Name: light preference assay to study innate and circadian regulated photobehavior in drosophila larvae
Uploaded: 2013-04-20T14:00:00
Description: Watch this Scientific Journal Video about Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae at JoVE.com

我们感谢我们的同事在弗里堡大学生物系，富有成效的讨论。我们感谢布卢明顿联合中心提供飞股票。财政支持这项工作是由瑞士国家科学基金会（PP00P3_123339）和威卢克斯基金会SGS

1。苗种培育<ol><li>保持飞菌株或大众文化的遗传杂交玉米粉培养基上，在25°C下12小时轻12小时黑暗周期中一只苍蝇孵化器配备有光线和定时器。 </li><li>稀释在水中，形成衬垫酵母的流体浆料（10克的背衬酵母稀释用3-4毫升蒸馏水H 2 O）。加一小滴玉米粉食品和覆盖小瓶。让至少一个小时的干燥，以避免成虫坚持酵母膏。将酵母水稀释一小滴表面上的食物可提高产卵。或者可以使用面包酵...

<ol>
	<li>Sawin-McCormack, E. P., Sokolowski, M. B., Campos, A. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Characterization+and+genetic+analysis+of+Drosophila+melanogaster+photobehavior+during+larval+development.">Characterization and genetic analysis of Drosophila melanogaster photobehavior during larval development.</a> J. Neurogenet. 10, 119-135 (1995).</li><li>Sprecher, S. G., Pichaud, F., Desplan, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Adult+and+larval+photoreceptors+use+different+mechanisms+to+specify+the+same+Rhodopsin+fates.">Adult and larval photoreceptors use different mechanisms to specify the same Rhodopsin fates.</a> Genes Dev. 21, 2182-2195 (2007).</li><li>Sprecher, S. G., Desplan, C. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Switch+of+rhodopsin+expression+in+terminally+differentiated+Drosophila+sensory+neurons.">Switch of rhodopsin expression in terminally differentiated Drosophila sensory neurons.</a> Nature. 454, 533-537 (2008).</li><li>Xiang, Y., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Light-avoidance-mediating+photoreceptors+tile+the+Drosophila+larval+body+wall.">Light-avoidance-mediating photoreceptors tile the Drosophila larval body wall.</a> Nature. 468, 921-926 (2010).</li><li>Diaz, N. N., Sprecher, S. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Photoreceptors:+unconventional+ways+of+seeing.">Photoreceptors: unconventional ways of seeing.</a> Curr. Biol. 21, R25-R27 (2011).</li><li>Emery, P., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Drosophila+CRY+is+a+deep+brain+circadian+photoreceptor.">Drosophila CRY is a deep brain circadian photoreceptor.</a> Neuron. 26, 493-504 (2000).</li><li>Mazzoni, E. O., Desplan, C., Blau, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Circadian+pacemaker+neurons+transmit+and+modulate+visual+information+to+control+a+rapid+behavioral+response.">Circadian pacemaker neurons transmit and modulate visual information to control a rapid behavioral response.</a> Neuron. 45, 293-300 (2005).</li><li>Keene, A. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Distinct+visual+pathways+mediate+Drosophila+larval+light+avoidance+and+circadian+clock+entrainment.">Distinct visual pathways mediate Drosophila larval light avoidance and circadian clock entrainment.</a> J. Neurosci. 31, 6527-6534 (2011).</li><li>Gong, Z. F., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Two+Pairs+of+Neurons+in+the+Central+Brain+Control+Drosophila+Innate+Light+Preference.">Two Pairs of Neurons in the Central Brain Control Drosophila Innate Light Preference.</a> Science. 330, 499-502 (2010).</li><li>Lilly, M., Carlson, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=smellblind:+a+gene+required+for+Drosophila+olfaction.">smellblind: a gene required for Drosophila olfaction.</a> Genetics. 124, 293-302 (1990).</li><li>Bodenstein, D., Demerec, 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+postembryonic+development+of+Drosophila.">The postembryonic development of Drosophila.</a> Biology of Drosophila. , 275-367 (1950).</li><li>Pittendrigh, C. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Circadian+systems:+Entrainment.">Circadian systems: Entrainment.</a> Biological Rhythms. 4 Handbook of Behavioral Neurobiology, 95-124 (1981).</li><li>Collins, B., Kane, E. A., Reeves, D. C., Akabas, M. H., Blau, J. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Balance+of+Activity+between+LN(v)s+and+Glutamatergic+Dorsal+Clock+Neurons+Promotes+Robust+Circadian+Rhythms+in+Drosophila.">Balance of Activity between LN(v)s and Glutamatergic Dorsal Clock Neurons Promotes Robust Circadian Rhythms in Drosophila.</a> Neuron. 74, 706-718 (2012).</li><li>Keene, A. C., Sprecher, S. 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光偏好测试幼虫的先天优势，感光特性。该法是很容易的建立，允许多次重复，以较低的成本和光传感和处理提供了有价值的信息。实验范式允许多少人喜欢浅色或深色的相对快速定量。这种偏好可以显示作为原油的百分比，或者偏好指数（县）。 PREF被表示为动物的差异，优选的光，优选黑暗除以动物的总的动物。 光偏好测试中的一个关键点是实验的时间。在这里，我们测...

