这项研究是药物滥用津贴的DA 006886，和DA 032270支持的研究所。

<ol><li>拧动线与驱动板提供的连接器线的双极步进电机。线的颜色上双极步进电动机是标准化的（ 图1）。 注:所描述的步进电机的分辨率为1.8°/步，面向0.346°/步。标准stereotax有旅游3 mm/360°。最终的分辨率为2.88微米/步。该电机还能够小数步进的。 <ol><li>绿色线连接到A +，黑线连接至A-，红色线连接到B +和蓝线连接到B-。 </li></ol></li><li>滑动耦合器在步进电机，注意对准安装孔，并用12倍 M3六角螺钉（20毫米）（ 图2）固定。 <ol><li>确保该耦合器被牢固地连接到电机。 注:3D模型不包括线程。部分标有螺纹尺寸，但它们必须被窃听他们毕竟制造。 </li></ol></li><li>从立体定位仪的使用小六角扳手所有3个轴的拇指握把取下固定螺丝。拇指握把有螺纹，所以将它们逆时针去除。保持PTFE垫圈代替在臂（ 图3）。 </li><li>用螺丝将衣领的螺纹端上的立体定位仪的武器的螺纹杆（ 图3）。 <ol><li>确保有衣领和PTFE O形圈之间没有间隙。这保证了坐标保持在机器人改变方向。 </li><li>固定衣领上使用3倍 NF10-32（1/4英寸）杯点紧定螺钉立体定位臂的线程。 </li><li>将每个电机和耦合器在衣领和立体定向武器。确保电机坐与臂平齐，并且在套环的固定螺丝孔与电动机轴的平面部分（ 图4）。 </li><li>安全合作使用该安装孔和6倍 NF10-32（1/2英寸）杯点紧定螺钉（ 图4）uplers到stereotax。 </li><li>固定套环，以使用3×NF10-32（1/4英寸）的固定螺钉（ 图4）的电机轴。 </li></ol></li><li>由每个控制器引脚设置为半步制备数控驱动板。 注:该步进电机驱动器来作为一个暴露的电路板。情况可以是内置的，尽管它不是必需的。另外，也可以使用许多不同的双极步进电机驱动器。如果是这样，请确保遵循针对购买的特定电路板的安装说明。 <ol><li>对齐所有6针每步进电机以同样的方式。半步允许在度/步（ 图5）步进分辨率的两倍。 </li><li>翻转针1到ON的位置，针2至关闭位置，引脚3 上的位置化，引脚4到关闭位置，引脚5到ON位置，和引脚6到关闭位置（ 图5）。 </li></ol></li><li>请将电机（X - Y - Z）为步进电机驱动器，以及12 V电源。正确的位置上标有驱动程序。此外，使用DB25cable（ 图6）连接步进驱动器到计算机的串行端口。 </li><li>安装数控铣床软件安装到个人计算机（这将需要位于手术区域）以下的默认指令。一旦安装完毕，打开开始配置软件。 <ol><li>配置软件与步进电机进行通信。 注:以下说明适用于只使用与TB6560的步进电机驱动器。 </li><li>点击进入该软件的菜单如下。打开→配置→端口和引脚→输出信号。在提示填写，以配合图7和命中适用。 </li><li>点击进入该软件的菜单如下。打开→配置→端口和引脚→输入信号。在提示填写，以配合图8和命中适用。 </li><li>点击进入该软件的菜单如下。打开→配置→端口和引脚→电机输出。在提示填写，以配合图9和命中适用。 </li><li>点击进入该软件的菜单如下。打开→配置→电机调谐。在提示匹配图10，然后单击保存轴设置填充。重复使用相同值的所有3个轴的上一步。 </li></ol></li><li>校准stereotax到CNC软件的规模。 注:该软件是专为标准铣床，所以衡量其单位不会成比例立体定位仪的旅行。<ol><li>电机"速度设定为每分钟1英寸，而"点动"立体定位仪的Z轴与上一页/下一页至毫米。 注:建议的最高点动速度是3500微米/秒，最大推荐的切削速度为500微米/秒。 </li><li>零的Z轴，以及操控立体定位仪1毫米。在Z轴中Mach3的行驶距离是"缩放常数"。机床坐标系是由"缩放常数"相乘头骨坐标（毫米）来确定。 </li><li>通过编程他们前往一些已知的距离执行所有3个轴的抽样测试，确保运动是准确的。如果stereotax行程过远或短，相应地修改比例常数。 注意:一旦缩放完成后，包括自定义脚本可用于生成G代码进行手术。但是，我们强烈建议用户熟悉G代码试图自动生成前手术。这是绝对必要的故障排除和修改自动手术。 </li></ol></li><li>使用超大探头支架装上微型马达钻立体定位仪。注:推荐的最小钻头转速为40,000 RPM。 </li><li>自动生成G代码与3头骨螺丝孔锐边开颅手术。 <ol><li>将所有的软件表中的自定义脚本到PC上的一个文件夹。 </li><li>打开脚本"SharpEdgeCraniotomy.m"并运行代码。 </li><li>选择两者的提示"是什么类型的手术你会被执行？" （ 图11）。 </li><li>选择自定义 ，定义骨窗的角落。在每个提示填写匹配图12。 </li><li>定义的开颅角落的X和Y位置。每个坐标必须以正确的顺序被输入，根据该示例在图13中</strong&gt;的信息。 </li><li>在提示符下输入3，产生3骷髅洞（ 图14）。 </li><li>选择定义使用坐标 ，并从图15中的模板输入每个孔的坐标。 注意:如果精确坐标并不重要，有一个选项来指向和点击孔的立场到大鼠头骨的图像。头寸将被自动生成。 </li><li>定义钻孔参数。对于第一次测试的手术，接受默认值。 注意:这些值依赖于步进电机，和动物的头骨。每鼠品种和目标位置有一个稍微不同颅骨厚度。对于使用该设备的初始几个手术，准备测试钻井深度和手动删除任何剩余的头骨碎片。的值可以被修改为将来手术（ 图16）。 </li><li>名称G代码，它会自动生成和SA粘弹性阻尼器的工作目录。 </li></ol></li><li>加载克码到数控铣床软件和测试颅骨插入立体定位仪器耳棒。 <ol><li>使用箭头键手动慢跑钻头前囟门。使用低速点动速度（〜5英寸/米），以确保准确性。 </li><li>启动钻头旋转时大于38,000转。 </li><li>按CycleStart;的stereotax将执行相同的切很多次传球，在不同深度。每遍之间，stereotax将暂停，所以外科医生可能会继续或中止切割。按继续循环 （按Alt-R）继续削减通行证。 </li></ol></li></ol>

