我们要感谢Astha Maltare产生的幼虫爬行数据。我们要感谢博士尼古拉斯·兰森小的手稿给他的意见。为ALS在约翰霍普金斯大学的罗伯特·帕卡德中心（百联）和肌萎缩性侧索硬化症协会（百联）;和支持这项工作是从国家精神卫生研究院（CDN），R01MH083689。

A.幼虫爬行法 1。幼虫收集<ol><li>成立8盎司一瓶苍蝇（10-15男性10-15女性）。 </li><li>让苍蝇奠定24小时的卵子，然后明确瓶苍蝇。 （转移到​​一个新瓶的成年人，并在必要时重复）。 </li><li>孵育瓶为3-4天，或直至三龄幼虫均可见。 </li><li>添加50 - 100毫升20％的蔗糖与幼虫的瓶子，让坐了20分钟。幼虫会浮到顶端。 </li><li>收集的幼虫用25毫升血清吸管尖端切断，成网状篮子的地方。 </li><li>洗两次去离子H 2 O幼虫在网篮幼虫为实验做好准备。 </li></ol> 2。治疗药物的幼虫<ol><li>用刷子刷到5毫升的烧杯中含有一种解决方案蔗糖5％+药物输送所需数量的幼虫。 </li><li>让至少15分钟的幼虫饲料。</li><li>网状筐倒入药物治疗的幼虫和冲洗。他们现在已经准备好使用。 </li></ol> 3。运动检测（测量总距离行驶或体壁收缩） <ol><li>用刷子个别运输幼虫到: <ol><li> 15厘米的培养皿中含有超过0.2厘米2格方格纸，2％琼脂糖（以前浇，并允许变硬）。 <ol><li>计数在1分钟内越过网格线的数目。 </li></ol></li><li>良好的玻璃夹层菜含有稀释的酵母膏溶液。 <ol><li>解剖显微镜下观察，计数在一分钟内蠕动收缩（全前后运动= 1收缩）。 </li></ol></li></ol></li><li>重复，直到幼虫所需的数字已经算。 </li></ol> B.快速迭代负geotaxis（环）协议此法德scribed由加尔加诺等 5。 <ol><li>收集到一个标准的小瓶含食品（或食物+药物）新出现的光CO 2麻醉和地方的成年男性苍蝇。 </li><li>在室温下保持苍蝇（台式〜22°）为2-3天，以便从CO 2回收（稳态药物浓度的积累，如果适当）。 </li><li> 无需麻醉准备聚苯乙烯瓶约25苍蝇转让。 </li><li>苍蝇瓶组装成环装置（图1）。 </li><li>让苍蝇的环境，不受干扰15-20分钟，以适应新环境。 </li><li>在此期间，地方的数码相机〜1米（在一个平台上，如果有必要的镜头对准中旬高度小瓶中心）在前面的仪器，集中和放大到仪器的摄像头，并设置一个计时器3.0秒。 </li><li>仔细举行的环器具用你的左手，以免区URB的苍蝇，并保持用右手的计时器。 </li><li>大幅挖掘设备上的三次替补的表面，确保水龙头击倒所有的苍蝇瓶的底部是很难足够。 </li><li>同时完成第三自来水，开始倒数计时3秒。 </li><li>在三秒钟拍摄一张照片。 </li><li> 1分钟复位定时器和启动。在此期间，重置设备上的摄像头和重点，并另设三秒钟的定时器通道。 </li><li> 1分钟后，重复步骤1.7-1.10 </li><li>共5-6次试验后，上传到电脑的图像，并使用您最喜爱的图像浏览器打开，并得分的平均身高为每个小瓶攀升。 </li><li>进行统计分析，对不同群体比较，平均身高增长。 </li></ol> C.求爱和交配实验<ol><li>在早晨的第一件事，明确生产FLI瓶要使用的ES。 </li><li>在一天的课程（每3-4小时），收集新出现的性幼稚的男性和女性: <ol><li>介质的小瓶或管分别放置男性。 </li><li>每瓶/管放置5-6女性。 </li><li>在25°C下12小时的光/暗为5天的隔离收集苍蝇。 </li><li>转移到交配轮室的一名女性。 </li><li>传送一个男性的交配轮腔。 </li><li>解剖显微镜下观察，对有下列行为: <ol><li>取向（男性对女性的定向） </li><li>攻丝（男性水龙头女性的） </li><li>永歌（男延伸和振动一翼） </li><li>舔（男舔女性生殖器） </li><li>冰壶（男性卷发下腹部） </li><li>交配尝试（冰壶活动，而试图装入女） </li></ol></li></ol></li><li>观察10分钟或直至成功交配，注意到每个行为发生的时间（潜伏期），从事直到交配求偶行为（计算的求偶指数），以及对数，成功地执行一个给定的行为的总时间（频率）。 100％的野生型对一般会在5分钟内交配。 </li><li>计算 求爱指数（CI）的时间除以花求爱除以直到交配的总时间。对于野生型对这个范围通常在0.6-0.8之间。 </li></ol> D.代表结果爬行法正常野生型幼虫会徘徊〜3厘米/分钟，和展览〜40-50体壁收缩，在一分钟内。最近，我们已经制定了相关的FUS / TLS-肌萎缩性侧索硬化症，幼虫爬行缺陷，减少寿命和成人攀岩减值6果蝇模型。我们有针对性的表达Øf野生型和突变形式的FUS / TLS的运动神经元（OK-371-GAL4驱动器），并进行了幼虫爬行检测。如下图所示，野生型幼虫爬至12厘米，而野生型的FUS的表达下降约6厘米的幼虫爬行能力。表达的FUS / TLS的ALS的致病突变R521C的动物展示在他们的爬行运动非常严重受损（图1），爬行大约只有1厘米/分钟。 负geotaxis环法年轻的野生型成蝇，应该有一个3秒的时间在4-5厘米（3秒的时间可以从调整，以适应不同品系或活动水平定义为一个给定的一个给定的平均身高平均爬升高度应变/处理）。保持在底部的苍蝇，被分配一个0值。不建议使用超过25％小瓶苍蝇，因为它然后变得难以确定每一个人的苍蝇来衡量高度的位置。 ñØ脱敏已被观察到，在间隔1分钟，除了我们已经聘请了连续6个试验。关键是不重用初始数据集后，在本实验聚苯乙烯测试小瓶收集，因为到用小瓶放在新苍蝇不会爬新鲜小瓶到相同的程度。 <img alt="图1" src="/files/ftp_upload/3795/3795fig1.jpg" /> 图1。安装环检测。数码相机被放置在含有聚苯乙烯瓶苍蝇的仪器前约1米;对焦和变焦相机到设备，并设置一个计时器，以3.0秒。 <img alt="图2" src="/files/ftp_upload/3795/3795fig2.jpg" /> 图2。代表数据果蝇幼虫爬行使用检测异位表达的UAS-Fu的野生和UAS-Fu的控制下运动神经元驱动（OK371-GAL4）R521C。 <p类"jove_content"&gt;

