作者还希望表达感谢克里斯刻录机，斯蒂芬·麦金尼，劳拉·托宾，珍妮Gilbreath，阿什利日Olley，梅根·奥费，以及梅根海德他们在微调工作，此法。

1。苍蝇的步态分析个人的制备<ol><li>转让苍蝇轻轻拍打成单独的空瓶子和盖子的小瓶用棉塞。让苍蝇品静置20-30分钟观察之前。它不麻醉前行为观察的时间内苍蝇与以前的研究发现麻醉曝光可以改变的行为相比，非麻醉苍蝇14是非常重要的。 </li><li>轻轻拍打飞出来的小瓶，然后将个别苍蝇在一个直径为5厘米的培养皿盖（5毫米高）的小缝隙，让空气通过。从下面照射并利用安装在上面的培养皿一个摄像头观察苍蝇。 </li></ol> 2。苍蝇扣押分析个人的制备<ol><li>喂两天老扣押敏感的苍蝇任何一种标准，酵母/玉米粉/琼脂培养基或标准介质直接混合药物。对于数据呈现在这里，苍蝇喂养或者1合F标准的媒体或1 g和二甲双胍25毫克混合标准介质。 </li><li>之后，苍蝇喂了两天的媒体或媒体加药，轻轻敲击将他们转移到个人的空瓶子和盖子的小瓶用棉塞。让苍蝇静置20分钟前观察。它不麻醉前行为观察的时间内苍蝇与以前的研究发现麻醉曝光可以改变的行为相比，非麻醉苍蝇14是非常重要的。 </li><li>含在实验室涡流混合器单一使用飞10秒的最高设置旋涡个人小瓶。将固定苍蝇在纸张的正下方安装在纸上面的一个摄像头一个空白页。 </li></ol> 3。视频录制<ol><li>记录使用HandyAvi软件程序的移动或SLA <a href="http://www.azcendant.com/download.htm">http://www.azcendant.com/download.htm</a> 。这个程序使用网络摄像头捕获基于用户设置的图像。 </li><li>选择捕捉选项卡下的延时图像选项，当窗口打开时，选择摄像头的捕获设备。对于大多数实验的640×480的视频帧的大小是足够的。 </li><li>选择英特尔IYUV编解码器的压缩和选择0.1秒/帧检化验或选择0.1〜0.5秒/帧的运动检测。在高级选项卡上，选择为每个捕获创建。bmp图像文件，并选择一个文件夹为程序存储，它需要（图像堆栈）的图像。 </li><li>放置纸张上一只死苍蝇。点击视频设置框并调整设置的设备设置选项卡下，这样有一个明亮的白色背景，并与暗飞了明显的反差。 </li><li>单击开始按钮，开始录制。一旦记录周期完成后，请单击停止按钮。图像堆栈现在应该选择的文件夹中。注:在录制之前，确保FOL德选择的图像堆栈是空的。 </li></ol> 4。使用ImageJ数据分析<ol><li>美国国立卫生研究院开放的免费图像处理软件ImageJ的，可从以下网址下载<a href="http://rsbweb.nih.gov/ij/download.html">http://rsbweb.nih.gov/ij/download.html</a> 。注:除了下载该程序，多轨插件，必须下载并安装于所示<a href="http://rsbweb.nih.gov/ij/plugins/index.html">http://rsbweb.nih.gov/ij/plugins/index.html</a> 。 </li><li>导入图像堆栈成的ImageJ通过点击文件选项卡，然后选择导入→图像序列。 <ol><li>选择包含​​图像堆栈的文件夹，突出图像中的一个，并选择开启。序列选项对话框将打开。点击排序数值名称，然后选择确定和图像堆栈将被加载到ImageJ的。 </li></ol></li><li>门槛都让每一个由一个白色的背景，是苍蝇的图像转换到一个黑点。要开始这个过程中，图像通过图像选项卡下选择类型→8位转换为8位格式。 <ol><li>再次单击图像选项卡上，选择调整→阈值。这将打开阈值对话框中，阈值截断点可以调整。 </li><li>通过扫描图像，看是否有多余的黑点，或者如果切片缺少一个黑点。如果是这种情况，调整阈值，以缓解该问题。通常情况下，阈值并不需要进行调整，因为程序默认为一个适当的值。 注:在堆栈中的每一张幻灯片应该只有一个黑点代表苍蝇的位置。 </li><li>选择适用于该阈值对话框和转换到面膜对话框将打开。不要单击任何选项，但只需点击OK阈值图像堆栈。 </li></ol></li><li>阈值化图像堆栈后，选择多轨插件在插件选项卡下，以测量苍蝇的运动以像素为单位。将出现对象跟踪对话框。 <ol><li>点击所有的四个选项在对话框中给定的，然后选择确定。在多轨插件，然后将附加坐标转换为黑点中的每个切片并在结果窗口，以及以像素为单位的总路径长度列出这些。路径窗口给出了飞了路径的图形显示。 </li><li> x和y坐标列表从结果窗口复制和粘贴数据到Excel电子表格。如果在堆栈中的任何幻灯片没有一个黑点在它的多轨程序将停止在该幻灯片。如果发生这种情况，就必须返回步骤4.2和重新阈堆栈，以确保在飞行寄存器作为一个黑点在所述的滑动。 </li></ol></li><li>为了转换像素的路径长度，以厘米，采取的是一个空白的白纸间隔开cm处两个点的单独图像。 <ol><li>遵循以上步骤（3.1-4.4.2），以确定在x，y像素坐标的两个点，并使用该信息来确定对应于一厘米的像素的数目。这个转换系数被用在步骤5，以产生在厘米运动数据。注意:一旦做到这一点，保持相机固定以后所有的记录是很重要的。每次摄像机移动或调整，需要重复此步骤，以像素转换为厘米。 </li><li>是由多轨插件生成的路径长度高估运动，因为在身体位置和光量/反射小幅波动会影响该阈值化的图像时产生的黑点的大小和中心。这种噪声必须以获得飞运动的精确估计被除去。 </li></ol></li></ol> 5。使用Excel数据分析<ol><li>通过导入一系列的X从记录的数据去除噪声，Y的多轨公关产生的坐标ogram成飞分析，由作者创建的Excel Visual Basic程序。 （该程序可通过联系作者。） </li><li>点击Click Me按钮，然后选择一个阈值截止。对于SLA分析选择为0.5厘米/秒的阈值，或者选择0.1-0.3厘米/秒之间的值作为运动分析。单击确定，程序将计算速度，时间和距离的运动。注:本程序采用一个滑动窗口分析，以评估在连续0.5-1.0秒箱中的数据和该窗口中计算速度。任何用户定义的截止频率以上的被视为动态运动。 </li></ol>

