Name: electrophysiological recording from drosophila labellar taste sensilla
Uploaded: 2014-02-26T23:00:00
Description: Watch this Scientific Journal Video about Electrophysiological Recording From Drosophila Labellar Taste Sensilla at JoVE.com

这项工作是由一个NRSA博士前补助1F31DC012985（到RD）和美国国立卫生研究院资助，以支持JC 我们要感谢琳内亚魏斯博士的帮助编制数字上的稿子，约瑟夫·莱恩博士有益的意见和弗雷德里克博士马里昂轮询的有益的技术咨询。我们也想感谢的四审稿意见非常有用。

以下协议符合美国耶鲁大学的所有动物护理指引。 1。试剂和设备的准备<ol><li>录音设备设置（ 图1A）。 <ol><li fo:keep-together.within-page="always">选择是无大的变化温度或湿度，并从分离的电气和机械噪声，如冰箱和离心机的来源一个房间钻机安装。 <img alt="图1" fo:content-width="5in" fo:src="/files/ftp_upload/51355/51355fig1highres.jpg" src="/files/ftp_upload/51355/51355fig1.jpg" /> <str...

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Labellar感受器由于形态和解剖组织的差异而有所不同在便于记录。有时候感器不响应任何促味剂，连一个已知可引起积极的回应。取决于感器类型与发生这种情况的频率而变化。 L感器是最一致响应和相对容易访问，因为它们的长度。一般来说，S感器是一致响应，但他们的短长并在唇瓣位置好好接触挑战性。余感器可以更容易地访问，这取决于制剂的角度，但是，它们更频繁响应。在任何给定的?...

味觉允许的昆虫以检测可溶性的化学物质种类繁多并且起着一种营养物质的接受了重要的作用，或一种有毒或毒性1的抑制。味道也被认为在择偶中发挥作用，通过信息素1-5的检测。这些重要而多样化的功能所做的昆虫味觉系统调查的令人信服的目标进入感觉系统如何翻译环境因素纳入有关的行为输出。 果蝇味觉系统的主要单位是味道的头发，或感器。分子通过孔在其尖端2,6进入感器。感器上发现的唇瓣，腿，翅缘，咽6。在唇瓣，感受器的数量和位置是千篇一律。有感器基于长度3形态类：长（L），中间体（I），和短（S<html> ）感器7,8。每个感器包含两个（I-型）或4（L-和S-型）味觉受体神经元（GRNS）9。不同GRNS应对不同类别的味觉刺激的：苦，糖，盐和渗透压7,10和表达味觉受体8,11-13的不同子集。只有我和S型感器含有苦响应GRNS 8,10。该GRNS项目的subesophageal神经节（SOG）和其活化味分子被传递到用于解码的更高中枢神经系统，从而导致行为反应6。数量相对较少的神经元和顺从分子和行为分析使果蝇味觉系统的味觉系统的一般调查一个很好的模型。相对难易程度，系统可以通过基因突变或GAL4-UAS表达系统来操作也可作为一种有价值的工具14,15。 ontent“&gt;因为这些感受器从唇瓣的表面突出，它们使电生理学优良的目标。的GRNS的烧成可以使用外记录进行监控。历史上，侧壁的记录方法，其使用插入玻璃电极感器来记录神经细胞的活动，26已被使用，但是，这种方法在技术上是挑战性的执行，并且难以进行长时间录制从每个制备的针尖记录方法，其测量神经元的响应与电极的那同时提供了一个促味，从此成为首选9,16的方法，它已被用来研究果蝇 8,10,17,18的味觉系统，以及其他一些昆虫种类19-23，它有由tastePROBE放大器的发展，这克服了针尖记录方法的主要缺点之一通过补偿得到了极大的便利参比电极和对昆虫感器之间的大的电势差，使得没有过多的放大或过滤24要被记录的GRN动作电位。另一个重要发展是采用tricholine柠檬酸作为记录电解质25。台泥抑制反应的渗透压敏感GRN和不刺激盐敏感GRN，使得由苦和糖味质生成的反应更容易进行分析25。 在这里，我们描述了如何在目前卡尔森实验室进行的果蝇 labellar感受器尖记录。该协议将解释如何建立一个合适的电钻机，如何准备飞，以及如何执行味道录音。我们也当使用这种出示由果蝇感器的子集，以及一些常见的问题和可能遇到的潜在解决方案的记录得出了一些有代表性的数据技术。

