多单位记录方法来描述蝗虫的神经活动（<em> Schistocerca美洲</em>）嗅觉电路
Summary
我们证明细胞外的多单元的记录技术中的前三个阶段的无脊椎动物的嗅觉路径表征气味诱发反应的变化。这些技术可以很容易地适应以及其他神经系统检查合奏活动。
Abstract
嗅觉检测和解释的许多生物的生存是至关重要的。值得注意的是，跨门类的物种都有着惊人的相似嗅觉系统在进化的时间1，化学传感，生物方法进行了优化。在昆虫的嗅觉系统，增味剂，转嗅觉受体神经元（ORN）的天线，这将化学刺激的动作电位的列车。从ORNs的感官输入，然后转发至触角叶（AL;类似于脊椎动物的嗅球结构）。在AL，神经表征气味采取的形式分布在歌舞团的主要神经元（PNS，也被称为投射神经元）2,3的时空放电模式。随后处理的AL输出由肯扬细胞（角质形成）在下游蘑菇体（MB），相关联的结构与嗅觉记忆和学习4,5。她的E，我们提出了电生理记录技术，在这些嗅觉电路的监控气味诱发的神经反应。
首先，我们提出一个单一的感受器记录的方法来研究人口的ORNs 6,7气味诱发反应。我们讨论使用生理盐水填充尖锐的玻璃吸管作为电极细胞外监控ORN反应。接下来，我们提出了一种方法来使用商业16通道的电极细胞外PN反应。类似的方法使用一个定制的8路双绞线四极管展示了肯扬细胞记录8。我们提供详细的实验装置，目前这些技术代表记录的痕迹。
Protocol
1。气味准备工作和实施稀释在矿物油中的气味的解决方案（体积），以达到所需的浓度水平。存储一个20毫升的混合物，矿物油和的加臭剂，在60毫升的玻璃瓶。成的橡胶止动件（表19），一个从底部和其他从顶部插入两个注射器针头，以提供一个入口和一个出口管线。该橡胶塞密封的玻璃瓶中，并附加一个自定义设计的活性炭过滤器的入口线（ 图1A）。 的碳过滤器是?…
Representative Results
在图3D中示出一个单一的ORN两种不同醇的气味诱发反应。根据记录位置（放置的电极）的多单元录音感器类型，可以实现的。 图6A中所示的原料外从AL记录波形。可以观察到的动作电位或尖峰的幅度变化，来自不同的PN，在此电压跟踪。虽然蝗虫触角叶投射神经元有兴奋和抑制神经元，唯一的PN生成钠尖峰，可以检测到细胞外3。这一观察表明?…
Discussion
大多数感官刺激，唤起组合分布在歌舞团的神经元的反应。因此，同时监控多神经元活动是有必要了解如何刺激特定的信息在大脑中的神经回路表示和处理。在这里，我们已经证明细胞外多单位记录技术的气味诱发反应的特点前三加工中心以及昆虫嗅觉通路。我们注意到，这里所介绍的技术已嗅觉编码的以前的研究中使用的一些，并成为一个标准的实践在此字段中3,6,13-17。相结合的技术?…
Declarações
The authors have nothing to disclose.
Acknowledgements
作者想感谢以下为这项工作提供资金：大方的启动资金从生物医学工程系，美国华盛顿大学，麦中心系统神经科学经费，办公室美国海军研究资助（批准编号：N000141210089）到BR
Materials
| Name | Company | Catalog Number | Comments |
| Electrophysiology Equipment | |||
| A.C. amplifier | GRASS | Model P55 | for single sensillum recordings |
| Audio monitor (model 3300) | A-M Systems | 940000 | |
| Custom-made 16 channel pre-amplifier and amplifier | Cal. Tech. Biology Electronics Shop | for AL and MB recordings | |
| Data acquisition unit | National Instruments | BNC-2090 | |
| Fiber optic light | WPI | SI-72-8 | |
| Light source 115 V | WPI | NOVA | |
| Manual micromanipulator | WPI | M3301R | for locust brain recordings |
| Stereomicroscope1 on boom stand | Leica | M80 | for locust brain recordings |
| Stereomicroscope2 | Leica | M205C | for single sensillum recordings |
| Vibration-isolation table | TMC | 63-500 series | |
| Motorized micromanipulator | Sutter Instruments | MP285/T | |
| Oscilloscope | Tektronix | TD2014B | |
| Electrodes/Construction Tools | |||
| 16-channel electrode | NeuroNexus | A2x2-tet-3mm-150-121 | for antennal lobe recordings |
| Borosilicate capillary tubes with filament, ID 0.69 mm | Sutter Instruments | BF120-69-10 | for making glass electrodes |
| Micropipette puller | Sutter Instruments | P-1000 | |
| Function generator | Multimeter Warehouse | SG1639A | for gold-plating electrodes |
| Gold plating solution (non cyanide) | SIFCO Industries | NC SPS 5355 | |
| Impedance tester | BAK Electronics Inc. | IMP-2 | for gold-plating electrodes |
| Switch rotary | Electroswitch | C7D0123N | for gold-plating electrodes |
| Pulse isolator | WPI | A365 | for gold-plating electrodes |
| Q series electrode holder | Warner Instruments | 64-1091 | |
| Silver wire 0.010″ diameter | A-M Systems | 782500 | ground electrode |
| 8 pin DIP IC socket | Digikey | ED90032-ND | |