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Genet. 34, 205-232 (2000).</li><li>Villella, A., Hall, J. C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurogenetics+of+courtship+and+mating+in+Drosophila.">Neurogenetics of courtship and mating in Drosophila.</a> Adv. Genet. 62, 67-184 (2008).</li></ol>

我们什么都没有透露。

果蝇的行为是严格受遗传和环境因素。我们和其他人，以前使用这里描述的实验，收集数据，检查飞的行为，并在果蝇 5-19仿照人类神经退行性疾病相关的基因。 第三龄幼虫爬行分析，慎重选择是关键的一步。如果用药物治疗，它会采取10-15分钟（或更取决于药物的类型和性质），以达到最大的效果，如果有良好的溶解性。因此，我们经常喂药物飞行15分钟，然后等待一个额外的15分钟前测试。然而，重要的是，保持在溶液中的药物浓度，治疗组之间的曝光时间，准确的比较。中枢神经系统活性药物通常都会有一个最大的效果持久〜45分钟。应洗好后，选择（或挖喂养）消除苍蝇食品和幼虫1分钟前开始爬行法允许水土不服。琼脂平板应保持在室温（〜22℃），一个小时，低温可以影响幼虫爬行能力。虽然幼虫爬行检测，可以提供重要的信息，有关活动水平，这是不适合的微妙协调财政赤字的分析。因此，作为一个筛选平台，它是最适当的检查活动的赤字总额的第一个通。 一个成年人的行为，涉及精细动作的协调是求偶和交配。这种行为已被用于研究人类疾病有关的行为的各个方面，涉及处理感官（嗅觉，视觉，听觉），除了罚款电机控制18。当一个男性发现女性，他发起了求爱仪式，在与取向的行为（转向和追逐女性）的刻板模式开始进展。这是其次，lickin wingsongg和攻牙女性genetalia，对女性的腹部卷曲，并在交配，它可以持续几十分钟20-22高潮。因为求爱的许多方面涉及的视觉线索，黑暗条件下导致了交配性能的退化，并应具有足够的光线进行检测，使苍蝇可以看到彼此。因此，白色眼睛的苍蝇通常我们描述实验中的表现极差，读者对规划的实验，他们利用这个协议的建议。如果用药物治疗的苍蝇，放入培养基药物苍蝇被隔离（如1％琼脂糖+蔗糖10％+的药物，而不是标准的食品与药物治疗时，以避免任何可能降解的药物中的微生物食品）。一般苍蝇被隔离到5毫升的试管中有300-500毫升的食品，在他们与棉花堵塞。 不要麻醉转移到交配轮的苍蝇。它，我绝对关键洗交配很好轮之间使用，以消除任何残留的费洛蒙（少量过夜alconox肥皂温水，用颤抖的，最低的48个小时去离子水冲洗，然后用颤抖的，无数的变化和水） 。执行我们所描述的实验时，另一个需要考虑的是天气。在我们的经验，苍蝇不会告上法庭，如果下雨或看起来会下雨。他们的表现最好在阳光明媚的日子，无论是实验室有窗户或没有。目前，我们的理论是，这种现象与大气压力，但我们还没有研究这个。 传统的的的负面geotaxis检测依赖于测量有多少苍蝇爬预定在10秒（中所述的高度<a href="https://www-jove-com.remotexs.ntu.edu.sg/details.php?id=2504">https://www-jove-com.remotexs.ntu.edu.sg/details.php?id=2504</a> ）以上。我们相信，环法有一定的优势超过了传统的检测。一个是throughput，为六个独立的复制可以同时测量标准检测相比，该系统一般可伸缩。另一个是灵敏度，因为平均身高被量化，而不是一个合格/不合格的绝对高度数，在规定的时间内攀升。使用这种方法，可以观察到更微妙的赤字。由于吞吐量的水​​平，该法更适合屏幕比传统的负面geotaxis检测更繁琐。此外，Garagano 等。（2005）描述了电脑计分法，如果将进一步增加吞吐量。此法的主要考虑是，它是必要的，苍蝇不麻醉测试前，新鲜瓶是用于给定的试验后，一批新的苍蝇测试之前。

<table border="1"><tbody><tr><td> 试剂名称 </td><td> 公司 </td><td> 目录编号 </td></tr><tr><td>蔗糖</td><td> Fisher Scientific则</td><td> S5-500 </td></tr><tr><td>琼脂糖</td><td> Invitrogen公司</td><td> 16500-500 </td></tr><tr><td> 6盎司果蝇瓶</td><td>杰纳西科学</td><td> 32-130 </td></tr><tr><td>油漆刷（＃1） </td><td>特德·佩拉，公司</td><td> 11859 </td></tr><tr><td> 飞食品组件 </td><td></td><td></td></tr><tr><td>棒子面</td><td> Fisher Scientific则</td><td> NC9109741 </td></tr><tr><td>洋菜</td><td>杰纳西科学</td><td> 66-104 </td></tr><tr><td>糖蜜</td><td> Fisher Scientific则</td><td> NC9349176 </td></tr><tr><td>丙酸</td><td>阿库罗斯福冈</td><td> 14930-0010 </td></tr><tr><td> tegosept </td><td>顶点</td><td> 20-258 </td></tr><tr><td>乙醇</td><td>费舍尔科学的</td><td> BP2818-4 </td></tr><tr><td>酵母</td><td>杰纳西科学</td><td> 62-107 </td></tr></tbody></table>

