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Biology

收获和准备果蝇胚胎的电生理记录和其他程序

Published: May 20, 2009
doi:
10.3791/1347
, ,
1Department of Biological Sciences,University of Illinois, 2Department of Biological Sciences,Vanderbilt University

Summary

这种技术公开的果蝇胚胎neuromusculature免疫组织化学或电生理记录。它是有用的，为研究神经肌肉发展的早期事件，或执行中的突变体，不能孵化电。

Abstract

果蝇胚胎发育和功能的神经科学的研究是首屈一指的遗传模型。传统上，这些领域都相当彼此孤立的，很大程度上是独立的历史和科学界。然而，这些通常是不同的领域之间的接口是发展计划的基本功能的电子信号的属性和功能的化学突触在神经回路形成的最后阶段分化的收购。该接口是一个极为重要的领域进行调查。在果蝇中，这些功能开发阶段发生在胚胎发育的最后三分之一的<8小时期间（在25 ° C） 。后期发展时期，长期被视为棘手的调查，由于一个艰难的，不透水的表皮角质层的沉积。一个突破性的进展是，可以在本地应用的角质层，使后期胚胎的控制清扫手术，聚合水胶中的应用。胚胎背侧纵切口，可以平放，露出腹侧神经线和体壁肌肉的实验研究。该系统已大量用于分离和鉴定基因突变体，损害胚胎的突触的形成，从而揭示了突触连接和功能的突触信号特性的规范和分化的分子机制。

Protocol

第1部分：设备和用品一个良好的解剖镜下胚胎解剖;建议40X放大倍率25X眼睛件，最大限度地增加放大。 胚胎及胚胎devitellinization手动选择需要精细镊子（5号）。 制定和修改细玻璃针的设备是必需的。拉玻璃针都需要剥离。我们喜欢实心玻璃夹层（持续更长的时间），但工程，以及标准的厚壁玻璃管（外径1-1.5毫米）。有些人用细的钨针，电解所需的清晰度在1M的NaOH使用自耦变…

Discussion

胚胎的精确分期是由于功能特性的只是几个小时的时间当然的迅速成熟的关键。几个问题复杂化分期。首先，大多数研究人员使用定时鸡蛋奠定阶段的胚胎，但产蛋时间可以千差万别，在不同的条件（Broadie等，1992）在动物。特别是，在有限的食物的女性倾向于保留奠定前长时间的受精卵。因此，至关重要的丰富的酵母饮食喂养前至少2天收集蛋（Broadie等，1992）“明确的”女性。此外，老年女性?…

Acknowledgements

KB是由美国国立卫生研究院授予GM54544支持。

Materials

Material Name Type Company Catalogue Number Comment
Embryo Collection Cages   Genesee Scientific (www.flystuff.com) 59-100 (for 60 mm dish; other sizes available) Cages can also be home made using punctured tri-pour beakers, as shown in video
Sylgard 184   Dow Corning Available from various companies Surgical glue adheres better to sylgard-coated coverslips
Fine glass tubing, outer diameter 1.0-1.5 mm   various   For pulling into fine glass needles for dissection and tubes for glue delivery
Plastic tubing, to attach to glass pipette for mouth suction and glue application   Tygon   Tubing inner diameter needs to match glass outer diameter.
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DrosophilaEmbryosElectrophysiological RecordingProceduresGenetic ModelEmbryonic DevelopmentFunctional NeuroscienceDevelopmental ProgramsElectrical Signaling PropertiesChemical SynapsesNeural Circuit FormationInvestigationLate Developmental PeriodEpidermal CuticleSurgical GlueControlled DissectionDorsal Longitudinal IncisionVentral Nerve CordBody Wall MusculatureGenetic MutantsSynapse FormationMolecular Mechanisms
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Featherstone, D. E., Chen, K., Broadie, K. Harvesting and Preparing Drosophila Embryos for Electrophysiological Recording and Other Procedures. J. Vis. Exp. (27), e1347, doi:10.3791/1347 (2009).
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