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			<td height="84" style="height:84px;width:168px;">Stereo Zoom Microscope</td>
			<td style="width:187px;">Olympus&#160;</td>
			<td style="width:211px;">SZX12 DFPLFL1.6x PF eyepieces: WHN10x-H/22</td>
			<td style="width:219px;">capable of ~150x magnification with long working distance table mount stand</td>
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		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Anti-vibration Table</td>
			<td style="width:187px;">Kinetic Systems</td>
			<td>BenchMate2210</td>
			<td></td>
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			<td height="21" style="height:21px;width:168px;">Micromanipulators</td>
			<td style="width:187px;">Narishige</td>
			<td>NMN-21</td>
			<td></td>
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			<td height="21" style="height:21px;width:168px;">Magnetic stands</td>
			<td style="width:187px;">ENCO</td>
			<td>Model #625-0930</td>
			<td></td>
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		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Reference Electrode Holder</td>
			<td style="width:187px;">Harvard Apparatus</td>
			<td style="width:211px;">ESP/W-F10N</td>
			<td>Can be mounted on 5ml serological pipette for extended range</td>
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			<td height="42" style="height:42px;width:168px;">&#160;Silver Wire</td>
			<td style="width:187px;">World Precision Instruments</td>
			<td style="width:211px;">AGW1510</td>
			<td>0.3-0.5mm diameter</td>
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			<td height="21" style="height:21px;width:168px;">Retort Stand</td>
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			<td>generic</td>
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			<td height="21" style="height:21px;width:168px;">Outlet Plastic Tube</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>generic, 1cm diameter</td>
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		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Flexible Plastic Tubing</td>
			<td style="width:187px;">Nalgene&#160;</td>
			<td>8000-0060</td>
			<td style="width:219px;">VI grade 1/4 in internal diameter&#160;</td>
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			<td height="21" style="height:21px;width:168px;">500 ml Conical Flask</td>
			<td style="width:187px;"></td>
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			<td>generic,&#160; with side arm</td>
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			<td height="42" style="height:42px;width:168px;">Aquarium Pump</td>
			<td style="width:187px;">Aquatic Gardens</td>
			<td style="width:211px;">Airpump 2000</td>
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			<td height="42" style="height:42px;width:168px;">Fiber Optic Light Source</td>
			<td style="width:187px;">Dolan-Jenner Industries</td>
			<td>Fiber-Lite 2100</td>
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			<td height="21" style="height:21px;width:168px;">White Card/Paper</td>
			<td style="width:187px;">Whatman</td>
			<td style="width:211px;">1001-110</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Digital Acquisition System</td>
			<td style="width:187px;">Syntech</td>
			<td style="width:211px;">IDAC-4</td>
			<td>Alternative: National Instruments NI-6251 &#160;</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Headstage</td>
			<td style="width:187px;">Syntech</td>
			<td style="width:211px;">DTP-1</td>
			<td>Tasteprobe</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Tasteprobe Amplifier</td>
			<td style="width:187px;">Syntech</td>
			<td style="width:211px;">DTP-1</td>
			<td>Tasteprobe</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Alligator Clips</td>
			<td style="width:187px;">Grainger</td>
			<td style="width:211px;">1XWN7</td>
			<td>Any brand is fine</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Insulated Electrical Wire</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>Generic</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Gold Connector Pins</td>
			<td style="width:187px;">World Precision Instruments</td>
			<td align="right" style="width:211px;">5482</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Personal Computer</td>
			<td style="width:187px;">Dell&#160;</td>
			<td style="width:211px;">Vostro</td>
			<td>Check for compatibility with digital acquisition system and software</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Acquisition Software</td>
			<td style="width:187px;">Syntech</td>
			<td style="width:211px;">Autospike</td>
			<td>Autospike works with IDAC-4; alternatively, use Labview with NI-6251</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Aluminum Foil and/or Faraday Cage</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>Electro-magnetic noise shielding</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;"></td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Borosilicate Glass Capillaries</td>
			<td style="width:187px;">World Precision Instruments</td>
			<td style="width:211px;">1B100F-4</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Pipette Puller</td>
			<td style="width:187px;">Sutter Instrument Company</td>
			<td style="width:211px;">Model P-87 Flaming/Brown Micropipette Puller</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;"></td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Beadle and Ephrussi Ringer Solution</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>See recipe in protocol section</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Tricholine citrate, 65%&#160;</td>
			<td style="width:187px;">Sigma</td>
			<td style="width:211px;">T0252-100G</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;"></td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Stereo Microscope</td>
			<td style="width:187px;">Olympus</td>
			<td style="width:211px;">VMZ 1x-4x</td>
			<td style="width:219px;">Capable of 10x-40x magnification</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Ice Bucket</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>Generic</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">p200 Pipette Tips</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>Generic</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Spinal Needle</td>
			<td style="width:187px;">Terumo</td>
			<td style="width:211px;">SN*2590</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">1ml Syringe</td>
			<td style="width:187px;">Beckton-Dickenson</td>
			<td align="right" style="width:211px;">301025</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Fly Aspirator</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>Assembled from P1000 pipette tips, flexible plastic tubing, and mesh</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Modeling Clay</td>
			<td style="width:187px;"></td>
			<td style="width:211px;"></td>
			<td>Generic</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:168px;">Forceps</td>
			<td style="width:187px;">Fine Science Tools By Dumont</td>
			<td style="width:211px;">11252-00</td>
			<td>#5SF (super-fine tips)</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">10ml Syringe&#160;</td>
			<td style="width:187px;">Beckton-Dickinson</td>
			<td align="right" style="width:211px;">301029</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:168px;">Plastic Tubing</td>
			<td style="width:187px;">Tygon</td>
			<td style="width:211px;">R-3603</td>
			<td></td>
		</tr>
	</tbody>
</table>

