Name: preparation of drosophila central neurons for in situ patch clamping
Uploaded: 2012-10-15T13:00:00
Description: Watch this Scientific Journal Video about Preparation of Drosophila Central Neurons for in situ Patch Clamping Video at JoVE.com

下面的描述是不特定的一个运动神经元。它可以用于与任何神经元。在这个例子中，我们使用的飞行运动神经元（MN5）支配抑制剂肌肉dorsolongitudinal翼（DLM）的的2 dorsalmost纤维。为了识别和可视化MN5我们使用UAS/GAL4系统表达GFP的飞行运动神经元（和其他一些神经元）。 1。解剖成年果蝇访问背部分的腹神经索（VNC） <ol><li>解剖成年果蝇背侧沿其背中线在Sylgard的?...

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当可视化的细胞绿色荧光蛋白的荧光蛋白，如，重要的是不超过制剂暴露太多的光。这可能会导致光损伤。我们使用100W HBO短弧汞灯泡，照明，我们还可以使用中性密度（ND）的0.8（的色度ND过滤器0.3和0.5）。为了能够判断清洗良好的能见度的质量是至关重要的。因此，ND过滤器可以去除约20秒，多次短时间。 应用了一些积极的压力时，细胞体“襟翼”一点。这有助于判断质量?...

<table border="1"><tbody><tr><td> 代理/项目 </td><td> 公司 </td><td> 目录号 </td></tr><tr><td>蛋白酶型第十四</td><td>美国Sigma Aldrich公司</td><td> P5147 </td></tr><tr><td> Microfil灵活的注射针</td><td>美国世界精密仪器</td><td> MF28G-5 </td></tr><tr><td>高硼硅玻璃毛细管，外径1.5毫米，1.0毫米，无灯丝ID </td><td>美国世界精密仪器</td><td> PG52151-4 </td></tr><tr><td> DiI荧光</td><td> Invitrogen公司的美国</td><td> D3899 </td></tr><tr><td> Sylgard的弹性体套装184（道康宁） </td><td> <a href="http://www.ellsworth.com" target="_blank">www.ellsworth.com</a> </td><td> 184 SIL ELAST套件</td></tr><tr><td> ND过滤器集（卸载） </td><td>浓度</td><td> 22000b系列</td></tr><tr><td>电极架1-HL-U </td><td>莫分子器件</td><td> 1，HL-U </td></tr></tbody></table>

preparation of drosophila central neurons for in situ patch clamping

短的世代时间和轻便的基因技术使果蝇优良的遗传模型的基础神经科学的研究。离子通道是神经元兴奋性的基础上，所有的行为，因为他们调解。电压门控离子通道的克隆果蝇的电压门控钾通道振动筛1,2。对理解离子通道和细胞膜的兴奋性神经系统功能的作用，它是有用的，结合强大的遗传工具，可以在果蝇在原位膜片钳记录。多年来，这样的录音已经阻碍了果蝇的中枢神经系统的小尺寸。此外，强大的鞘的神经胶质细胞和胶原蛋白构成障碍补丁吸液管进入中央神经元。本套去除任何在成年果蝇中枢神经系统的神经元膜片钳记录的一个必要的先决条件。近年来科学家们一直在能够进行原位膜片钳记录神经元在成人大脑3,4和腹神经索的胚胎，幼虫7,8,9,10，5,6和成年果蝇 11,12,13,14。一个稳定的千兆密封是一个很好的补丁的主要的先决条件，并取决于与细胞膜，以避免漏电流的补丁移液器的清洁接触。因此，从成年果蝇神经元全细胞原位膜片钳记录，必须彻底清洗。神经节鞘在第一步骤中，将被处理酶，并机械地除去，使靶细胞可访问。在第二步骤中，细胞膜具有进行研磨，使得没有神经胶质细胞，胶原蛋白或其他材料的层，可能会扰乱千兆密封件形成。本文介绍了如何编写一个确定的中央神经元在果蝇的腹神经索，飞行运动神经元的5（MN5 15），体细胞整个C埃尔膜片钳记录。识别和知名度的神经元是通过有针对性的表达GFP MN5。我们不打算解释膜片钳技术。

Watch this Scientific Journal Video about Preparation of Drosophila Central Neurons for in situ Patch Clamping Video at JoVE.com

Preparation of Drosophila Central Neurons for in situ Patch Clamping Video (Video) | JoVE

原位补丁钳用于神经元的电生理特性的完整电路。在果蝇的遗传模型膜片钳是困难的，因为中枢神经系统，周围环绕着一个强大的护套。本文介绍的步骤，取出护套和清洁随后的膜片钳记录神经元。

制备<em&gt;果蝇</em&gt;中央神经元<em在原位</em&gt;膜片钳

Short generation times and facile genetic techniques make the fruit fly Drosophila melanogaster an excellent genetic model in fundamental neuroscience ...

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Preparation of Drosophila Central Neurons for in situ Patch Clamping

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