscular atrophy. Measures include hind limb clasping, ledge test, gait and kyphosis. Each measure is recorded on a scale of 0-3, with a combined total of 0-12 for all four measures. The results effectively discriminate between affected and non-affected individuals, while also quantifying the temporal progression of neurodegenerative disease phenotypes. Measures may be analyzed individually or combined into a composite phenotype score for greater statistical power. The ideal combination of the four described measures will depend upon the disorder in question. We present an example of the protocol used to assess disease severity in a transgenic mouse model of spinocerebellar ataxia type 7 (SCA7).
Albert R. La Spada and Gwenn A. Garden contributed to this manuscript equally.

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebellar ataxia. Measures include hind limb clasping, ledge test, gait and kyphosis. This protocol effectively discriminates between affected and non-affected individuals, and detects the progression of affected individuals over time.

一个简单的综合评分系统评估小脑共济失调的小鼠模型的表型

We describe a protocol for the rapid and sensitive quantification of disease severity in mouse models of cerebella ataxia. It is derived from previously ...

科研

神经科学

The Vermicelli and Capellini Handling Tests: Simple quantitative measures of dexterous forepaw function in rats and mice

Previous characterizations of rodent eating behavior have revealed that they use coordinated forepaw movements to manipulate food pieces. We have extended upon this work to develop a simple quantitative measure of forepaw dexterity that is sensitive to lateralized impairments and age-dependent changes. Rodents learn skillful forepaw and digit movements to manage thin pasta pieces, which they eagerly consume. We have previously described methods for quantifying vermicelli handling in rats and showed that the measures are very sensitive to forelimb impairments resulting from unilateral ischemic lesions, middle cerebral artery occlusions and unilateral striatal dopamine depletion [Allred, R.P., Adkins, D.L., Woodlee, M.T., Husbands, L.C., Maldonado M.A., Kane, J.R., Schallert, T. &amp; Jones, T.A. The Vermicelli Handling Test: a simple quantitative measure of dexterous forepaw function in rats. J. Neurosci. Methods 170, 229-244 (2008)]. Here we present a more detailed protocol for this test in rats and compare it with a newly developed version for mice, the Capellini Handling Test. Rats and mice are videotaped while handling short lengths of uncooked vermicelli or capellini pasta, respectively, with a camera positioned to optimize the view of paw movements. Slow motion video playback allows for the identification of forepaw adjustments, defined as any distinct removal and replacement of the paw, or of any number of digits, on the pasta piece after eating commences. Forepaw adjustments per piece are averaged over trials per each testing session. Repeated testing permits sensitive quantitative analysis of changes in forepaw dexterity over time. Protocols for pre-testing habituation and handling practice, as well as procedures for characterizing atypical handling patterns, are described. Because rats and mice perform the pasta handling tests slightly differently, species-specific differences in administration and scoring of these tests are highlighted. All animal use was in accordance with protocols approved by the University of Texas at Austin Animal Care and Use Committee.

The Vermicelli and Capellini Handling Tests of forepaw dexterity take advantage of the natural inclination of rodents to manipulate food items using skillful forepaw and digit movements. Animals are videotaped while handling short strands of uncooked dry pasta. Slow motion video playback allows for the quantification of forepaw adjustments.

粉条和Capellini处理测试：在大鼠和小鼠的灵巧的前爪功能简单的定量措施

Previous characterizations of rodent eating behavior have revealed that they use coordinated forepaw movements to manipulate food pieces. We have extended ...