在这里，我们描述了一个基于行为的检测暗（或光）幼虫偏好。幼虫的反应与一个强大和刻板避光反应在觅食阶段（L1至L3月初）1。幼虫避光行为的测定法的目的是评估和比较亮或暗的幼虫自由移动，在涂有琼脂的陪替氏培养皿中的一组的偏好。这种行为的检测不仅提供信息的敏感性，一体化和时间的视觉系统可塑性，它进一步提供了光敏感性及其过程控制的昼夜系统的提示。 果蝇幼虫眼（也被称为Bowlig的器官; BO），光感的主要器官。每只眼睛是由12个光感受器（PR），8 PRS表达绿色，：敏感rhodopsin6（RH6）和四个永久居民表达了对蓝色敏感rhodopsin5（RH5）2,3。此外PRS，ALSØIV级的multidendritic神经元，涵盖幼虫体壁，已经确定以应对4,5有害光线强度。它也被称为位于中央的幼虫脑起搏器的神经元表达光敏蛋白隐花色素（喊），作为时钟固有的蓝色光传感器大脑内的6,7。有趣的野生型动物photophobicity显示一昼夜成分在不同时间点的过程中，白天和夜间的时候，用这个方法测试。对L3幼虫觅食表现出较强的photophobicity在黎明和photophobicity的黄昏时进行测试时光黑暗的偏好7。有趣的是，只有RH5-PRS都需要避免轻，而RH6​​-PRS是可有可无的。两者RH5-PR和RH6-PRS参与重置分子钟光8。惊魂途径，必须协调与其他光感的途径，以协调在一个适当的行为输出一天的过程中。乙酰胆碱在PRS光回避行为，以及夹带的分子时钟起着至关重要的作用。从PRS昼夜起搏器神经元阻断神经递质乙酰胆碱减少光黑暗偏好测定8避光反应。采用相同的检测，两个对称的神经元对最近发现的第三龄幼虫果蝇 9开灯偏好。这两对神经元功能在后期幼体阶段，当动物留下的食物大概是找到一个合适的的pupariation网站。然而，视觉通路如何相互作用，一昼夜地控制幼虫的视觉行为的问题在很大程度上仍然未回答。光偏好测定允许昼夜时间点之间的比较，飞线和昼夜状态下不同光质。该法是很容易准备，价格低廉，一直是有用的，我以前Ñ​​几个实验室的描述和研究光派生行为在幼虫。