| Borosilicate capillary tubes with filament, ID 0.58 mm | Warner Instruments | 64-0787 | twisted wire tetrode construction |
| Heat gun | Weller | 6966C | |
| Rediohm-800 wire | Kanthal Precision Technologies | PF002005 | |
| Titer plate shaker | Thermo Scientific | 4625Q | twisting wires |
| Carbide scissors, 4.5″ | Biomedical Research Instr | 25-1000 | for cutting twisted tetrode wires |
| Fine point tweezers | HECO | 91-EF5-SA | for teasing tetrode wires apart |
| Odor Delivery | |||
| 6 ml syringe | Kendall | 1180600777 | for custom designed activated carbon filter |
| Brown odor bottles | Fisher | 08-912-165 | |
| Charcoal | BuyActivatedCharcoal.com | GAC-48C | |
| Desiccant | Drierite | 23005 | |
| Drierite gas drying jar | Fischer Scientific | 09-204 | |
| Heat shrink tubing | 3M | EPS-200 | odor filter preparation |
| Hypodermic needle aluminum hub, gauge 19 | Kendall | 8881-200136 | for providing inlet and outlet lines for odor bottles |
| Mineral oil | Mallinckrodt Chemicals | 6357-04 | for odor dilution |
| Nalgene plastic tubing, 890 FEP | Thermo Scientific | 8050-0310 | for carrier gas delivery |
| Pneumatic picopump | WPI | sys-pv820 | for odor delivery |
| Polyethylene tubing ID 0.86 mm | Intramedic | 427421 | for odor bottle outlet connections and saline profusion tubing |
| Stoppers | Lab Pure | 97041 | for sealing odor bottles |
| Time tape | PDC | T-534-RP | |
| Tubing luer | Cole-Parmer | 30600-66 | |
| Vacuum tube | McMaster-Carr | 5488K66 | |
| Preparation/Dissection | |||
| 100 x 15 mm petri dish | VWR International | 89000-304 | |
| 18 AWG copper stranded wire | Lapp Kabel | 4510013 | wire insulation is used as rubber gaskets |
| 22 AWG stranded hookup wire | AlphaWire | 1551 | brain platform |
| Batik wax | Jacquard | 7946000 | |
| Dental periphery Wax | Henry-Schein Dental | 6652151 | |
| Electrowaxer | Almore International | 66000 | |
| Epoxy, 5 min | Permatex | 84101 | |
| Hypodermic needle aluminum hub | Kendall | 8881-200136 | |
| Protease from Streptomyces griseus | Sigma-Aldrich | P5147 | for desheathing locust brain |
| Suture thread non-sterile | Fisher | NC9087024 | for tying the abdomen after gut removal |
| Vetbond | 3M | 1469SB | for sealing amputation sites |
| Dumont #1 forceps (coarse) | WPI | 500335 | |
| Dumont #5 titanium forceps (fine) | WPI | 14096 | |
| Dumont #5SF forceps (super-fine) | WPI | 500085 | desheathing locust brain |
| 10 cm dissecting scissors | WPI | 14393 | for removing legs and wings |
| Vannas scissors (fine) | WPI | 500086 | for removing cuticle, cutting the foregut |
| Saline Profusion | |||
| Extension set with rate flow regulator | Moore Medical | 69136 | for regulating saline flow |
| IV administration set with Y injection site | Moore Medical | 73190 | for regulating saline flow |
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Citar este artigo
Saha, D., Leong, K., Katta, N., Raman, B. Multi-unit Recording Methods to Characterize Neural Activity in the Locust (Schistocerca Americana) Olfactory Circuits. J. Vis. Exp. (71), e50139, doi:10.3791/50139 (2013).