electrophysiological recording from drosophila labellar taste sensilla

昆虫的外围味道响应可以有力地调查与电生理技术。这里介绍的方法使研究人员直接和定量测量味觉反应，反映了感官输入，昆虫的神经系统从它的环境味觉刺激接收。该协议概述了在利用这项技术的所有关键步骤。在组装的电装备，如选择的必要设备和一个合适的环境记录的关键步骤，圈定。我们还描述了记录作出适当的参考和记录电极如何准备，并促味剂的解决方案。我们详细地描述用于通过插入玻璃参比电极成飞，以固定所述长鼻制备昆虫的方法。我们显示通过味觉神经元响应于一个糖和一个苦味化合物烧制的电脉冲的痕迹。该协议的某些方面为technically具有挑战性的，我们包括可能​​遇到的一些常见的技术挑战，如缺乏信号或噪音过大的系统，和潜在的解决方案广泛的描述。该技术具有局限性，如不能交付时间上复杂的刺激，观察背景发射前夕刺激交付，或使用不溶于水的呈味物质方便。尽管有这些限制，这项技术（包括在协议中引用的微小变化）是一种标准，用于记录果蝇神经元对味道的化合物广泛接受的过程。

图5A示出了一个L形感器的一种糖，蔗糖的反应。同一感器不给苦化合物反应，黄连， 图5B示出了一个I型感器，它包含一个苦响应神经元，显示响应于小檗碱较大的幅度峰值，并响应于蔗糖较小的幅度峰值。 L感器显示一个最小的背景响应溶剂对照，台泥，而我感器显示给TCC几乎没有响应（ 图5）。有关labellar GRNS的盐和水反应的更多信息，请参阅广井10。</...