<table border="1">
 <tbody>
 <tr>
 <td>Name of Reagent/Material</td>
 <td>Company</td>
 <td>Catalog Number</td>
 <td>Comments</td>
 </tr>
 <tr>
 <td>Agar</td>
 <td>Sigma-Aldrich</td>
 <td>A5093-500G</td>
 <td>2.5%; Sigma-Aldrich, 9471 Buchs, Switzerland </td>
 </tr>
 <tr>
 <td>Petri dishes</td>
 <td>Greiner Bio-One GmbH</td>
 <td>633180</td>
 <td>90-mm diameter; Greiner Bio-One GmbH, 4550 Kremsmeinster, Austria</td>
 </tr>
 <tr>
 <td>LEDs Lamp</td>
 <td>OSARAM </td>
 <td>80012 White</td>
 <td>LED lamp, 80012 White</td>
 </tr>
 <tr>
 <td>Environment Meter</td>
 <td>PCE</td>
 <td>PCE EM882</td>
 <td>Lux, Temp, RH%</td>
 </tr>
 <tr>
 <td>Thermostatic cabinet</td>
 <td>Aqua Lytic (Liebherr)</td>
 <td>ET636-6</td>
 <td></td>
 </tr>
 <tr>
 <td>Light timer</td>
 <td>Timer T</td>
 <td>6185.104</td>
 <td>230V/50HZ (check specifications for your country)</td>
 </tr>
 <tr>
 <td>Universal thermostat</td>
 <td>Conrad</td>
 <td>UT200</td>
 <td></td>
 </tr>
 <tr>
 <td>Humidifier</td>
 <td>Boneco</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Balck tape</td>
 <td>Tesa</td>
 <td></td>
 <td>5 cm </td>
 </tr>
 <tr>
 <td>Glue</td>
 <td>Uhu</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>lncubator lamp</td>
 <td>Phillips</td>
 <td>Softtone</td>
 <td>5W</td>
 </tr>
 <tr>
 <td>Timer clock</td>
 <td>Ziliss</td>
 <td></td>
 <td>Ziliss, Switzerland</td>
 </tr>
 <tr>
 <td>Excel Software</td>
 <td>Microsoft</td>
 <td></td>
 <td>Excel</td>
 </tr>
 <tr>
 <td>Origin Software 8.5</td>
 <td>OriginLab</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Backer Yeast</td>
 <td>Migros Switzerland</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Iron support stand 17X28CM</td>
 <td>Fisher Scientific</td>
 <td>S47808</td>
 <td></td>
 </tr>
 <tr>
 <td>Acetic acid</td>
 <td>Sigma Aldrich</td>
 <td>A6283-100ML</td>
 <td>20% acetic acid dilluted in H2O</td>
 </tr>
 <tr>
 <td>Red light lamp</td>
 <td>Phillips</td>
 <td>PFE712E*8C</td>
 <td></td>
 </tr>
 <tr>
 <td>Spatula </td>
 <td>Fisher Scientific</td>
 <td>14-373-25A</td>
 <td></td>
 </tr>
 <tr>
 <td>Power supply</td>
 <td>EA</td>
 <td>EA PS 2042-06B</td>
 <td>Optional</td>
 </tr>
 <tr>
 <td>Aluminium foil</td>
 <td>Prix Coop</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Heater</td>
 <td>GOON</td>
 <td>NSB200C</td>
 <td></td>
 </tr>
 <tr>
 <td>Microwave Oven</td>
 <td>Intertronic</td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Standard corn meal fly food</td>
 <td></td>
 <td></td>
 <td></td>
 </tr>
 <tr>
 <td>Destilled water </td>
 <td></td>
 <td></td>
 <td></td>
 </tr>
 </tbody>
</table>

light preference assay to study innate and circadian regulated photobehavior in drosophila larvae