<ol>
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作者没有竞争经济利益披露。

使用自动化手术设备有助于消除一些在神经科学研究中最常见的问题。首先，刀具路径是100％可重复的。每一个切割保证在相对位置前囟门一样。其次，它应该减少实验者错误。虽然许多研究人员都高度熟练的外科医生，它需要实践的一个特殊的量变得一个称职的外科医生。该设备将允许新的学生能够快速，轻松地进行高精度的手术。第三，电动手术设备应减少执行一个实验6所需的动物?...

立体定向手术啮齿动物被用在各种各样的神经科学应用，包括lesioning 1，离子电渗疗法2，微丝植入3，刺激4，和薄颅骨成像5。不过，也有面向那些谁想要应用这些技术，包括用于执行精确的立体定向手术的陡峭的学习曲线和人为错误的几率高主要障碍。人为错误包括测量和计算失误，以及低精度和可复制人类的动作。在努力减少这些干扰错误，立体定向外科医生将受益于一个系统，它可以确保所有的外科手术跨学科完全相同进行。错误的减少，也是一种方法，通过它可以调查尽量少用动物学科，美国国立卫生研究院的动物实验6的主要目标。在一个理想的世界里，所有的stereotactic手术将完全复制中的实验和实验室之间。为了解决这个问题，公司已经开发出新的超精密stereotaxics，和数字显示器读取测量结果。若要移除人体运动的错误，电动微型机器人和stereotaxics进行商业化生产，但其成本高，让人望而却步，以在有限的预算的实验室。此外，他们的软件是完全私有的，并且不能由研究者修改，以适应新类型的手术。 经济实惠的解决方案，以人为错误的问题是从实验室现有的模型建立一个机器人stereotax，使用行业标准的数控设备。因为一个新兴的数控爱好者社区，该材料​​是显著更便宜比科研设备。这允许一个建立一个准确的数控立体定位仪，这也是高度灵活和便宜。随着数控加工及G-Code，individu的基本知识ALS可以设定任何立体定向手术，他们想象，如果没有专有软件的限制。而且，为了加快生产的G代码进行简单的手术，这个协议包括软件，允许用户设计手术（锐边开颅手术，颅骨变薄窗口，钻孔和植入物降低）内的点，然后单击菜单。这些程序输出一个完整的G-代码，可以直接从CNC软件运行。 总之，机动立体升级是非常适合那些谁拥有权益的增加手术的准确性和可复制性，同时保留一个开源平台的灵活性和低成本。