methods to assay drosophila behavior

果蝇 ，果蝇，已用于研究的范围广泛的人类疾病，如癌症，心血管疾病及各种神经系统疾病的分子机制。我们已经优化了简单和强大的行为分析，确定幼虫的运动，成年的爬坡能力（环法），对果蝇的求偶行为。这些行为的分析是研究遗传和环境因素对飞行为的作用，广泛适用。幼虫爬行能力，能可靠地用于确定在果蝇幼虫爬行能力的早期阶段的变化，并检查他们的运动，药物或人类疾病基因的影响（转基因果蝇）。幼虫爬行检测变得更适用，如果一个基因的表达或取消导致蛹或成人阶段的杀伤力，因为这些苍蝇不活到成年，他们可以评估。这基本也可用于ssay配合明亮的光线或应力研究果蝇幼虫额外的行为反应。求偶行为已被广泛用于研究遗传基础的性行为，也可用于检查活动和协调，以及学习和记忆。 果蝇求偶行为涉及的各种感官刺激，包括视觉，听觉和化学感应信号交换之间的男性和女性，导致一系列复杂的特点以及电机最终成功交配行为。传统的成人攀岩检测（负geotaxis），单调乏味，劳动密集，耗时2-4不同试验之间的显着变化，。快速迭代5负geotaxis（环）法有许多优势，更广泛采用的协议，提供了一个重复性好，灵敏度高，高通量的方法来量化成人运动和消极geotaxis behaviors。在环法，几个基因型或药物治疗，可以同时使用大量的动物测试，高通量的方法筛选试验更适合。

Watch this Scientific Journal Video about Methods to Assay Drosophila Behavior at JoVE.com

Methods to Assay Drosophila Behavior

果蝇是一种基因和行为听话的模型系统已用于了解一个多世纪1的基础上的许多重要的生物过程的分子和细胞， 果蝇已经得到了很好的利用获得到飞行为的遗传基础的见解。

对检测方法果蝇行为

Drosophila melanogaster, the fruit fly, has been used to study molecular mechanisms of a wide range of human diseases such as cancer, cardiovascular ...

Nanyang Technological University

Research

JoVE Journal

Neuroscience

Methods to Assay Drosophila Behavior

42.7K 次观看.  Louisiana State University Health Sciences Center. 果蝇是一种基因和行为听话的模型系统已用于了解一个多世纪1的基础上的许多重要的生物过程的分子和细胞， 果蝇已经得到了很好的利用获得到飞行为的遗传基础的见解。

视频: Methods to Assay Drosophila Behavior

Single Sensillum Recordings in the Insects Drosophila melanogaster and Anopheles gambiae

The sense of smell is essential for insects to find foods, mates, predators, and oviposition sites3. Insect olfactory sensory neurons (OSNs) are enclosed in sensory hairs called sensilla, which cover the surface of olfactory organs. The surface of each sensillum is covered with tiny pores, through which odorants pass and dissolve in a fluid called sensillum lymph, which bathes the sensory dendrites of the OSNs housed in a given sensillum. The OSN dendrites express odorant receptor (OR) proteins, which in insects function as odor-gated ion channels4, 5. The interaction of odorants with ORs either increases or decreases the basal firing rate of the OSN. This neuronal activity in the form of action potentials embodies the first representation of the quality, intensity, and temporal characteristics of the odorant6, 7.
Given the easy access to these sensory hairs, it is possible to perform extracellular recordings from single OSNs by introducing a recording electrode into the sensillum lymph, while the reference electrode is placed in the lymph of the eye or body of the insect. In Drosophila, sensilla house between one and four OSNs, but each OSN typically displays a characteristic spike amplitude. Spike sorting techniques make it possible to assign spiking responses to individual OSNs. This single sensillum recording (SSR) technique monitors the difference in potential between the sensillum lymph and the reference electrode as electrical spikes that are generated by the receptor activity on OSNs1, 2, 8. Changes in the number of spikes in response to the odorant represent the cellular basis of odor coding in insects. Here, we describe the preparation method currently used in our lab to perform SSR on Drosophila melanogaster and Anopheles gambiae, and show representative traces induced by the odorants in a sensillum-specific manner.

