Name: measurement of metabolic rate in drosophila using respirometry
Uploaded: 2014-06-24T15:00:00
Description: Watch this Scientific Journal Video about Measurement of Metabolic Rate in Drosophila using Respirometry at JoVE.com

我们要感谢德国马普学会为我们的研究提供资金。

呼吸计1。准备<ol><li>削减1,000微升枪头，用刀片，允许插入50微升毛细管微量的，试图让枪头尽量伸直。 </li><li>将一块泡沫进入吸管和推下来的枪头。 </li><li>加入少量的CO 2吸收剂的和由第二片泡沫的包含它。 </li><li>在其中微量插入移液管尖端的地方涂上胶水。 </li><li>离开呼吸过夜，让胶水干。 一个呼吸的原理图如图1A所示。 </li></ol> 2，准备测量室<...

<ol>
	<li>Van Voorhies, W. A., Khazaeli, A. A., Curtsinger, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Testing+the+&amp;#34;rate+of+living&amp;#34;+model:+further+evidence+that+longevity+and+metabolic+rate+are+not+inversely+correlated+in+Drosophila+melanogaster.">Testing the &amp;#34;rate of living&amp;#34; model: further evidence that longevity and metabolic rate are not inversely correlated in Drosophila melanogaster.</a> J Appl Physiol. 97, 1915-1922 (2004).</li><li>Ross, R. E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Age-specific+decrease+in+aerobic+efficiency+associated+with+increase+in+oxygen+free+radical+production+in+Drosophila+melanogaster.">Age-specific decrease in aerobic efficiency associated with increase in oxygen free radical production in Drosophila melanogaster.</a> Journal of Insect Physiology. 46, 1477-1480 (2000).</li><li>van Voorhies, W. A., Khazaeli, A. A., Curtsinger, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Selected+contribution:+long-lived+Drosophila+melanogaster.+lines+exhibit+normal+metabolic+rates.">Selected contribution: long-lived Drosophila melanogaster. lines exhibit normal metabolic rates.</a> J Appl Physiol. 95, 2605-2613 (2003).</li><li>Hulbert, A. J., 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=Metabolic+rate+is+not+reduced+by+dietary-restriction+or+by+lowered+insulin/IGF-1+signalling+and+is+not+correlated+with+individual+lifespan+in+Drosophila+melanogaster.">Metabolic rate is not reduced by dietary-restriction or by lowered insulin/IGF-1 signalling and is not correlated with individual lifespan in Drosophila melanogaster.</a> Experimental Gerontology. 39, 1137-1143 (2004).</li><li>Ueno, T., Tomita, J., Kume, S., Kume, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dopamine+modulates+metabolic+rate+and+temperature+sensitivity+in+Drosophila+melanogaster.">Dopamine modulates metabolic rate and temperature sensitivity in Drosophila melanogaster.</a> PLoS ONE. 7, (2012).</li><li>Takeuchi, K., 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=Changes+in+temperature+preferences+and+energy+homeostasis+in+dystroglycan+mutants.">Changes in temperature preferences and energy homeostasis in dystroglycan mutants.</a> Science. 323, 1740-1743 (2009).</li><li>Kucherenko, M. M., Marrone, A. K., Rishko, V. M., Magliarelli Hde, F., Shcherbata, H. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Stress+and+muscular+dystrophy:+a+genetic+screen+for+dystroglycan+and+dystrophin+interactors+in+Drosophila.+identifies+cellular+stress+response+components.">Stress and muscular dystrophy: a genetic screen for dystroglycan and dystrophin interactors in Drosophila. identifies cellular stress response components.</a> Developmental Biology. 352, 228-242 (2011).</li><li>Marrone, A. K., Kucherenko, M. M., Wiek, R., Gopfert, M. C., Shcherbata, H. R. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Hyperthermic+seizures+and+aberrant+cellular+homeostasis+in+Drosophila+dystrophic.+muscles.">Hyperthermic seizures and aberrant cellular homeostasis in Drosophila dystrophic. muscles.</a> Scientific Reports. 1, 47 (2011).</li><li>Khazaeli, A. A., Van Voorhies, W., Curtsinger, J. W. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Longevity+and+metabolism+in+Drosophila+melanogaster:+genetic+correlations+between+life+span+and+age-specific+metabolic+rate+in+populations+artificially+selected+for+long+life.">Longevity and metabolism in Drosophila melanogaster: genetic correlations between life span and age-specific metabolic rate in populations artificially selected for long life.</a> Genetics. 169, 231-242 (2005).</li><li>Elia, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Energy+equivalents+of+CO2+and+their+importance+in+assessing+energy+expenditure+when+using+tracer+techniques.">Energy equivalents of CO2 and their importance in assessing energy expenditure when using tracer techniques.</a> The American Journal of Physiology. 260, (1991).</li><li>Schindelin, J., 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=Fiji:+an+open-source+platform+for+biological-image+analysis.">Fiji: an open-source platform for biological-image analysis.</a> Nature Methods. 9, 676-682 (2012).</li><li>Bharucha, K. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+epicurean+fly:+using+Drosophila+melanogaster.+to+study+metabolism.">The epicurean fly: using Drosophila melanogaster. to study metabolism.</a> Pediatric Research. 65, 132-137 (2009).</li><li>Rajan, A., Perrimon, N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Of+flies+and+men:+insights+on+organismal+metabolism+from+fruit+flies.">Of flies and men: insights on organismal metabolism from fruit flies.</a> BMC Biology. 11, 38 (2013).</li></ol>