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			<td height="44" style="height:44px;width:141px;">1x Standard U Frame Stereotax</td>
			<td style="width:137px;">Kopf</td>
			<td style="width:117px;"><a href="http://www.kopfinstruments.com/Stereotaxic/900.html">Kopf</a></td>
			<td style="width:355px;">This protocol should work with most existing stereotaxic devices.</td>
		</tr>
		<tr height="65">
			<td height="65" style="height:65px;width:141px;">3x 12 V, 1.6 A, 233 oz-inch Geared Bipolar Stepper Motor</td>
			<td style="width:137px;">Phidgets</td>
			<td style="width:117px;"><a href="http://www.robotshop.com/geared-bipolar-stepper-motor-3v-233-oz-in.html">Robot Shop</a></td>
			<td style="width:355px;">Any high torque geared stepper motor should do.&#160;</td>
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		<tr height="73">
			<td height="73" style="height:73px;width:141px;">1x 3 Axis CNC Stepper Motor Driver Board Controller</td>
			<td style="width:137px;">Toshiba</td>
			<td style="width:117px;"><a href="http://www.ebay.com/itm/CNC-Router-3-Axis-TB6560-3-5A-Stepper-Motor-Driver-Board-For-Engraving-Machine-/300981888819">Ebay</a></td>
			<td style="width:355px;">Any 3 Axis CNC driver should do. Linked Item includes Mach3 CNC software.&#160;</td>
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		<tr height="94">
			<td height="94" style="height:94px;width:141px;">2x Arm Couplers: medial-lateral (ML) &#38; dorsal-ventral (DV)</td>
			<td style="width:137px;">custom machined</td>
			<td style="width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/3D%20Models%20-%20CNC%20Stereotax%20Parts.zip">Part Drawings</a></td>
			<td style="width:355px;">These must be machined by your local machine shop. (costs will vary)</td>
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		<tr height="76">
			<td height="76" style="height:76px;width:141px;">1x anterior-posterior (AP) Coupler</td>
			<td style="width:137px;">custom machined</td>
			<td style="width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/3D%20Models%20-%20CNC%20Stereotax%20Parts.zip">Part Drawings</a></td>
			<td style="width:355px;">These must be machined by your local machine shop. (costs will vary)</td>
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			<td height="172" rowspan="3" style="height:172px;width:141px;">3x Motor to Stereotax Collar</td>
			<td rowspan="3" style="width:137px;">custom machined</td>
			<td style="width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/3D%20Models%20-%20CNC%20Stereotax%20Parts.zip">Part Drawings</a></td>
			<td rowspan="3" style="width:355px;">These must be machined by your local machine shop. (costs will vary)</td>
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		<tr height="23">
			<td height="23" style="height:23px;width:117px;"></td>
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		<tr height="22">
			<td height="22" style="height:22px;width:117px;"><a href="http://eden.rutgers.edu/~mrcoffey/images/Collar.html">View in Browser</a></td>
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		<tr height="43">
			<td height="65" rowspan="2" style="height:65px;width:141px;">12x NF10-32 Cup Point Set Screws</td>
			<td rowspan="2" style="width:137px;">McMaster Carr</td>
			<td style="width:117px;"><a href="http://www.mcmaster.com/#92313A829">&#189;&#8221; Length</a></td>
			<td rowspan="2" style="width:355px;">You will need 6 of each.</td>
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			<td height="22" style="height:22px;width:117px;"><a href="http://www.mcmaster.com/#92313A825">&#188;&#8221; Length</a></td>
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			<td height="46" style="height:46px;width:141px;">12x M3 Socket Head Screws (20 mm)</td>
			<td style="width:137px;">McMaster Carr</td>
			<td style="width:117px;"><a href="http://www.mcmaster.com/#92855A319">20mm Length</a></td>
			<td style="width:355px;">You will need 4 for each motor</td>
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			<td height="48" style="height:48px;width:141px;">1x Micro-Motor Drill&#160;</td>
			<td style="width:137px;">Buffalo Dental</td>
			<td style="width:117px;"><a href="http://www.buffalodental.com/pdfnew/x50.pdf">X50</a></td>
			<td style="width:355px;">Any Micromotor drill will work.&#160; At least 38,000 rpm recommended</td>
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			<td height="53" style="height:53px;width:141px;">1x 12 V DC Power Supply</td>
			<td style="width:137px;">12 Volt Adapters</td>
			<td style="width:117px;"><a href="http://www.12vadapters.com/adapter/power-supply/12v/7-amp-7a.html">12v DC &#8211; 7 Amp</a></td>
			<td style="width:355px;">Any 12 V DC PSU should work (ensure amperage rating is higher than the sum of the motors&#8217; amperage).</td>
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			<td height="44" style="height:44px;width:141px;">1x Extra Large Probe Holder</td>
			<td style="width:137px;">Stoelting</td>
			<td><a href="http://www.stoeltingco.com/neuroscience/accessories/stereotaxic-probe-holders/extra-large-probe-holder-551.html">Stoelting</a></td>
			<td style="width:355px;"></td>
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			<td height="44" style="height:44px;width:141px;">1x Grade B Rat Skull</td>
			<td style="width:137px;">Skulls Unlimited</td>
			<td><a href="http://www.skullsunlimited.com/record_variant.php?id=4009">Skulls Unlimited</a></td>
			<td style="width:355px;"></td>
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			<td height="21" style="height:21px;width:141px;">Mach 3 Mill</td>
			<td style="width:137px;">ArtSoft USA</td>
			<td style="width:117px;"><a href="http://www.machsupport.com/ccount/click.php?id=1">Trial Download</a></td>
			<td style="width:355px;">Any Standard CNC controlling software should work.</td>
		</tr>
		<tr height="44">
			<td height="44" style="height:44px;width:141px;">Surgery Designer</td>
			<td style="width:137px;">Kevin Coffey &#38; David Barker</td>
			<td style="width:117px;"><a href="http://www.mathworks.com/matlabcentral/fileexchange/40282-robot-assisted-stereotactic-surgery">MATLAB File Exchange</a></td>
			<td style="width:355px;">These codes are available to modify. We accept no responsibility for your use or modification of code.</td>
		</tr>
	</tbody>
</table>