Single Sensillum Recordings in the Insects Drosophila melanogaster and Anopheles gambiae

Electrophysiological responses of olfactory sensory neurons to odorants can be measured in insects using single sensillum recordings. In this video article we will demonstrate how to perform single sensillum recordings in the antennae of the vinegar fly (Drosophila melanogaster) and the maxillary palps of the malaria mosquito (Anopheles gambiae).

在昆虫单感器记录果蝇按蚊</em

The sense of smell is essential for insects to find foods, mates, predators, and oviposition sites3. Insect olfactory sensory neurons (OSNs) are enclosed ...

科研

神经科学

Methods for Study of Neuronal Morphogenesis: Ex vivo RNAi Electroporation in Embryonic Murine Cerebral Cortex

The cerebral cortex directs higher cognitive functions. This six layered structure is generated in an inside-first, outside-last manner, in which the first born neurons remain closer to the ventricle while the last born neurons migrate past the first born neurons towards the surface of the brain1. In addition to neuronal migration2, a key process for normal cortical function is the regulation of neuronal morphogenesis3. While neuronal morphogenesis can be studied in vitro in primary cultures, there is much to be learned from how these processes are regulated in tissue environments.
We describe techniques to analyze neuronal migration and/or morphogenesis in organotypic slices of the cerebral cortex4,6. A pSilencer modified vector is used which contains both a U6 promoter that drives the double stranded hairpin RNA and a separate expression cassette that encodes GFP protein driven by a CMV promoter7-9. Our approach allows for the rapid assessment of defects in neurite outgrowth upon specific knockdown of candidate genes and has been successfully used in a screen for regulators of neurite outgrowth8. Because only a subset of cells will express the RNAi constructs, the organotypic slices allow for a mosaic analysis of the potential phenotypes. Moreover, because this analysis is done in a near approximation of the in vivo environment, it provides a low cost and rapid alternative to the generation of transgenic or knockout animals for genes of unknown cortical function. Finally, in comparison with in vivo electroporation technology, the success of ex vivo electroporation experiments is not dependant upon proficient surgery skill development and can be performed with a shorter training time and skill.

Methods for Study of Neuronal Morphogenesis: Ex vivo RNAi Electroporation in Embryonic Murine Cerebral Cortex

To conduct a rapid assessment of the function of genes in the development of cerebral cortex, we describe methods involving the ex vivo electroporation of plasmids co-expressing inhibitory RNA (RNAi) and GFP in murine embryonic cortex. This protocol is amenable to the study of various aspects of neurodevelopment such as neurogenesis, neuronal migration and neuronal morphogenesis including dendrite and axon outgrowth.

神经元形态的研究方法:<em&gt;体外</em&gt; RNAi技术在胚胎小鼠大脑皮质的电穿孔

The cerebral cortex directs higher cognitive functions. This six layered structure is generated in an inside-first, outside-last manner, in which the ...

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.
The Drosophila larva is an established model system to study thermal nociception, a sensory response to potentially harmful temperatures that is evolutionarily conserved across species1,2. The advantages of Drosophila for such studies are the relative simplicity of its nervous system and the sophistication of the genetic techniques that can be used to dissect the molecular basis of the underlying biology4-6 In Drosophila, as in all metazoans, the response to noxious thermal stimuli generally involves a "nocifensive" aversive withdrawal to the presented stimulus7. Such stimuli are detected through free nerve endings or nociceptors and the amplitude of the organismal response depends on the number of nociceptors receiving the noxious stimulus8. In Drosophila, it is the class IV dendritic arborization sensory neurons that detect noxious thermal and mechanical stimuli9 in addition to their recently discovered role as photoreceptors10. These neurons, which have been very well studied at the developmental level, arborize over the barrier epidermal sheet and make contacts with nearly all epidermal cells11,12. The single axon of each class IV neuron projects into the ventral nerve cord of the central nervous system11 where they may connect to second-order neurons that project to the brain.
Under baseline conditions, nociceptive sensory neurons will not fire until a relatively high threshold is reached. The assays described here allow the investigator to quantify baseline behavioral responses or, presumably, the sensitization that ensues following tissue damage. Each assay provokes distinct but related locomotory behavioral responses to noxious thermal stimuli and permits the researcher to visualize and quantify various aspects of thermal nociception in Drosophila larvae. The assays can be applied to larvae of desired genotypes or to larvae raised under different environmental conditions that might impact nociception. Since thermal nociception is conserved across species, the findings gleaned from genetic dissection in Drosophila will likely inform our understanding of thermal nociception in other species, including vertebrates.

