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Biologie

在电生理记录果蝇胚胎

Published: May 21, 2009
doi:
10.3791/1348
, ,
1Department of Biological Sciences,University of Illinois, 2Department of Biological Sciences,Vanderbilt University

Summary

从电生理记录<em>果蝇</em>胚胎允许发展肌肉和神经元的电性能，以及在谷氨酸能神经肌肉接头及中枢胆碱能和GABA能突触功能突触的特性分析。

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。全细胞膜片钳技术，然后可以记录单独识别的神经元和躯体肌肉。这些记录配置已被用来跟踪的外观和成熟的离子流和行动中的神经元和肌肉的潜在传播。影响这些电气特性的遗传突变体进行了表征，揭示了离子通道及相关的信号复合物的分子组成，并开始探索功能分化的分子机制。一个特别的重点一直是突触连接的大会，无论是在中枢神经系统和外围。谷氨酸能神经肌肉接头（NMJ）是最容易结合光学成像和电生理记录。玻璃吸电极是用于刺激周围神经兴奋结的电流电压钳位肌肉（EJC）录制。此记录已配置使用，以图表的突触功能的分化，并跟踪突触前谷氨酸释放属性的外观和成熟。此外，突触后性能可以独立检测，直接通过谷氨酸iontophoretic或施加压力的肌肉表面，测量谷氨酸受体字段的外观和成熟。因此，可以监视元素前和突触后单独或合并在胚胎突触。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 果蝇胚胎的电生理记录，首先需要在胚胎夹层技术的能力，这是在另一朱庇特的视频。 果蝇胚胎的电生理记录使用标准的膜片钳记录配置。膜片钳记录设备和软件适用于许多其他准备工作也适合记录果蝇胚胎。由于果蝇胚胎粘过程中剥离的盖玻片，录音室应接受盖玻片。编制和方式，它是安装在盖玻片厚度需要一个优秀的DIC的光学?…

Discussion

果蝇胚胎的电生理记录需要手工操作和解剖。该制剂的健康，以及随之而来的录音质量，取决于一个能够迅速而整齐地准备录制脆弱的胚胎组织，然后执行实验。实验者应精通胚胎解剖和膜片钳电，然后再尝试一次都解决。

录音可以做的“修订标准”或“血淋巴样”（HL），salines几个变种之一。这两个生理盐水类之间的主要区别是：1）Na +浓度（与70毫米HL 120-…

Divulgations

The authors have nothing to disclose.

Acknowledgements

KB是由美国国立卫生研究院授予GM54544支持。

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Tags

Electrophysiological RecordingDrosophila EmbryoGenetic ModelEmbryonic DevelopmentFunctional NeuroscienceDevelopmental ProgramsFunctional Electrical Signaling PropertiesFunctional Chemical SynapsesNeural Circuit FormationLate-stage EmbryogenesisEpidermal CuticleWater-polymerizing Surgical GlueControlled DissectionDorsal Longitudinal IncisionVentral Nerve CordBody Wall MusculatureWhole-cell Patch-clamp TechniquesIndividually-identifiable NeuronsSomatic MusclesIonic CurrentsAction Potentials
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Chen, K., Featherstone, D. E., Broadie, K. Electrophysiological Recording in the Drosophila Embryo. J. Vis. Exp. (27), e1348, doi:10.3791/1348 (2009).
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