<ol>
	<li>Stone, B., Evans, L., Coleman, J., Kuebler, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Genetic+and+pharmacological+manipulations+that+alter+metabolism+suppress+seizure-like+activity+in.">Genetic and pharmacological manipulations that alter metabolism suppress seizure-like activity in.</a> 1496, 94-103 (2013).</li><li>Benzer, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+mutants+of+Drosophila+isolated+by+countercurrent+distribution.">Behavioral mutants of Drosophila isolated by countercurrent distribution.</a> Proc. Natl. Acad. Sci. U.S.A. 58, 1112-1119 (1967).</li><li>Slawson, J. B., Kim, E. Z., Griffith, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Resolution+Video+Tracking+of+Locomotion+in+Adult+Drosophila+melanogaster.">High-Resolution Video Tracking of Locomotion in Adult Drosophila melanogaster.</a> J. Vis. Exp. (24), (2009).</li><li>Spink, A. J., Tegelenbosch, R. A., Buma, M. O., Noldus, L. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+EthoVision+video+tracking+systema+tool+for+behavioral+phenotyping+of+transgenic+mice.">The EthoVision video tracking systema tool for behavioral phenotyping of transgenic mice.</a> Physiol. Behav. 73, 731-744 (2001).</li><li>Kohlhoff, K. J., Jahn, T. R., Lomas, D. A., Dobson, C. M., Crowther, D. C., Vendruscolo, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+iFly+tracking+system+for+an+automated+locomotor+and+behavioural+analysis+of+Drosophila+melanogaster.">The iFly tracking system for an automated locomotor and behavioural analysis of Drosophila melanogaster.</a> Integr. Biol. 3, 755-760 (2011).</li><li>Gomez-Marin, A., Partoune, N., Stephens, G. J., Louis, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Automated+tracking+of+animal+posture+and+movement+during+exploration+and+sensory+orientation+behaviors.">Automated tracking of animal posture and movement during exploration and sensory orientation behaviors.</a> PLoS ONE. 7, (2002).</li><li>Wolf, F. W., Rodan, A. R., Tsai, L. T., Heberlein, U. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Resolution+analysis+of+ethanol-induced+locomotor+stimulation+in+Drosophila.">High-Resolution analysis of ethanol-induced locomotor stimulation in Drosophila.</a> J. Neurosci. 22, 11035-11044 (2002).</li><li>Martin, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+portrait+of+locomotor+behaviour+in+Drosophila+determined+by+a+video-tracking+paradigm.">A portrait of locomotor behaviour in Drosophila determined by a video-tracking paradigm.</a> Behav. Processes. 67, 207-219 (2004).</li><li>Meunier, N., Belgacem, Y. H., Martin, J. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Regulation+of+feeding+behaviour+and+locomotor+activity+by+takeout+in+Drosophila.">Regulation of feeding behaviour and locomotor activity by takeout in Drosophila.</a> J. Exp. Biol. 210, 1424-1434 (2007).</li><li>Koudounas, S., Green, E. W., Clancy, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Reliability+and+variability+of+sleep+and+activity+as+biomarkers+of+ageing+in+Drosophila.">Reliability and variability of sleep and activity as biomarkers of ageing in Drosophila.</a> Biogerontology. 13, 489-499 (2012).</li><li>Catterson, J. H., Knowles-Barley, S., James, K., Heck, M. M., Harmar, A. J., Hartley, P. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dietary+modulation+of+Drosophila+sleep-wake+behaviour.">Dietary modulation of Drosophila sleep-wake behaviour.</a> PLoS One. 5, (2010).</li><li>Donelson, N., Kim, E. Z., Slawson, J. B., Vecsey, C. G., Huber, R., Griffith, L. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=High-Resolution+positional+tracking+for+long-term+analysis+of+Drosophila+sleep+and+locomotion+using+the+''Tracker''+program.">High-Resolution positional tracking for long-term analysis of Drosophila sleep and locomotion using the ''Tracker'' program.</a> PLos ONE. 7, (2012).</li><li>Duboff, B., Fridovich-Keil, J. L. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Mediators+of+a+long-term+movement+abnormality+in+a+Drosophila+melanogaster+model+of+classic+galactosemia.">Mediators of a long-term movement abnormality in a Drosophila melanogaster model of classic galactosemia.</a> Dis. Model Mech. 5, 796-803 (2012).</li><li>Seiger, M. B., Kink, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+effect+of+anesthesia+on+the+photoresponses+of+four+sympatric+species+of+Drosophila.">The effect of anesthesia on the photoresponses of four sympatric species of Drosophila.</a> Behav. Genet. 23, 99-104 (1993).</li><li>Pavlidis, P., Ramaswami, M., Tanouye, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Drosophila+easily+shocked+gene:+a+mutation+in+a+phospholipid+synthetic+pathway+causes+seizure,+neuronal+failure,+and+paralysis.">The Drosophila easily shocked gene: a mutation in a phospholipid synthetic pathway causes seizure, neuronal failure, and paralysis.</a> Cell. 79, 2333 (1994).</li><li>Royden, C. S., Pirrotta, V., Jan, L. Y. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+tko+locus,+site+of+a+behavioral+mutation+in+D.+melanogaster,+codes+for+a+protein+homologous+to+prokaryotic+ribosomal+protein+S12.">The tko locus, site of a behavioral mutation in D. melanogaster, codes for a protein homologous to prokaryotic ribosomal protein S12.</a> Cell. 51, 165-173 (1987).</li><li>Parker, L., Padilla, M., Du, Y., Dong, K., Tanouye, M. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+as+a+model+for+epilepsy:+bss+is+a+gain-of-function+mutation+in+the+para+sodium+channel+gene+that+leads+to+seizures.">Drosophila as a model for epilepsy: bss is a gain-of-function mutation in the para sodium channel gene that leads to seizures.</a> Genetics. , 187-534 (2011).</li><li>Hardie, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Role+of+AMP-activated+protein+kinase+in+the+metabolic+syndrome+and+in+heart+disease.">Role of AMP-activated protein kinase in the metabolic syndrome and in heart disease.</a> FEBS Lett. 582, 81-89 (2008).</li><li>Fulgencio, J. P., Kohl, C., Girard, J., Pegorier, J. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Effect+of+metformin+on+fatty+acid+and+glucose+metabolism+in+freshly+isolated+hepatocytes+and+on+specific+gene+expression+in+cultured+hepatocytes.">Effect of metformin on fatty acid and glucose metabolism in freshly isolated hepatocytes and on specific gene expression in cultured hepatocytes.</a> Biochem. Pharma. 62, 439-446 (2001).</li></ol>