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of thermal nociceptors1,2 while the second involves a wholesale (global) activation of most or all such neurons3. Together, these techniques allow visualization and quantification of the behavioral functions of Drosophila nociceptive sensory neurons.

局部和全局的方法评估热痛觉<em&gt;果蝇</em&gt;幼虫

In this article, we demonstrate assays to study thermal nociception in Drosophila larvae. One assay involves spatially-restricted (local) stimulation of ...

Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster

A growing number of genetically encoded tools are becoming available that allow non-invasive manipulation of the neural activity of specific neurons in Drosophila melanogaster1. Chief among these are optogenetic tools, which enable the activation or silencing of specific neurons in the intact and freely moving animal using bright light. Channelrhodopsin (ChR2) is a light-activated cation channel that, when activated by blue light, causes depolarization of neurons that express it. ChR2 has been effective for identifying neurons critical for specific behaviors, such as CO2 avoidance, proboscis extension and giant-fiber mediated startle response2-4. However, as the intense light sources used to stimulate ChR2 also stimulate photoreceptors, these optogenetic techniques have not previously been used in the visual system. Here, we combine an optogenetic approach with a mutation that impairs phototransduction to demonstrate that activation of a cluster of loom-sensitive neurons in the fly's optic lobe, Foma-1 neurons, can drive an escape behavior used to avoid collision. We used a null allele of a critical component of the phototransduction cascade, phospholipase C-&beta;, encoded by the norpA gene, to render the flies blind and also use the Gal4-UAS transcriptional activator system to drive expression of ChR2 in the Foma-1 neurons. Individual flies are placed on a small platform surrounded by blue LEDs. When the LEDs are illuminated, the flies quickly take-off into flight, in a manner similar to visually driven loom-escape behavior. We believe that this technique can be easily adapted to examine other behaviors in freely moving flies.

Optogenetic Stimulation of Escape Behavior in Drosophila melanogaster

Genetically encoded optogenetic tools enable noninvasive manipulation of specific neurons in the Drosophila brain. Such tools can identify neurons whose activation is sufficient to elicit or suppress particular behaviors. Here we present a method for activating Channelrhodopsin2 that is expressed in targeted neurons in freely walking flies.

Optogenetic刺激的逃避行为<em&gt;果蝇</em

A  growing number of genetically encoded tools are becoming available that allow non-invasive  manipulation of the neural activity of specific neurons in ...

A Single-fly Assay for Foraging Behavior in Drosophila

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster. In a light-tight box illuminated by red light that is invisible to fruit flies, a camera linked to custom data acquisition software monitors the position of six flies simultaneously. Each fly is confined to walk in individual arenas containing a food odor at the center. The testing arenas rest on a porous floor that functions to prevent odor accumulation. Latency to locate the odor source, a metric that reflects olfactory sensitivity under different physiological states, is determined by software analysis. Here, we discuss the critical mechanics of running this behavioral paradigm and cover specific issues regarding fly loading, odor contamination, assay temperature, data quality, and statistical analysis.

A Single-fly Assay for Foraging Behavior in Drosophila

In this video article, we describe an automated assay to measure the effect of hunger or satiety on olfactory dependent food search behavior in the adult fruit fly Drosophila melanogaster.

单苍蝇检测方法的觅食行为的<em&gt;果蝇</em

For many animals, hunger promotes changes in the olfactory system in a manner that facilitates the search for appropriate food sources. In this video ...