The NeuroStar TMS Device: Conducting the FDA Approved Protocol for Treatment of Depression

The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic fields. These rapidly alternating fields induce electrical currents within localized, targeted regions of the cortex which are associated with various physiological and functional brain changes.1,2,3
In 2007, O'Reardon et al., utilizing the NeuroStar device, published the results of an industry-sponsored, multisite, randomized, sham-stimulation controlled clinical trial in which 301 patients with major depression, who had previously failed to respond to at least one adequate antidepressant treatment trial, underwent either active or sham TMS over the left dorsolateral prefrontal cortex (DLPFC). The patients, who were medication-free at the time of the study, received TMS five times per week over 4-6 weeks.4
The results demonstrated that a sub-population of patients (those who were relatively less resistant to medication, having failed not more than two good pharmacologic trials) showed a statistically significant improvement on the Montgomery-Asberg Depression Scale (MADRS), the Hamilton Depression Rating Scale (HAMD), and various other outcome measures. In October 2008, supported by these and other similar results5,6,7, Neuronetics obtained the first and only Food and Drug Administration (FDA) approval for the clinical treatment of a specific form of medication-refractory depression using a TMS Therapy device (FDA approval K061053).
In this paper, we will explore the specified FDA approved NeuroStar depression treatment protocol (to be administered only under prescription and by a licensed medical profession in either an in- or outpatient setting).

In this article, we examine the methodology and considerations relevant to the FDA approved depression treatment protocol using the Neuronetics NeuroStar TMS device.

NeuroStar TMS的设备进行美国FDA批准用于治疗抑郁症的议定书“

The Neuronetics NeuroStar Transcranial Magnetic Stimulation (TMS) System is a class II medical device that produces brief duration, pulsed magnetic ...

A Simple Way to Measure Ethanol Sensitivity in Flies

Low doses of ethanol cause flies to become hyperactive, while high doses are sedating. The sensitivity to ethanol-induced sedation of a given fly strain is correlated with that strain s ethanol preference, and therefore sedation is a highly relevant measure to study the genetics of alcohol responses and drinking. We demonstrate a simple way to expose flies to ethanol and measure its intoxicating effects. The assay we describe can determine acute sensitivity, as well as ethanol tolerance induced by repeat exposure. It does not require a technically involved setup, and can therefore be applied in any laboratory with basic fly culture tools.

A simple assay to measure the sedating effects of ethanol on Drosophila flies, based on the loss of righting reflex, is described.

一个简单的方法来衡量在果蝇的乙醇灵敏度

Low doses of ethanol cause flies to become hyperactive, while high doses are sedating. The sensitivity to ethanol-induced sedation of a given fly strain ...

Ex utero Electroporation and Whole Hemisphere Explants: A Simple Experimental Method for Studies of Early Cortical Development

Cortical development involves complex interactions between neurons and non-neuronal elements including precursor cells, blood vessels, meninges and associated extracellular matrix. Because they provide a suitable organotypic environment, cortical slice explants are often used to investigate those interactions that control neuronal differentiation and development. Although beneficial, the slice explant model can suffer from drawbacks including aberrant cellular lamination and migration. Here we report a whole cerebral hemisphere explant system for studies of early cortical development that is easier to prepare than cortical slices and shows consistent organotypic migration and lamination. In this model system, early lamination and migration patterns proceed normally for a period of two days in vitro, including the period of preplate splitting, during which prospective cortical layer six forms. We then developed an ex utero electroporation (EUEP) approach that achieves ~80% success in targeting GFP expression to neurons developing in the dorsal medial cortex.
The whole hemisphere explant model makes early cortical development accessible for electroporation, pharmacological intervention and live imaging approaches. This method avoids the survival surgery required of in utero electroporation (IUEP) approaches while improving both transfection and areal targeting consistency. This method will facilitate experimental studies of neuronal proliferation, migration and differentiation.

Ex utero Electroporation and Whole Hemisphere Explants: A Simple Experimental Method for Studies of Early Cortical Development

This protocol describes an improved explant procedure that involves ex utero electroporation, dissection and culture of entire cerebral hemispheres from the embryonic mouse. The preparation facilitates pharmacological studies and assays of gene function during early cortical development.

前子宫穿孔和整个半球的外植体早期皮质发育的研究：一个简单的实验方法

Cortical development involves complex interactions between neurons and non-neuronal elements including precursor cells, blood vessels, meninges and ...