作者宣称，他们有没有竞争的财务权益。

当检查的运动或运动模式在果蝇中 ，它才能够提取关于移动的距离信息，运动的速度和运动模式是有用的。为了提取这些信息，昂贵的设备在传统上一直采用的往往是望而却步那些希望谨慎4,8,9使用这样的检测更小的实验室或实验室。 此处所描述的测定法可用于确定行驶的距离（ 图2和6）的运动，速度（ 图4和7）</st...

鉴于其寿命短和可用的健壮的遗传工具， 果蝇是用于研究各种疾病的病因学和各种生物过程中的潜在生理学极好的模型系统。在许多情况下，有利的是测量的影响，行为，遗传或药理操作对运动在这些模型有机体1,2。 有很多种方法，这些方法通常用于测量在两维3-7苍蝇的运动。这些系统可同时支持多个苍蝇的跟踪，并且可以测量速度，记录的路径长度，并记录时间的苍蝇已经使用了移动的百分比。它们已被用于研究在各种情况下，包括药物对运动和苍蝇的运动6-9的性二态性质的影响的运动。这些系统的主要缺点是跟踪系统的成本统或相应的软件和照相机。在某些情况下，这可以在数千美元运行。成本是一个实验室，将只需要有限的使用这样的系统，例如，量化新分离的突变体的机车图案的特别关注。 一个更简单的，但不太可靠的方法是使用其中记录的运动的基础上多少次穿过蝇被放置在闭合管10,11的中途的红外光束路径的系统。虽然这种系统可以提供运动和睡眠 - 觉醒周期的有价值的信息，他们可以超过或低于估计的运动，因为他们无法捕捉苍蝇的实际路径。例如，苍蝇，表现出在管的端部相当的运动将登记为低运动苍蝇虽然额外高分辨率的方法已被用来尝试规避这些限制12。 简单，便宜还都爬不动了测量geotaxi设备秒，苍蝇的向上运动，通过单管或一系列管2,13的。虽然这样的系统很便宜，可以很容易地识别趋地性的缺陷，他们无法捕捉到许多其他方面的运动，将感兴趣的研究者。 对于许多实验室，低成本的强大的分析系统，该系统简单的设置和操作将是一个有利的工具表征果蝇品系行为上的差异。在这里，我们描述了可以设置为不到两百元，并能提供有关的路径，速度苍蝇的运动和时间信息的检测。以证明该测定的功效，我们提出的数据表明，它可用于标识:1）在一个爆炸敏感（BS）突变体易受癫痫发作和药物二甲双胍2）的能力，一个自发的缺陷而通常用于治疗II型糖尿病，以减少发作样活性（SLA）Ⅰ的强度n两个学士学位突变体。

<table border="0" cellpadding="0" cellspacing="0" style="width:1117px;" width="1117">
	<colgroup>
		<col />
		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="68">
			<td height="68" style="height:68px;width:251px;">Name of the Reagent/Equipment</td>
			<td style="width:152px;">Company</td>
			<td style="width:169px;">Catalog Number/Model</td>
			<td style="width:545px;">Comments</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">WebCam</td>
			<td>Logitech</td>
			<td>Pro900</td>
			<td>Any quality webcam will suffice.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">HandiAVI time-lapse software</td>
			<td>Azcendant software</td>
			<td></td>
			<td>Latest version can be found at http://www.azcendant.com/</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">ImageJ</td>
			<td>NIH</td>
			<td></td>
			<td>Latest version can be found at http://rsbweb.nih.gov/ij/download.html</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Multiracker Plug-In</td>
			<td>NIH</td>
			<td></td>
			<td>Latest version can be found at http://rsbweb.nih.gov/ij/plugins/index.html</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Vortexer</td>
			<td>VWR</td>
			<td>Vortex Genie 2</td>
			<td>Most standard size vortexers such as the Vortex Genie 2 will suffice.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">5cm petri dish cover</td>
			<td>LabM Limited</td>
			<td>D011</td>
			<td>Smaller or larger petri dish covers can be used for an arena in movement assay.</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;">Light box</td>
			<td colspan="2">Built from scrap material.</td>
			<td>Illumination is used for the locomotion assay. Depending on the room lighting, it is possible to perform the assay without the light box.&#160;</td>
		</tr>
	</tbody>
</table>

a low-cost method for analyzing seizure-like activity and movement in drosophila

视频跟踪系统已被广泛地用于分析果蝇的运动和检测各种异常的机车行为。虽然这些系统可以提供丰富的行为信息，这些系统的成本和复杂性可能是令人望而却步的许多实验室。我们已经开发出用于测量果蝇机车行为，并扣押运动低成本的检测。该系统使用网络摄像捕获，可以使用廉价的和免费软件的组合来跟踪移动的距离被处理的图像，运动和运动的持续时间在指定的时间范围的平均流速。为了证明该系统的效用，我们研究了D组的果蝇突变体，该邦敏感（BS）瘫痪，这比野生型果蝇3-10倍更易患癫痫样活性（SLA）。使用这种新颖的系统，我们能够检测到该BS的Mutan吨的一声毫无意义的（BSS）具有探索性的运动在一个新的环境比野生型果蝇较低水平。此外，该系统被用来确定该药物二甲双胍，其通常被用于治疗II型糖尿病，降低了SLA的在BS中的突变体的强度。

这里所描述的技术已被以前用来分析在果蝇突变体学士学位1 SLA差异。这里给出的结果涉及的BS菌株容易感到震惊（EAS），轰的一声毫无意义的（BSS），以及技术性击倒（TKO）。在EAS基因编码参与磷脂酰乙醇胺合成15乙醇胺激酶和将军澳基因编码线粒体riboprotein 16。该BSS突变是麻痹 （ 对 ）电压门控钠通道17的等?...