Dyeing Insects for Behavioral Assays: the Mating Behavior of Anesthetized Drosophila

Mating experiments using Drosophila have contributed greatly to the understanding of sexual selection and behavior. Experiments often require simple, easy and cheap methods to distinguish between individuals in a trial. A standard technique for this is CO2 anaesthesia and then labelling or wing clipping each fly. However, this is invasive and has been shown to affect behavior. Other techniques have used coloration to identify flies. This article presents a simple and non-invasive method for labelling Drosophila that allows them to be individually identified within experiments, using food coloring. This method is used in trials where two males compete to mate with a female. Dyeing allowed quick and easy identification. There was, however, some difference in the strength of the coloration across the three species tested. Data is presented showing the dye has a lower impact on mating behavior than CO2 in Drosophila melanogaster. The impact of CO2 anaesthesia is shown to depend on the species of Drosophila, with D. pseudoobscura and D. subobscura showing no impact, whereas D. melanogaster males had reduced mating success. The dye method presented is applicable to a wide range of experimental designs.

Dyeing Insects for Behavioral Assays: the Mating Behavior of Anesthetized Drosophila

This protocol describes a simple, cost effective way to individually identify Drosophila or other insects. Demonstration data investigating mating success across three species of Drosophila show that this method is comparable or better than the use of CO2 anaesthesia.

染色昆虫行为检测:麻醉的交配行为<em&gt;果蝇</em

Mating experiments using Drosophila have contributed greatly to the understanding of sexual selection and behavior. Experiments often require simple, easy ...

Using Linear Agarose Channels to Study Drosophila Larval Crawling Behavior

Drosophila larval crawling is emerging as a powerful model to study neural control of sensorimotor behavior. However, larval crawling behavior on flat open surfaces is complex,&#160;including: pausing, turning, and meandering. This complexity in the repertoire of movement hinders detailed analysis of the events occurring during a single crawl stride cycle. To overcome this obstacle, linear agarose channels were made that constrain larval behavior to straight, sustained, rhythmic crawling. In principle, because agarose channels and the Drosophila larval body are both optically clear, the movement of larval structures labeled by genetically-encoded fluorescent probes can be monitored in intact, freely-moving larvae. In the past, larvae were placed in linear channels and crawling at the level of whole organism, segment, and muscle were analyzed1. In the future, larvae crawling in channels can be used for calcium imaging to monitor neuronal activity. Moreover, these methods can be used with larvae of any genotype and with any researcher-designed channel. Thus the protocol presented below is widely applicable for studies using the Drosophila larva as a model to understand motor control.

Using Linear Agarose Channels to Study Drosophila Larval Crawling Behavior

The Drosophila larva is a powerful model system to study neural control of behavior. This publication describes the use of linear agarose channels to elicit sustained bouts of linear crawling and methods to quantify the dynamics of larval structures during repetitive crawling behavior.

用线性琼脂糖渠道研究<em&gt;果蝇</em&gt;幼虫爬行行为

Drosophila larval crawling is emerging as a powerful model to study neural control of sensorimotor behavior. However, larval crawling behavior on flat ...

A Simple Way to Measure Alterations in Reward-seeking Behavior Using Drosophila melanogaster

We describe a protocol for measuring ethanol self-administration in fruit flies (Drosophila melanogaster) as a proxy for changes in reward states. We demonstrate a simple way to tap into the fly reward system, modify experiences related to natural reward, and use voluntary ethanol consumption as a measure for changes in reward states. The approach serves as a relevant tool to study the neurons and genes that play a role in experience-mediated changes of internal state. The protocol is composed of two discrete parts: exposing the flies to rewarding and nonrewarding experiences, and assaying voluntary ethanol consumption as a measure of the motivation to obtain a drug reward. The two parts can be used independently to induce the modulation of experience as an initial step for further downstream assays or as an independent two-choice feeding assay, respectively. The protocol does not require a complicated setup and can therefore be applied in any laboratory with basic fly culture tools.

A Simple Way to Measure Alterations in Reward-seeking Behavior Using Drosophila melanogaster

We describe a protocol for inducing rewarding and nonrewarding experiences in fruit flies (Drosophila melanogaster) using voluntary ethanol consumption as a measure for changes in reward states.