building an open-source robotic stereotaxic instrument

该协议包括必要的设计和软件升级现有立体定位仪的机器人（CNC）立体定位仪为大约$ 1,000（不包括钻头），使用行业标准的步进电机和数控控制软件。每个轴具有可变速度控制，并且可以同时地或独立地操作。机器人的灵活性和开放的编码系统（G代码），使其能够执行不支持由商业系统的自定义任务。其应用包括，但不限于，钻孔，尖锐边缘开颅手术，颅骨变薄，并降低电极或插管。为了加快G-编码的简单手术的写作，我们已经开发的自定义脚本，允许个人设计一个手术，没有编程知识。然而，为用户获得最大的电动stereotax的，这将是有益的知识渊博的数学规划和G-编码（简单的编冲压数控加工）。 建议钻孔速度大于40000转。步进电机的分辨率为1.8°/步，面向0.346°/步。标准stereotax有2.88微米/步的分辨率。建议的最高切割速度为500微米/秒。建议的最高点动速度是3500微米/秒。建议的最高钻头大小是惠普2。

设计中的方法，手术的最终结果将是一只老鼠颅骨用锋利的边缘开颅手术，以及3骷髅洞（ 图17）。注意，用来演示手术颅骨比原型大鼠颅骨宽得多。锋利的边缘开颅可用于插入一个微丝阵列进入大脑，对于高密度的神经录音。数控stereotax也可用于降低非常精确阵列。软件包含在本协议中，允许外科医生来定义微丝的参数，或套管降低。锋利的边缘开颅手术也可用于揭示运动皮层的大?...

Watch this Scientific Journal Video about Building An Open-source Robotic Stereotaxic Instrument at JoVE.com

Building An Open-source Robotic Stereotaxic Instrument

该协议包括了设计和必要的软件来升级现有的立体定位仪的机器人（控制计算机数值; CNC）立体定位仪在1000美元左右（不包括钻头）。

建设一个开放源码的机器人立体定位仪

This protocol includes the designs and software necessary to upgrade an existing stereotaxic instrument to a robotic (CNC) stereotaxic instrument for ...

building-an-open-source-robotic-stereotaxic-instrument

Research

JoVE Journal

Neuroscience

14.8K 次观看.  Rutgers, The State University of New Jersey. 该协议包括了设计和必要的软件来升级现有的立体定位仪的机器人（控制计算机数值; CNC）立体定位仪在1000美元左右（不包括钻头）。 

视频: Building An Open-source Robotic Stereotaxic Instrument

Rodent Stereotaxic Surgery and Animal Welfare Outcome Improvements for Behavioral Neuroscience