Watch this Scientific Journal Video about A Low-cost Method for Analyzing Seizure-like Activity and Movement in Drosophila at JoVE.com

A Low-cost Method for Analyzing Seizure-like Activity and Movement in Drosophila

使用自由和廉价的软件程序的网络摄像头和相结合的，在果蝇机车图案可以被分析以检测速度，距离，以及各马达的活动时间的差别。

在分析癫痫样活动和运动的一种低成本方法<em&gt;果蝇</em

Video tracking systems have been used widely to analyze Drosophila melanogaster movement and detect various abnormalities in locomotive behavior. While ...

a-low-cost-method-for-analyzing-seizure-like-activity-movement

Research

JoVE Journal

Neuroscience

A Low-cost Method for Analyzing Seizure-like Activity and Movement in Drosophila

13.5K 次观看.  Franciscan University of Steubenville. 使用自由和廉价的软件程序的网络摄像头和相结合的，在果蝇机车图案可以被分析以检测速度，距离，以及各马达的活动时间的差别。 

视频: A Low-cost Method for Analyzing Seizure-like Activity and Movement in Drosophila

Assaying Locomotor Activity to Study Circadian Rhythms and Sleep Parameters in Drosophila

Most life forms exhibit daily rhythms in cellular, physiological and behavioral phenomena that are driven by endogenous circadian (&equiv;24 hr) pacemakers or clocks. Malfunctions in the human circadian system are associated with numerous diseases or disorders. Much progress towards our understanding of the mechanisms underlying circadian rhythms has emerged from genetic screens whereby an easily measured behavioral rhythm is used as a read-out of clock function. Studies using Drosophila have made seminal contributions to our understanding of the cellular and biochemical bases underlying circadian rhythms. The standard circadian behavioral read-out measured in Drosophila is locomotor activity. In general, the monitoring system involves specially designed devices that can measure the locomotor movement of Drosophila. These devices are housed in environmentally controlled incubators located in a darkroom and are based on using the interruption of a beam of infrared light to record the locomotor activity of individual flies contained inside small tubes. When measured over many days, Drosophila exhibit daily cycles of activity and inactivity, a behavioral rhythm that is governed by the animal's endogenous circadian system. The overall procedure has been simplified with the advent of commercially available locomotor activity monitoring devices and the development of software programs for data analysis. We use the system from Trikinetics Inc., which is the procedure described here and is currently the most popular system used worldwide. More recently, the same monitoring devices have been used to study sleep behavior in Drosophila. Because the daily wake-sleep cycles of many flies can be measured simultaneously and only 1 to 2 weeks worth of continuous locomotor activity data is usually sufficient, this system is ideal for large-scale screens to identify Drosophila manifesting altered circadian or sleep properties.

Assaying Locomotor Activity to Study Circadian Rhythms and Sleep Parameters in Drosophila

We describe procedures for recording daily locomotor activity rhythms of Drosophila and subsequent data analysis. Locomotor activity rhythms are a reliable behavioral output of animal circadian clocks and are used as the standard readout of clock function when studying circadian mutants or examining how the environment regulates the circadian system.

含量自发活动，研究昼夜节律和睡眠参数果蝇</em

Most life forms exhibit daily rhythms in cellular, physiological and behavioral phenomena that are driven by endogenous circadian (&equiv;24 hr) ...

科研

神经科学

A Low Cost Setup for Behavioral Audiometry in Rodents

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the experiments. Such parameters may be physiological response characteristics of the auditory pathway, e.g. via brainstem audiometry (BERA). But these methods allow only indirect and uncertain extrapolations about the auditory percept that corresponds to these physiological parameters. To assess the perceptual level of hearing, behavioral methods have to be used. A potential problem with the use of behavioral methods for the description of perception in animal models is the fact that most of these methods involve some kind of learning paradigm before the subjects can be behaviorally tested, e.g. animals may have to learn to press a lever in response to a sound. As these learning paradigms change perception itself 1,2 they consequently will influence any result about perception obtained with these methods and therefore have to be interpreted with caution. Exceptions are paradigms that make use of reflex responses, because here no learning paradigms have to be carried out prior to perceptual testing. One such reflex response is the acoustic startle response (ASR) that can highly reproducibly be elicited with unexpected loud sounds in na&iuml;ve animals. This ASR in turn can be influenced by preceding sounds depending on the perceptibility of this preceding stimulus: Sounds well above hearing threshold will completely inhibit the amplitude of the ASR; sounds close to threshold will only slightly inhibit the ASR. This phenomenon is called pre-pulse inhibition (PPI) 3,4, and the amount of PPI on the ASR gradually depends on the perceptibility of the pre-pulse. PPI of the ASR is therefore well suited to determine behavioral audiograms in na&iuml;ve, non-trained animals, to determine hearing impairments or even to detect possible subjective tinnitus percepts in these animals. In this paper we demonstrate the use of this method in a rodent model (cf. also ref. 5), the Mongolian gerbil (Meriones unguiculatus), which is a well know model species for startle response research within the normal human hearing range (e.g. 6).