在这个协议中，我们描述了一种廉价，可靠的方法，用于测量CO 2的生产中的苍蝇。我们发现，这个实验是容易的，能够快速进行，并产生可再现的数据是在与其他研究1，6,9协议。这里列出的协议，可以很容易地修改，以适应任何实验室的预算和可用的材料。每个个体呼吸的结构可以只要该室保持气密进行调整。然而，更长，更薄微量提供比短的更精确。只要没有显著环境温度和?...

生化的Kreb的周期产生的ATP通过乙从碳水化合物，脂肪，以及产生CO 2的蛋白衍生的氧化。在果蝇中 ，O 2的输入是直接相关的CO 2输出，并反映代谢1的电平。因此，CO 2输出的测量已成功应用于涉及到老龄化和新陈代谢2-5的研究。在这里，我们的实验室已修改先前设计的实验装置，使CO 2的产量可达18样品中，而不需要任何专门的设备测量。其他和我们先前已经用这种方法来显示代谢率有缺陷的肌营养不良症相关蛋白，Dystroglycan基因（DG）6-8差异苍蝇。 O用于氧化代谢2转化成CO 2，这是因为排出的呼吸废物。营建管理手工制作的呼吸计化描述，它允许对O 2的消耗速率的测定。苍蝇被放置在一个密闭容器内利用吸收排出的CO 2的物质，从气相有效消除它。在气体体积（减小压力）的变化是由流体的附着在封闭式呼吸的玻璃毛细管的位移测量。 这种技术比其他的主要优点是成本。以前的研究已经测量的CO 2产量果蝇使用气体分析仪及技术先进的呼吸测量系统1,9。尽管更复杂的设备，在这里所描述的方法的灵敏度是类似于报告的值（ 表1）。此外，其他几个小组已经使用这种技术的变化，以确定相对代谢率在果蝇 4-6。因此，该测定法可以用来生成reliab乐，有关果蝇的代谢没有购买专门设备，可以设定在任何实验室，可用于教育目的重复性的数据。 一般而言，接受的技术，以确定生物体的新陈代谢是测量CO 2的生产时，O 2消耗，或两者3,4,9。不过，它可以假定1当量的O 2产生1当量的CO 2，CO 2的精确率，生成依赖的代谢底物利用10上。因此，为了准确地确定代谢率在能量单位，有必要同时测量O 2的消耗和CO 2的产生。由于这个原因，这里描述的方法是特别相关的比较，在动物中，而不是绝对值之间的CO 2产量的差异。我们的技术集成了多种动物的CO 2生产过一段恬E（1-2小时），因此返回的平均动物的活动。如果有理由相信，本实验的动物比对照动物少活性的测定可以反映不同水平的活性，而不一定代谢。

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measurement of metabolic rate in drosophila using respirometry

代谢紊乱是一个常见的​​问题影响到人类的健康。因此，了解调节新陈代谢机制是一个重要的科学任务。在人类许多疾病的致病基因有一只苍蝇同源，使果蝇的研究涉及不同疾病的发展信号通路的良好模型。此外， 果蝇的可追踪性简化了遗传筛选确定了可调节新陈代谢新的治疗靶点提供帮助。为了执行这样一个屏幕的简单和快速的方法，以确定变化苍蝇的代谢状态是必要的。在一般情况下，二氧化碳是生产的底物氧化和能量消耗，提供约代谢状态的信息的良好指标。在这个协议中，我们介绍一个简单的方法来测量从苍蝇的CO 2的输出。该技术可以潜在地有助于影响代谢率的遗传扰动的识别。

为了证明该方法是我们测量的CO 2产量从野生型（ 俄勒冈州R）的雄蝇在18，25，和29°C和苍蝇突变为危险品敏感。苍蝇提出了在25℃，然后转移到实验温度连续5天前测量。如预期此变温动物物种，CO 2的生成量随温度的升高（ 图2）。我们已经在过去表明，无糖饮食降低了野生型的代谢率和DG突变果蝇7。 DG的损失导致增加新陈代谢水平?...

Watch this Scientific Journal Video about Measurement of Metabolic Rate in Drosophila using Respirometry at JoVE.com

Measurement of Metabolic Rate in Drosophila using Respirometry

代谢率的测量<em&gt;果蝇</em&gt;使用呼吸运动

代谢紊乱都属于在人体中的最常见的疾病之一。该基因听话的模式生物D.果蝇可用于鉴定调节代谢的新基因。本文介绍一种比较简单的方法，它允许通过测量它们的CO 2的生产研究代谢率的苍蝇。

Metabolic disorders are a frequent problem affecting human health. Therefore, understanding the mechanisms that regulate metabolism is a crucial ...

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Measurement of Metabolic Rate in Drosophila using Respirometry

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