Stereotaxic surgery for the implantation of cannulae into specific brain regions has for many decades been a very successful experimental technique to investigate the effects of locally manipulated neurotransmitter and signaling pathways in awake, behaving animals. Moreover, the stereotaxic implantation of electrodes for electrophysiological stimulation and recording studies has been instrumental to our current understanding of neuroplasticity and brain networks in behaving animals. Ever-increasing knowledge about optimizing surgical techniques in rodents1-4, public awareness concerning animal welfare issues and stringent legislation (e.g., the 2010 European Union Directive on the use of laboratory animals5) prompted us to refine these surgical procedures, particularly with respect to implementing new procedures for oxygen supplementation and the continuous monitoring of blood oxygenation and heart rate levels during the surgery as well as introducing a standardized protocol for post-surgical care. Our observations indicate that these modifications resulted in an increased survival rate and an improvement in the general condition of the animals after surgery (e.g. less weight loss and a more active animal). This video presentation will show the general procedures involved in this type of stereotaxic surgery with special attention to our several modifications. We will illustrate these surgical procedures in rats, but it is also possible to perform this type of surgery in mice or other small laboratory animals by using special adaptors for the stereotaxic apparatus6.

Stereotaxic surgery on rodents allows for targeted administration of drugs or electrical stimulation and recordings in awake, behaving animals. In this video presentation we will demonstrate recent procedural refinements to this long-standing procedure that successfully improved survival rate and reduced post-surgical weight loss.

啮齿动物的立体定向手术和动物福利成果改进行为神经科学

Stereotaxic surgery for the implantation of cannulae into specific brain regions has for many decades been a very successful experimental technique to ...

科研

神经科学

Stereotaxic Surgery for Excitotoxic Lesion of Specific Brain Areas in the Adult Rat

Many behavioral functions in mammals, including rodents and humans, are mediated principally by discrete brain regions. A common method for discerning the function of various brain regions for behavior or other experimental outcomes is to implement a localized ablation of function. In humans, patient populations with localized brain lesions are often studied for deficits, in hopes of revealing the underlying function of the damaged area. In rodents, one can experimentally induce lesions of specific brain regions.
Lesion can be accomplished in several ways. Electrolytic lesions can cause localized damage but will damage a variety of cell types as well as traversing fibers from other brain regions that happen to be near the lesion site. Inducible genetic techniques using cell-type specific promoters may also enable site-specific targeting. These techniques are complex and not always practical depending on the target brain area. Excitotoxic lesion using stereotaxic surgery, by contrast, is one of the most reliable and practical methods of lesioning excitatory neurons without damaging local glial cells or traversing fibers.
Here, we present a protocol for stereotaxic infusion of the excitotoxin, N-methyl-D-aspartate (NMDA), into the basolateral amygdala complex. Using anatomical indications, we apply stereotaxic coordinates to determine the location of our target brain region and lower an injection needle in place just above the target. We then infuse our excitotoxin into the brain, resulting in excitotoxic death of nearby neurons. While our experimental subject of choice is a rat, the same methods can be applied to other mammals, with the appropriate adjustments in equipment and coordinates.
This method can be used on a variety of brain regions, including the basolateral amygdala1-6, other amygdala nuclei6, 7, hippocampus8, entorhinal cortex9 and prefrontal cortex10. It can also be used to infuse biological compounds such as viral vectors1, 11. The basic stereotaxic technique could also be adapted for implantation of more permanent osmotic pumps, allowing more prolonged exposure to a compound of interest.

Targeted ablation of specific brain region(s) by infusion of an excitotoxin using stereotaxic coordinates is described. This technique could also be adapted for infusion of other chemicals into the rat brain.

特定脑区的兴奋毒性损伤的成年大鼠立体定向手术治疗

Many behavioral functions in mammals, including rodents and humans, are mediated principally by discrete brain regions. A common method for discerning the ...

Stereotaxic Infusion of Oligomeric Amyloid-beta into the Mouse Hippocampus

Alzheimer&#8217;s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the over-production of amyloid-beta and the deposition of amyloid-beta plaques in brain regions of the afflicted individuals. Throughout the years scientists have generated numerous Alzheimer&#8217;s disease mouse models that attempt to replicate the amyloid-beta pathology. Unfortunately, the mouse models only selectively mimic the disease features. Neuronal death, a prominent effect in the brains of Alzheimer&#8217;s disease patients, is noticeably lacking in these mice. Hence, we and others have employed a method of directly infusing soluble oligomeric species of amyloid-beta - forms of amyloid-beta that have been proven to be most toxic to neurons - stereotaxically into the brain. In this report we utilize male C57BL/6J mice to document this surgical technique of increasing amyloid-beta levels in a select brain region. The infusion target is the dentate gyrus of the hippocampus because this brain structure, along with the basal forebrain that is connected by the cholinergic circuit, represents one of the areas of degeneration in the disease. The results of elevating amyloid-beta in the dentate gyrus via stereotaxic infusion reveal increases in neuron loss in the dentate gyrus within 1 week, while there is a concomitant increase in cell death and cholinergic neuron loss in the vertical limb of the diagonal band of Broca of the basal forebrain. These effects are observed up to 2 weeks. Our data suggests that the current amyloid-beta infusion model provides an alternative mouse model to address region specific neuron death in a short-term basis. The advantage of this model is that amyloid-beta can be elevated in a spatial and temporal manner.