A fast and inexpensive method for the behavioral determination of hearing parameters like hearing thresholds, hearing impairments or phantom perceptions (subjective tinnitus) is described. It uses pre-pulse inhibition of the acoustic startle response and can be easily implemented in a personal computer using a programmable AD/DA-converter and a piezo sensor.

在啮齿动物的行为测听的低成本设置

In auditory animal research it is crucial to have precise information about basic hearing parameters of the animal subjects that are involved in the ...

A Human-machine-interface Integrating Low-cost Sensors with a Neuromuscular Electrical Stimulation System for Post-stroke Balance Rehabilitation

A stroke is caused when an artery carrying blood from heart to an area in the brain bursts or a clot obstructs the blood flow to brain thereby preventing delivery of oxygen and nutrients. About half of the stroke survivors are left with some degree of disability. Innovative methodologies for restorative neurorehabilitation are urgently required to reduce long-term disability. The ability of the nervous system to reorganize its structure, function and connections as a response to intrinsic or extrinsic stimuli is called neuroplasticity. Neuroplasticity is involved in post-stroke functional disturbances, but also in rehabilitation. Beneficial neuroplastic changes may be facilitated with non-invasive electrotherapy, such as neuromuscular electrical stimulation (NMES) and sensory electrical stimulation (SES). NMES involves coordinated electrical stimulation of motor nerves and muscles to activate them with continuous short pulses of electrical current while SES involves stimulation of sensory nerves with electrical current resulting in sensations that vary from barely perceivable to highly unpleasant. Here, active cortical participation in rehabilitation procedures may be facilitated by driving the non-invasive electrotherapy with biosignals (electromyogram (EMG), electroencephalogram (EEG), electrooculogram (EOG)) that represent simultaneous active perception and volitional effort. To achieve this in a resource-poor setting, e.g., in low- and middle-income countries, we present a low-cost human-machine-interface (HMI) by leveraging recent advances in off-the-shelf video game sensor technology. In this paper, we discuss the open-source software interface that integrates low-cost off-the-shelf sensors for visual-auditory biofeedback with non-invasive electrotherapy to assist postural control during balance rehabilitation. We demonstrate the proof-of-concept on healthy volunteers.

A novel low-cost human-machine interface for interactive post-stroke balance rehabilitation system is presented in this article. The system integrates off-the-shelf low-cost sensors towards volitionally driven electrotherapy paradigm. The proof-of-concept software interface is demonstrated on healthy volunteers.

人机界面集成的低成本传感器，具有神经肌肉电刺激系统中风后康复平衡

A stroke is caused when an artery carrying blood from heart to an area in the brain bursts or a clot obstructs the blood flow to brain thereby preventing ...

A Simple and Inexpensive Method for Determining Cold Sensitivity and Adaptation in Mice

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life. The cold plantar assay allows the objective and inexpensive assessment of cold sensitivity in mice, and can quantify both analgesia and hypersensitivity. Mice are acclimated on a glass plate, and a compressed dry ice pellet is held against the glass surface underneath the hindpaw. The latency to withdrawal from the cooling glass is used as a measure of cold sensitivity.
Cold sensation is also important for survival in regions with seasonal temperature shifts, and in order to maintain sensitivity animals must be able to adjust their thermal response thresholds to match the ambient temperature. The Cold Plantar Assay (CPA) also allows the study of adaptation to changes in ambient temperature by testing the cold sensitivity of mice at temperatures ranging from 30 &#176;C to 5 &#176;C. Mice are acclimated as described above, but the glass plate is cooled to the desired starting temperature using aluminum boxes (or aluminum foil packets) filled with hot water, wet ice, or dry ice. The temperature of the plate is measured at the center using a filament T-type thermocouple probe. Once the plate has reached the desired starting temperature, the animals are tested as described above.
This assay allows testing of mice at temperatures ranging from innocuous to noxious. The CPA yields unambiguous and consistent behavioral responses in uninjured mice and can be used to quantify both hypersensitivity and analgesia. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

The Cold Plantar Assay (CPA) measures cold responsiveness between 30 &#176;C and 5 &#176;C, and can also measure cold adaptation. This protocol describes how to use the CPA to measure cold hypersensitivity, analgesia, and adaptation in mice.

测定冷灵敏度和适应小鼠一种简单廉价的方法

Cold hypersensitivity is a serious clinical problem, affecting a broad subset of patients and causing significant decreases in quality of life.  The cold ...