光作为环境信号来控制各级动物行为。 果蝇幼虫的神经系统是作为一个独特的模型，回答基本问题，如何快速昼夜行为之间的光信息处理和共享。 果蝇幼虫显示刻板的避税行为，当暴露在光线下。探讨可比简单的明暗偏好测试可应用于光依赖行为。参与传感和处理视觉输入的神经通路在脊椎动物和节肢动物部分重叠与处理光的昼夜信息。迷人的光传感系统和昼夜系统如何互动，以保持输出协调行为的问题，仍然在很大程度上未被果蝇是影响生物模型来处理这些问题，由于在大脑中的神经元数量少，遗传工具的可用性。神经manipul的BULLETIN。呈现明暗偏好测定允许视觉行为包括昼夜的控制趋光性了一系列的调查。

继上述协议，我们测试了两种不同的昼夜倍CT0和CT12明暗偏好早期幼虫第三体育场野生型广东-S飞。成人12小时光照，12小时黑暗饲养，并留下12小时产卵。幼虫生长的前两天，在相同的光线暗的政权。因为我们想在测试昼夜调制恒定的条件下（自由运行的生物钟），幼虫，然后转移到不断的黑暗未来三天，直到进行测试（ 图3A）。 这里，我们用350勒克司，因为它已...

Watch this Scientific Journal Video about Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae at JoVE.com

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

在这里，我们描述了果蝇幼虫光暗偏好测试。此法提供了先天和昼夜节律调节的光传感和处理感光特性的信息。

光偏好的检测，以研究的先天和昼夜调节感光特性<em&gt;果蝇</em&gt;幼虫

Light acts as environmental signal to control animal behavior at various levels. The Drosophila larval nervous system is used as a unique model to answer ...

Research

JoVE Journal

Neuroscience

Light Preference Assay to Study Innate and Circadian Regulated Photobehavior in Drosophila Larvae

Laser Capture Microdissection of Drosophila Peripheral Neurons

Laser Capture Microdissection of Drosophila Peripheral Neurons

激光捕获显微切割技术的果蝇外周神经元

The dendritic arborization (da) neurons of the Drosophila peripheral nervous system (PNS) provide an excellent model system in which to investigate the ...

科研

神经科学

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

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

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) ...

Measuring Circadian and Acute Light Responses in Mice using Wheel Running Activity

测量小鼠使用车轮运行活动的昼夜和急性对光反应

Circadian rhythms are physiological functions that cycle over a period of approximately 24 hours (circadian- circa: approximate and diem: day)1, 2.  They ...

Methods to Assay Drosophila Behavior

Methods to Assay Drosophila Behavior

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 ...

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

Local and Global Methods of Assessing Thermal Nociception in Drosophila Larvae

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 ...

Visualization of Proprioceptors in Drosophila Larvae and Pupae

Visualization of Proprioceptors in Drosophila Larvae and Pupae

可视化本体感受器<em&gt;果蝇</em&gt;幼虫和蛹

Proprioception is the ability to sense the motion, or position, of body parts by responding to stimuli arising within the body. In fruitflies and other ...

Appetitive Associative Olfactory Learning in Drosophila Larvae

Appetitive Associative Olfactory Learning in Drosophila Larvae

欲求联想嗅觉学习<em&gt;果蝇</em&gt;幼虫

In the following we describe the methodological  details of appetitive associative olfactory learning in Drosophila larvae. The setup, in combination with ...

Recording and Analysis of Circadian Rhythms in Running-wheel Activity in Rodents

运行轮活动在啮齿类动物的昼夜节律的记录和分析

When rodents have free access to a running wheel in their home cage, voluntary use of this wheel will depend on the time of day1-5. Nocturnal rodents, ...

Foraging Path-length Protocol for Drosophila melanogaster Larvae

Foraging Path-length Protocol for Drosophila melanogaster Larvae

觅食路径长度协议<em&gt;果蝇</em&gt;幼虫

The Drosophila melanogaster larval path-length phenotype is an established measure used to study the genetic and environmental contributions to behavioral ...

Taste Preference Assay for Adult Drosophila

Taste Preference Assay for Adult Drosophila

口味偏好测定成人<em&gt;果蝇</em

Olfactory and gustatory perception of the environment is vital for animal survival.  The most obvious application of these chemosenses is to be able to ...