一个简单的方法来衡量在改建悬赏寻求行为使用<em&gt;果蝇</em

We describe a protocol for measuring ethanol self-administration in fruit flies (Drosophila melanogaster) as a proxy for changes in reward states. We ...

Quantification of Drosophila Grooming Behavior

Drosophila grooming behavior is a complex multi-step locomotor program that requires coordinated movement of both forelegs and hindlegs. Here we present a grooming assay protocol and novel chamber design that is cost-efficient and scalable for either small or large-scale studies of Drosophila grooming. Flies are dusted all over their body with Brilliant Yellow dye and given time to remove the dye from their bodies within the chamber. Flies are then deposited in a set volume of ethanol to solubilize the dye. The relative spectral absorbance of dye-ethanol samples for groomed versus ungroomed animals are measured and recorded. The protocol yields quantitative data of dye accumulation for individual flies, which can be easily averaged and compared across samples. This allows experimental designs to easily evaluate grooming ability for mutant animal studies or circuit manipulations. This efficient procedure is both versatile and scalable. We show work-flow of the protocol and comparative data between WT animals and mutant animals for the Drosophila type I Dopamine Receptor (DopR).

This protocol describes a scalable individual grooming assay technique in Drosophila that yields robust, quantitative data to measure grooming behavior. The method is based on comparing the difference in dye accumulation on the bodies of un-groomed versus groomed animals over a set period of time.

果蝇美容行为的量化

Drosophila grooming behavior is a complex multi-step locomotor program that requires coordinated movement of both forelegs and hindlegs. Here we present a ...

High-throughput Analysis of Locomotor Behavior in the Drosophila Island Assay

Advances in next-generation sequencing technologies contribute to the identification of (candidate) disease genes for movement disorders and other neurological diseases at an increasing speed. However, little is known about the molecular mechanisms that underlie these disorders. The genetic, molecular, and behavioral toolbox of Drosophila melanogaster makes this model organism particularly useful to characterize new disease genes and mechanisms in a high-throughput manner. Nevertheless, high-throughput screens require efficient and reliable assays that, ideally, are cost-effective and allow for the automatized quantification of traits relevant to these disorders. The island assay is a cost-effective and easily set-up method to evaluate Drosophila locomotor behavior. In this assay, flies are thrown onto a platform from a fixed height. This induces an innate motor response that enables the flies to escape from the platform within seconds. At present, quantitative analyses of filmed island assays are done manually, which is a laborious undertaking, particularly when performing large screens.
This manuscript describes the &#34;Drosophila Island Assay&#34; and &#34;Island Assay Analysis&#34; algorithms for&#160;high-throughput, automated data processing and quantification of island assay data. In the setup, a simple webcam connected to a laptop collects an image series of the platform while the assay is performed. The &#34;Drosophila Island Assay&#34; algorithm developed for the open-source software Fiji processes these image series and quantifies, for each experimental condition, the number of flies on the platform over time. The &#34;Island Assay Analysis&#34; script, compatible with the free software R, was developed to automatically process the obtained data and to calculate whether treatments/genotypes are statistically different. This greatly improves the efficiency of the island assay and makes it a powerful readout for basic locomotion and flight behavior. It can thus be applied to large screens investigating fly locomotor ability, Drosophila models of movement disorders, and drug efficacy.

High-throughput Analysis of Locomotor Behavior in the Drosophila Island Assay

The island assay is a relatively new, cost-effective assay that can be used to evaluate the basic locomotor behavior of Drosophila melanogaster. This manuscript describes algorithms for automatic data processing and objective quantification of island assay data, making this assay a sensitive, high-throughput readout for large genetic or pharmacological screens.

对检测方法果蝇行为

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此视频中的章节

在昆虫单感器记录果蝇按蚊

神经元形态的研究方法:

局部和全局的方法评估热痛觉

Optogenetic刺激的逃避行为

单苍蝇检测方法的觅食行为的

染色昆虫行为检测:麻醉的交配行为

用线性琼脂糖渠道研究

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