A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

Traumatic brain injury (TBI) affects millions of people each year, causing impairment of physical, cognitive, and behavioral functions and death. Studies using Drosophila have contributed important breakthroughs in understanding neurological processes. Thus, with the goal of understanding the cellular and molecular basis of TBI pathologies in humans, we developed the High Impact Trauma (HIT) device to inflict closed head TBI in flies. Flies subjected to the HIT device display phenotypes consistent with human TBI such as temporary incapacitation and progressive neurodegeneration. The HIT device uses a spring-based mechanism to propel flies against the wall of a vial, causing mechanical damage to the fly brain. The device is inexpensive and easy to construct, its operation is simple and rapid, and it produces reproducible results. Consequently, the HIT device can be combined with existing experimental tools and techniques for flies to address fundamental questions about TBI that can lead to the development of diagnostics and treatments for TBI. In particular, the HIT device can be used to perform large-scale genetic screens to understand the genetic basis of TBI pathologies.

A Method to Inflict Closed Head Traumatic Brain Injury in Drosophila

Here we describe a method to inflict closed head traumatic brain injury (TBI) in Drosophila. This method provides a gateway to investigate the cellular and molecular mechanisms that underlie TBI pathologies using the vast array of experimental tools and techniques available for flies.

方法对敌人造成闭合性颅脑创伤性脑损伤<em&gt;果蝇</em

Traumatic brain injury (TBI) affects millions of people each year, causing impairment of physical, cognitive, and behavioral functions and death. Studies ...

Direct-current Stimulation and Multi-electrode Array Recording of Seizure-like Activity in Mice Brain Slice Preparation

Cathodal transcranial direct-current stimulation (tDCS) induces suppressive effects on drug-resistant seizures. To perform effective actions, the stimulation parameters (e.g., orientation, field strength, and stimulation duration) need to be examined in mice brain slice preparations. Testing and arranging the orientation of the electrode relative to the position of the mice brain slice are feasible. The present method preserves the thalamocingulate pathway to evaluate the effect of DCS on anterior cingulate cortex seizure-like activities. The results of the multichannel array recordings indicated that cathodal DCS significantly decreased the amplitude of the stimulation-evoked responses and duration of 4-aminopyridine and bicuculline-induced seizure-like activity. This study also found that cathodal DCS applications at 15 min caused long-term depression in the thalamocingulate pathway. The present study investigates the effects of DCS on thalamocingulate synaptic plasticity and acute seizure-like activities. The current procedure can test the optimal stimulation parameters including orientation, field strength, and stimulation duration in an in vitro mouse model. Also, the method can evaluate the effects of DCS on cortical seizure-like activities at both the cellular and network levels.

Studies have shown that cathodal transcranial direct-current stimulation can produce suppressive effects on drug-resistant seizures. In this study, an in vitro experimental setup was devised in which the direct-current stimulation and multielectrode array recording of seizure-like activity were evaluated in mice brain slice preparation. The direct-current stimulation parameters were evaluated.

小鼠脑切片准备扣押样活性的直接电流刺激和多电极阵列记录

Cathodal transcranial direct-current stimulation (tDCS) induces suppressive effects on drug-resistant seizures. To perform effective actions, the ...

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 ...

Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light

The nervous system has the remarkable ability to adapt and respond to various stimuli. This neural adjustment is largely achieved through plasticity at the synaptic level. The Active Zone (AZ) is the region at the presynaptic membrane that mediates neurotransmitter release and is composed of a dense collection of scaffold proteins. AZs of Drosophila melanogaster (Drosophila) photoreceptors undergo molecular remodeling after prolonged exposure to natural ambient light. Thus the level of neuronal activity can rearrange the molecular composition of the AZ and contribute to the regulation of the functional output.
Starting from the light exposure set-up preparation to the immunohistochemistry, this protocol details how to quantify the number, the spatial distribution, and the delocalization level of synaptic molecules at AZs in Drosophila photoreceptors. Using image analysis software, clusters of the GFP-fused AZ component Bruchpilot were identified for each R8 photoreceptor (R8) axon terminal. Detected Bruchpilot spots were automatically assigned to individual R8 axons. To calculate the distribution of spot frequency along the axon, we implemented a customized software plugin. Each axon&#39;s start-point and end-point were manually defined and the position of each Bruchpilot spot was projected onto the connecting line between start and end-point. Besides the number of Bruchpilot clusters, we also quantified the delocalization level of Bruchpilot-GFP within the clusters. These measurements reflect in detail the spatially resolved synaptic dynamics in a single neuron under different environmental conditions to stimuli.

Analyzing Synaptic Modulation of Drosophila melanogaster Photoreceptors after Exposure to Prolonged Light

Here we show how to quantify the number and spatial distribution of synaptic active zones in Drosophila melanogaster&#160;photoreceptors, highlighted with a genetically encoded molecular marker, and their modulation after prolonged exposure to light.