Here, we present a protocol for direct stereotaxic brain infusion of amyloid-beta. This methodology provides an alternative in vivo mouse model to address the short-term effects of amyloid-beta on brain neurons.

低聚淀粉样蛋白-β的立体灌注到小鼠海马

Alzheimer&#8217;s disease is a neurodegenerative disease affecting the aging population. A key neuropathological feature of the disease is the ...

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method described here enables the investigator to measure long-lasting (from minutes to hours) high-quality intracellular responses from single Drosophila R1-R6 photoreceptors and Large Monopolar Cells (LMCs) to light stimuli. Because the recording system has low noise, it can be used to study variability among individual cells in the fly eye, and how their outputs reflect the physical properties of the visual environment. We outline all key steps in performing this technique. The basic steps in constructing an appropriate electrophysiology set-up for recording, such as design and selection of the experimental equipment are described. We also explain how to prepare for recording by making appropriate (sharp) recording and (blunt) reference electrodes. Details are given on how to fix an intact fly in a bespoke fly-holder, prepare a small window in its eye and insert a recording electrode through this hole with minimal damage. We explain how to localize the center of a cell's receptive field, dark- or light-adapt the studied cell, and to record its voltage responses to dynamic light stimuli. Finally, we describe the criteria for stable normal recordings, show characteristic high-quality voltage responses of individual cells to different light stimuli, and briefly define how to quantify their signaling performance. Many aspects of the method are technically challenging and require practice and patience to master. But once learned and optimized for the investigator's experimental objectives, it grants outstanding in vivo neurophysiological data.

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Sharp microelectrodes enable accurate electrophysiological characterization of photoreceptor and visual interneuron output in living Drosophila. Here we show how to use this method to record high-quality voltage responses of individual cells to controlled light stimulation. This method is ideal for studying neural information processing in insect compound eyes.

为记录细胞内电压响应电法<em&gt;果蝇</em&gt;光感受器和中间对光刺激<em&gt;在体内</em

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method ...

Non-restraining EEG Radiotelemetry: Epidural and Deep Intracerebral Stereotaxic EEG Electrode Placement

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and neurodegenerative diseases as well as epilepsies. This powerful technique does not only provide valuable insights into the underlying pathophysiological mechanisms, i.e., the etiopathogenesis of CNS related diseases, it also facilitates the development of new translational, i.e., therapeutic approaches. Whereas competing techniques that make use of recorder systems used in jackets or tethered systems suffer from their unphysiological restraining to semi-restraining character, radiotelemetric EEG recordings overcome these disadvantages. Technically, implantable EEG radiotelemetry allows for precise and highly sensitive measurement of epidural and deep, intracerebral EEGs under various physiological and pathophysiological conditions. First, we present a detailed protocol of a straight forward, successful, quick and efficient technique for epidural (surface) EEG recordings resulting in high-quality electrocorticograms. Second, we demonstrate how to implant deep, intracerebral EEG electrodes, e.g., in the hippocampus (electrohippocampogram). For both approaches, a computerized 3D stereotaxic electrode implantation system is used. The radiofrequency transmitter itself is implanted into a subcutaneous pouch in both mice and rats. Special attention also has to be paid to pre-, peri- and postoperative treatment of the experimental animals. Preoperative preparation of mice and rats, suitable anesthesia as well as postoperative treatment and pain management are described in detail.

Non-restraining EEG radiotelemetry is a valuable methodological approach to record in vivo long-term electroencephalograms from freely moving rodents. This detailed protocol describes stereotaxic epidural and deep intracerebral electrode placement in different brain regions in order to obtain reliable recordings of CNS rhythmicity and CNS related behavioral stages.

非约束脑电遥测:硬膜外和脑深部脑电立体定位放置电极

Implantable EEG radiotelemetry is of central relevance in the neurological characterization of transgenic mouse models of neuropsychiatric and ...