Visualizing the Effects of a Positive Early Experience, Tactile Stimulation, on Dendritic Morphology and Synaptic Connectivity with Golgi-Cox Staining

To generate longer-term changes in behavior, experiences must be producing stable changes in neuronal morphology and synaptic connectivity. Tactile stimulation is a positive early experience that mimics maternal licking and grooming in the rat. Exposing rat pups to this positive experience can be completed easily and cost-effectively by using highly accessible materials such as a household duster. Using a cross-litter design, pups are either stroked or left undisturbed, for 15 min, three times per day throughout the perinatal period. To measure the neuroplastic changes related to this positive early experience, Golgi-Cox staining of brain tissue is utilized. Owing to the fact that Golgi-Cox impregnation stains a discrete number of neurons rather than all of the cells, staining of the rodent brain with Golgi-Cox solution permits the visualization of entire neuronal elements, including the cell body, dendrites, axons, and dendritic spines. The staining procedure is carried out over several days and requires that the researcher pay close attention to detail. However, once staining is completed, the entire brain has been impregnated and can be preserved indefinitely for ongoing analysis. Therefore, Golgi-Cox staining is a valuable resource for studying experience-dependent plasticity.

This paper describes the procedures for tactile stimulation of rat pups and subsequent Golgi-Cox staining of neuronal morphology. Tactile stimulation is a positive experience that is administered in the perinatal period by stroking pups with a household duster. Golgi-cox staining is a reliable procedure permitting the visualization of entire neurons.

可视化的一个积极的早期经验，触觉刺激的影响，对树突状形态和突触连接与高尔基染色考克斯

To generate longer-term changes in behavior, experiences must be producing stable changes in neuronal morphology and synaptic connectivity. Tactile ...

A Simple Behavioral Assay for Testing Visual Function in Xenopus laevis

Measurement of the visual function in the tadpoles of the frog, Xenopus laevis, allows screening for blindness in live animals. The optokinetic response is a vision-based, reflexive behavior that has been observed in all vertebrates tested. Tadpole eyes are small so the tail flip response was used as alternative measure, which requires a trained technician to record the subtle response. We developed an alternative behavior assay based on the fact that tadpoles prefer to swim on the white side of a tank when placed in a tank with both black and white sides. The assay presented here is an inexpensive, simple alternative that creates a response that is easily measured. The setup consists of a tripod, webcam and nested testing tanks, readily available in most Xenopus laboratories. This article includes a movie showing the behavior of tadpoles, before and after severing the optic nerve. In order to test the function of one eye, we also include representative results of a tadpole in which each eye underwent retinal axotomy on consecutive days. Future studies could develop an automated version of this assay for testing the vision of many tadpoles at once.

A Simple Behavioral Assay for Testing Visual Function in Xenopus laevis

Xenopus laevis tadpoles prefer swimming on the white side of a black/white tank. This behavior is guided by their vision. Based on this behavior, we present a simple assay to test the visual function of tadpoles.

一个简单的行为分析在检测视觉功能<em&gt;非洲爪蟾</em

Measurement of the visual function in the tadpoles of the frog, Xenopus laevis, allows screening for blindness in live animals. The optokinetic response ...

A Simple Alternative to Stereotactic Injection for Brain Specific Knockdown of miRNA

MicroRNAs (miRNAs) are key regulators of gene expression. In the brain, vital processes like neurodevelopment and neuronal functions depend on the correct expression of microRNAs. Perturbation of microRNAs in the brain can be used to model neurodegenerative diseases by modulating neuronal cell death. Currently, stereotactic injection is used to deliver miRNA knockdown agents to specific location in the brain. Here, we discuss strategies to design antagomirs against miRNA with locked nucleotide modifications (LNA). Subsequently describe a method for brain specific delivery of antagomirs, uniformly across different regions of the brain. This method is simple and widely applicable since it overcomes the surgery, associated injury and limitation of local delivery in stereotactic injections. We prepared a complex of neurotropic, cell-penetrating peptide Rabies Virus Glycoprotein (RVG) with antagomir against miRNA-29 and injected through tail vein, to specifically deliver in the brain. The antagomir design incorporated features that allow specific targeting of the miRNA and formation of non-covalent complexes with the peptide. The knock-down of the miRNA in neuronal cells, resulted in apoptotic cell death and associated behavioural defects. Thus, the method can be used for acute models of neuro-degeneration through the perturbation of miRNAs.