分析突触调制<em&gt;果蝇</em&gt;曝光时间延长后，光光感受器

The nervous system has the remarkable ability to adapt and respond to various stimuli. This neural adjustment is largely achieved through plasticity at ...

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the pathophysiological basis of these diseases, it is necessary to precisely characterize in which way the processing of information is disturbed between the different neuronal parts of the circuit. Using extracellular in vivo electrophysiological recordings, it is possible to accurately delineate neuronal activity within a neuronal network. The application of this method has several advantages over alternative techniques, e.g., functional magnetic resonance imaging and calcium imaging, as it allows a unique temporal and spatial resolution and does not rely on genetically engineered organisms. However, the use of extracellular in vivo recordings is limited since it is an invasive technique that cannot be universally applied. In this article, a simple and easy to use method is presented with which it is possible to simultaneously record extracellular potentials such as local field potentials and multiunit activity at multiple sites of a network. It is detailed how a precise targeting of subcortical nuclei can be achieved using a combination of stereotactic surgery and online analysis of multi-unit recordings. Thus, it is demonstrated, how a complete network such as the hyperdirect cortico-basal ganglia loop can be studied in anesthetized animals in vivo.

Acute In Vivo Electrophysiological Recordings of Local Field Potentials and Multi-unit Activity from the Hyperdirect Pathway in Anesthetized Rats

In this study, the methodology is presented on how to perform multi-site in vivo electrophysiological recordings from the hyperdirect pathway under urethane anesthesia.

急性<em&gt;体内</em&gt;麻醉大鼠超导途径局部场电位和多单位活动的电生理记录

Converging evidence shows that many neuropsychiatric diseases should be understood as disorders of large-scale neuronal networks. To better understand the ...

In vivo Calcium Imaging in Mouse Inferior Olive

Inferior olive (IO), a nucleus in the ventral medulla, is the only source of climbing fibers that form one of the two input pathways entering the cerebellum. IO has long been proposed to be crucial for motor control and its activity is currently considered to be at the center of many hypotheses of both motor and cognitive functions of the cerebellum. While its physiology and function have been relatively well studied on single-cell level in vitro, presently there are no reports on the organization of the IO network activity in living animals. This is largely due to the extremely challenging anatomical location of the IO, making it difficult to subject to conventional fluorescent imaging methods, where an optic path must be created through the entire brain located dorsally to the region of interest.
Here we describe an alternative method for obtaining state-of-the-art -level calcium imaging data from the IO network. The method takes advantage of the extreme ventral location of the IO and involves a surgical procedure for inserting a gradient-refractive index (GRIN) lens through the neck viscera to come into contact with the ventral surface of the calcium sensor GCaMP6s-expressing IO in anesthetized mice. A representative calcium imaging recording is shown to demonstrate the feasibility to record IO neuron activity after the surgery. While this is a non-survival surgery and the recordings must be conducted under anesthesia, it avoids damage to life-critical brainstem nuclei and allows conducting large variety of experiments investigating spatiotemporal activity patterns and input integration in the IO. This procedure with modifications could be used for recordings in other, adjacent regions of the ventral brainstem.

In vivo Calcium Imaging in Mouse Inferior Olive

We present a protocol to expose the brainstem of adult mouse from the ventral side. By using a gradient-refractive index lens with a miniature microscope, calcium imaging can be used to examine the activity of inferior olive neural somata in vivo.

在分析癫痫样活动和运动的一种低成本方法

摘要

探索更多视频

此视频中的章节

含量自发活动，研究昼夜节律和睡眠参数果蝇

在啮齿动物的行为测听的低成本设置

人机界面集成的低成本传感器，具有神经肌肉电刺激系统中风后康复平衡

测定冷灵敏度和适应小鼠一种简单廉价的方法

方法对敌人造成闭合性颅脑创伤性脑损伤

小鼠脑切片准备扣押样活性的直接电流刺激和多电极阵列记录

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

分析突触调制

急性

在体内小鼠劣质橄榄中的钙成像

在分析癫痫样活动和运动的一种低成本方法

摘要

探索更多视频

此视频中的章节

含量自发活动，研究昼夜节律和睡眠参数果蝇

在啮齿动物的行为测听的低成本设置

人机界面集成的低成本传感器，具有神经肌肉电刺激系统中风后康复平衡

测定冷灵敏度和适应小鼠一种简单廉价的方法

方法对敌人造成闭合性颅脑创伤性脑损伤

小鼠脑切片准备扣押样活性的直接电流刺激和多电极阵列记录

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

分析突触调制

急性

在体内 小鼠劣质橄榄中的钙成像

在体内小鼠劣质橄榄中的钙成像