Automated Analysis of C. elegans Swim Behavior Using CeleST Software

Dissecting the neuronal and neuromuscular circuits that regulate behavior remains a major challenge in biology. The nematode Caenorhabditis elegans has proven to be an invaluable model organism in helping to tackle this challenge, from inspiring technological approaches,&#160;building the human brain connectome, to actually shedding light on the specific molecular drivers of basic functional patterns. The bulk of the behavioral studies in C. elegans have been performed on solid substrates. In liquid, animals exhibit behavioral patterns that include movement at a range of speeds in 3D, as well as partial body movements, such as a posterior curl without anterior shape change, which introduce new challenges for quantitation. The steps of a simple procedure, and use of a software that enables high-resolution analysis of C. elegans swim behavior, are presented here. The software, named CeleST, uses a specialized computer program that tracks multiple animals simultaneously and provides novel measures of C. elegans locomotion in liquid (swimming). The measures are mostly grounded in animal posture and based on mathematics used in computer vision and pattern recognition, without computational requirements for threshold cut-offs. The software tool can be used to both assess overall swimming prowess in hundreds of animals from combined small batch trials and to reveal novel phenotypes even in well-characterized genetic mutants. The preparation of specimens for analysis with CeleST is simple and low-tech, enabling wide adaptation by the scientific community. Use of the computational approach described here should therefore contribute to the greater understanding of behavior and behavioral circuits in the C. elegans model.

Automated Analysis of C. elegans Swim Behavior Using CeleST Software

An efficient and simple methodology for computer-based analysis of nematode swimming behavior in liquid is described. The method requires little to no investment for C. elegans laboratories. The hardware used is standard, and the computer software for the behavioral analysis (CeleST) is an open source one.

的自动分析<em&gt;℃。线虫</em&gt;游泳行为使用天书软件

Dissecting the neuronal and neuromuscular circuits that regulate behavior remains a major challenge in biology. The nematode Caenorhabditis elegans has ...

High-density Electroencephalographic Acquisition in a Rodent Model Using Low-cost and Open-source Resources

Advanced electroencephalographic analysis techniques requiring high spatial resolution, including electrical source imaging and measures of network connectivity, are applicable to an expanding variety of questions in neuroscience. Performing these kinds of analyses in a rodent model requires higher electrode density than traditional screw electrodes can accomplish. While higher-density electroencephalographic montages for rodents exist, they are of limited availability to most researchers, are not robust enough for repeated experiments over an extended period of time, or are limited to use in anesthetized rodents.1-3 A proposed low-cost method for constructing a durable, high-count, transcranial electrode array, consisting of bilaterally implantable headpieces is investigated as a means to perform advanced electroencephalogram analyses in mice or rats.

Procedures for headpiece fabrication and surgical implantation necessary to produce high signal to noise, low-impedance electroencephalographic and electromyographic signals are presented. While the methodology is useful in both rats and mice, this manuscript focuses on the more challenging implementation for the smaller mouse skull. Freely moving mice are only tethered to cables via a common adapter during recording. One version of this electrode system that includes 26 electroencephalographic channels and 4 electromyographic channels is described below.

Instructions for the low-cost construction and surgical implantation of a chronic transcranial high-density electroencephalographic montage into mice are provided. Signal recording, extraction, and processing techniques are also described.

高密度的脑电图采集在啮齿动物模型使用低成本的开源资源

Advanced electroencephalographic analysis techniques requiring high spatial resolution, including electrical source imaging and measures of network ...

Stereotaxic Surgery for Genetic Manipulation in Striatal Cells of Neonatal Mouse Brains

Many genes are expressed in embryonic brains, and some of them are continuously expressed in the brain after birth. For such persistently expressed genes, they may function to regulate the developmental process and/or physiological function in neonatal brains. To investigate neurobiological functions of specific genes in the brain, it is essential to inactivate genes in the brain. Here, we describe a simple stereotaxic method to inactivate gene expression in the striatum of transgenic mice at neonatal time windows. AAV-eGFP-Cre viruses were microinjected into the striatum of Ai14 reporter gene mice at postnatal day (P) 2 by stereotaxic brain surgery. The tdTomato reporter gene expression was detected in P14 striatum, suggesting a successful Cre-loxP mediated DNA recombination in AAV-transduced striatal cells. We further validated this technique by microinjecting AAV-eGFP-Cre viruses into P2Foxp2fl/fl mice. Double labeling of GFP and Foxp2 showed that GFP-positive cells lacked Foxp2 immunoreactivity in P9 striatum, suggesting the loss of Foxp2 protein in AAV-eGFP-Cre transduced striatal cells. Taken together, these results demonstrate an effective genetic deletion by stereotaxically microinjected AAV-eGFP-Cre viruses in specific neuronal populations in the neonatal brains of floxed transgenic mice. In conclusion, our stereotaxic technique provides an easy and simple platform for genetic manipulation in neonatal mouse brains. The technique can not only be used to delete genes in specific regions of neonatal brains, but it also can be used to inject pharmacological drugs, neuronal tracers, genetically modified optogenetics and chemogenetics proteins, neuronal activity indicators and other reagents into the striatum of neonatal mouse brains.