MicroRNAs play crucial roles in the brain and are potential targets for modeling neuro-degeneration. However, perturbing miRNA levels is challenging due to the short length of miRNA and inaccessibility of the brain tissue. This video presents a method for antagomir design and brain specific delivery using a neuropeptide in mice.

一个简单的替代，以立体定向注射miRNA的脑特异性敲除

MicroRNAs (miRNAs) are key regulators of gene expression.  In the brain, vital processes like neurodevelopment and neuronal functions depend on the ...

A Simple Flight Mill for the Study of Tethered Flight in Insects

Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have shown that flight mills are valuable tools for the experimental study of insect flight behavior, allowing researchers to examine how factors such as age, host plants, or population source can influence an insects' propensity to disperse. Flight mills allow researchers to measure components of flight such as speed and distance flown. Lack of detailed information about how to build such a device can make their construction appear to be prohibitively complex. We present a simple and relatively inexpensive flight mill for the study of tethered flight in insects. Experimental insects can be tethered with non-toxic adhesives and revolve around an axis by means of a very low friction magnetic bearing. The mill is designed for the study of flight in controlled conditions as it can be used inside an incubator or environmental chamber. The strongest points are the very simple electronic circuitry, the design that allows sixteen insects to fly simultaneously allowing the collection and analysis of a large number of samples in a short time and the potential to use the device in a very limited workspace. This design is extremely flexible, and we have adjusted the mill to accommodate different species of insects of various sizes.

Flight in insects is influenced by a number of factors and the propensity to disperse is an important variable in understanding insect ecology and biological control strategies. We describe the construction and use of a simple, relatively inexpensive, and flexible flight mill for measuring parameters of tethered flight in insects.

一个简单的飞行磨的系留飞行的昆虫研究

Flight in insects can be long-range migratory flights, intermediate-range dispersal flights, or short-range host-seeking flights. Previous studies have ...

A Simple Approach to Induce Experimental Autoimmune Neuritis in C57BL/6 Mice for Functional and Neuropathological Assessments

Experimental autoimmune neuritis (EAN) is a well-appreciated experimental model of autoimmune peripheral demyelinating diseases. EAN disease is induced by immunizing mice with neurogenic peptides to direct an inflammatory attack toward components of the peripheral nervous system (PNS). Recent advances have enabled the induction of EAN in the relatively resistant C57BL/6 mouse line using myelin protein zero (P0)106-125 or P0180-199 peptides delivered in adjuvant combined with the injection of pertussis toxin. The ability to induce EAN in the C57BL/6 strain allows for the use of the numerous genetic tools that exist on this mouse background, and thus allows the sophisticated study of disease pathogenesis and interrogation of the mechanistic action of novel therapeutics in combination with transgenic approaches. In this study, we demonstrate a simple approach to successfully induce EAN using the P0180-199 peptide in C57BL/6 mice. We also outline a protocol for the assessment of functional deficits that occur in this model, accompanied by an array of neuropathological features. Thus, this model is a powerful experimental model to study the pathogenesis of human peripheral demyelinating neuropathies, and to determine the efficacy of potential therapies that aim to promote myelin repair and protect against nerve damage in autoimmune neuritis.

This report outlines a simple approach to successfully induce experimental autoimmune neuritis (EAN) using the myelin protein zero (P0)180-199 peptide in combination with Freund&#39;s complete adjuvant and pertussis toxin. We present a sophisticated paradigm capable of accurately assessing the extent of functional deficits and neuropathology that occur in this EAN.

C57BL/6 小鼠自身免疫性神经炎的功能和病理评价的简易方法

Experimental autoimmune neuritis (EAN) is a well-appreciated experimental model of autoimmune peripheral demyelinating diseases. EAN disease is induced by ...