Watch this Scientific Journal Video about Electrophysiological Recording From Drosophila Labellar Taste Sensilla at JoVE.com

Electrophysiological Recording From Drosophila Labellar Taste Sensilla

本协议描述的由labellar味觉神经元在果蝇发射动作电位反应细胞外记录。

电生理记录从<em&gt;果蝇</em&gt; Labellar味感器

The peripheral taste response of insects can be powerfully investigated with electrophysiological techniques. The method described here allows the ...

Research

JoVE Journal

Neuroscience

Electrophysiological Recording From Drosophila Labellar Taste Sensilla

Electrophysiological Measurements from a Moth Olfactory System

从蛾的嗅觉系统的电生理测量

Insect olfactory systems provide unique opportunities for recording odorant-induced responses in the forms of electroantennograms (EAG) and single ...

科研

神经科学

Simultaneous Recording of Calcium Signals from Identified Neurons and Feeding Behavior of Drosophila melanogaster

Simultaneous Recording of Calcium Signals from Identified Neurons and Feeding Behavior of Drosophila melanogaster

从查明的神经元和取食行为的钙信号的同时记录果蝇</em

To study neuronal networks in terms of their function in behavior, we must analyze how neurons operate when each behavioral pattern is generated.  Thus, ...

Forebrain Electrophysiological Recording in Larval Zebrafish

斑马鱼幼虫的前脑的电生理记录

Epilepsy affects nearly 3 million people in the United States and up to 50 million people worldwide. Defined as the occurrence of spontaneous unprovoked ...

Simultaneous Pre- and Post-synaptic Electrophysiological Recording from Xenopus Nerve-muscle Co-cultures

Simultaneous Pre- and Post-synaptic Electrophysiological Recording from Xenopus Nerve-muscle Co-cultures

同时前和突触后电生理记录<em&gt;爪蟾</em神经肌肉文化

Much information about the coupling of  presynaptic ionic currents with the release of neurotransmitter has been obtained  from invertebrate preparations, ...

Simultaneous Electrophysiological Recording and Calcium Imaging of Suprachiasmatic Nucleus Neurons

Simultaneous electrophysiological and fluorescent imaging recording methods were used to study the role of changes of membrane potential or current in ...

Electrophysiological Recording in the Brain of Intact Adult Zebrafish

在完整的成年斑马鱼大脑电生理记录

Previously, electrophysiological studies in adult zebrafish have been limited to slice preparations or to eye cup preparations and electrorentinogram ...

A Method for Systematic Electrochemical and Electrophysiological Evaluation of Neural Recording Electrodes

对于系统电化学神经记录电极和电生理评价方法

New materials and designs for neural implants are typically tested separately, with a demonstration of performance but without reference to other implant ...

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

Electrophysiological Method for Recording Intracellular Voltage Responses of Drosophila Photoreceptors and Interneurons to Light Stimuli In Vivo

为记录细胞内电压响应电法<em&gt;果蝇</em&gt;光感受器和中间对光刺激<em&gt;在体内</em

Voltage responses of insect photoreceptors and visual interneurons can be accurately recorded with conventional sharp microelectrodes. The method ...

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 ...

Electrophysiological Method for Whole-cell Voltage Clamp Recordings from Drosophila Photoreceptors

Electrophysiological Method for Whole-cell Voltage Clamp Recordings from Drosophila Photoreceptors

全细胞电压钳记录的电生理方法<em&gt;果蝇</em&gt;光感受器

Whole-cell voltage clamp recordings from Drosophila melanogaster photoreceptors have revolutionized the field of invertebrate visual transduction, ...