We describe a protocol of stereotaxic surgery with a homemade head-fixed device for microinjecting reagents into the striatum of neonatal mouse brains. This technique allows genetic manipulation in neuronal cells of specific regions of neonatal mouse brains.

新生小鼠脑巴细胞基因调控的立体定向手术

Many genes are expressed in embryonic brains, and some of them are continuously expressed in the brain after birth. For such persistently expressed genes, ...

An Alternative Approach to Study Primary Events in Neurodegeneration Using Ex Vivo Rat Brain Slices

Despite&#160;numerous studies that attempt to develop reliable animal models&#160;which reflecting the primary processes underlying neurodegeneration, very few have been widely accepted. Here, we propose&#160;a new procedure&#160;adapted from the well-known ex vivo brain slice technique, which offers a closer in vivo-like scenario than in vitro preparations, for investigating the early events triggering cell degeneration, as observed in Alzheimer&#39;s disease (AD). This variation consists of simple and easily reproducible steps, which enable preservation of the anatomical cytoarchitecture of the selected brain region and its local functionality in a physiological milieu. Different anatomical areas can be obtained from the same brain, providing the opportunity to perform multiple experiments with the treatments in question in a site-, dose-, and time-dependent manner. Potential limitations&#160;which could affect the outcomes related to this methodology are related to the conservation of the tissue, i.e., the maintenance of its anatomical integrity during the slicing and incubation steps and the section thickness, which can influence the biochemical and immunohistochemical analysis. This approach can be employed for different purposes, such as exploring molecular mechanisms involved in physiological or pathological conditions, drug screening, or dose-response assays. Finally, this protocol could also reduce the number of animals employed in behavioral studies. The application reported here has been recently described and tested for the first time on ex vivo rat brain slices containing the basal forebrain (BF), which is one of the cerebral regions primarily affected in AD. Specifically, it has been demonstrated that the administration of a toxic peptide derived from the C-terminus of acetylcholinesterase (AChE) could prompt an AD-like profile, triggering, along the antero-posterior axis of the BF, a differential expression of proteins altered in AD, such as the alpha7 nicotinic receptor (&#945;7-nAChR), phosphorylated Tau (p-Tau), and amyloid beta (A&#946;).

An Alternative Approach to Study Primary Events in Neurodegeneration Using Ex Vivo Rat Brain Slices

We present a method which can&#160;provide further insights into the early events underlying neurodegeneration and based on the established ex-vivo brain technique, combining the advantages of in vivo and in vitro experiments. Moreover, it represents a unique opportunity for direct comparison of treated and untreated group in the same anatomical plane.

利用体外大鼠脑切片研究变性主要事件的一种替代方法

Despite&#160;numerous studies that attempt to develop reliable animal models&#160;which reflecting the primary processes underlying neurodegeneration, ...

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset (Callithrix jacchus)

Marmosets (Callithrix jacchus) are small non-human primates that are gaining popularity in biomedical and preclinical research, including the neurosciences. Phylogenetically, these animals are much closer to humans than rodents. They also display complex behaviors, including a wide range of vocalizations and social interactions. Here, an effective stereotaxic neurosurgical procedure for implantation of recording electrode arrays in the common marmoset is described. This protocol also details the pre- and postoperative steps of animal care that are required to successfully perform such a surgery. Finally, this protocol shows an example of local field potential and spike activity recordings in a freely behaving marmoset 1 week after the surgical procedure. Overall, this method provides an opportunity to study the brain function in awake and freely behaving marmosets. The same protocol can be readily used by researchers working with other small primates. In addition, it can be easily modified to allow other studies requiring implants, such as stimulating electrodes, microinjections, implantation of optrodes or guide cannulas, or ablation of discrete tissue regions.

Stereotaxic Surgery for Implantation of Microelectrode Arrays in the Common Marmoset Callithrix jacchus

This work presents a protocol to perform a stereotaxic, neurosurgical implantation of&#160;microelectrode arrays in the common marmoset. This method specifically enables electrophysiological recordings in freely behaving animals but can be easily adapted to any other similar neurosurgical intervention in this species (e.g., cannula for drug administration or electrodes for brain stimulation).