昆虫机的混合动力系统：一个自由飞翔的甲壳虫无线电远程控制（<em> Mercynorrhina torquata</em>）
Summary
This protocol describes the process of constructing an insect-machine hybrid system and carrying out wireless electrical stimulation of the flight muscles required to control the turning motion of a flying insect.
Abstract
启用无线电数字电子设备的兴起，促使利用小型无线神经肌肉录像机和刺激为研究飞行昆虫的行为。借助该技术使用本协议中所述活虫平台昆虫机混合动力系统的开发。此外，该协议提出的系统结构和用于在不受限制的昆虫评估飞行肌的功能自由飞行的实验程序。为了演示，我们有针对性的第三腋骨片（3AX）肌肉来控制和实现飞行甲虫向左或向右转向。薄银线电极植入上的甲虫两侧的肌肉3AX。这些被连接到一个无线背包的输出（ 即神经肌肉电刺激器）装上甲虫的前胸。肌肉是由交替的刺激侧（左或右），或者改变stimulatio在自由飞行刺激N频点。甲虫转向同侧当肌肉受到刺激并展出，以增加频率的召。植入过程和三维运动捕获相机系统的体积校准需要小心进行，以避免损坏肌肉和丢失标记的轨道，分别。该方法是非常有益的，研究昆虫飞行，因为它有助于揭示感兴趣的飞行肌的功能在自由飞行。
Introduction
An insect-machine hybrid system, often referred to as a cyborg insect or biobot, is the fusion of a living insect platform with a miniature mounted electronic device. The electronic device, which is wirelessly commanded by a remote user, outputs an electrical signal to electrically stimulate neuromuscular sites in the insect via implanted wire electrodes to induce user desired motor actions and behaviors. In the early stages of this research field, researchers were limited to conducting wireless recording of the muscular action of an insect, using simple analog circuits comprised of surface-mounted components1-3. The development of system-on-a-chip technology with radio frequency functionality enabled not only the wireless recording of neuromuscular signals but also the electrical stimulation of the neuromuscular sites in living insects. At present, a built-in radio microcontroller is small enough to be mounted on living insects without causing any obstructions to their locomotion4-13.
The development of the built-in radio microcontroller allows researchers to determine electrical stimulation protocols to induce desired motor actions to control the locomotion of the insect of interest. On the ground, researchers have demonstrated walking control by stimulating the neuromuscular sites of cockroaches4,12,14, spiders15, and beetles16,17. In the air, the initiation and cessation of flight were achieved using different methods such as the stimulation of the optic lobes (the massive neural cluster of a compound eye) in beetles7,9 and brain sub-regions in bees18, whereas turning control has been demonstrated by stimulating the antennae muscles and nervous system of the abdomens in moths11,19 and the flight muscles of beetles7,9,13. In most cases, a built-in radio microcontroller was integrated on a custom-designed printed circuit board to produce a miniature wireless stimulator (backpack), which was mounted on the insect of interest. This allows wireless electrical stimulation to be applied to a freely walking or flying insect. Such a microcontroller-mounted insect is what is referred to as an insect-machine hybrid system.
This study describes the experimental protocols for building an insect-machine hybrid system, wherein a living beetle is employed as the insect platform, and instructs on how to operate the robot and test its flight control systems. The third axillary sclerite (3Ax) muscle was chosen as the muscle of interest for electrical stimulation and demonstration of left or right turning control13. A pair of thin silver wire electrodes was implanted in both the left and right 3Ax muscles. Moreover, a backpack was mounted on the living beetle. The other ends of the wire electrode were connected to the output pins of the microcontroller. The backpack was small enough for the beetle to carry in flight. Thus, this allows an experimentalist to remotely stimulate the muscle of interest of an insect in free flight and investigate its reactions to the stimulations.
Protocol
Representative Results
Discussion
植入过程是重要的，因为它影响了实验的可靠性。电极应在3毫米或取决于甲虫（避免与附近的肌肉接触）的大小更小的深度插入到肌肉。如果电极触摸附近的肌肉，可能由于附近的肌肉的收缩发生不期望的马达动作和行为。两个电极应该很好对齐，以确保不会发生短路。当熔融和使用烙铁回流蜂蜡，该实验者有要小心和焊料尽快，由于肌肉可以通过用高温下长时间接触被燃烧，导致肌肉的故障。…
Disclosures
The authors have nothing to disclose.
Acknowledgements
This material is based on the works supported by Nanyang Assistant Professorship (NAP, M4080740), Agency for Science, Technology and Research (A*STAR) Public Sector Research Funding (PSF, M4070190), A*STAR-JST (The Japan Science and Technology Agency) joint grant (M4070198), and Singapore Ministry of Education (MOE2013-T2-2-049). The authors would like to thank Mr. Roger Tan Kay Chia, Prof. Low Kin Huat, Mr. Poon Kee Chun, Mr. Chew Hock See, Mr. Lam Kim Kheong and Dr. Mao Shixin at School of MAE for their support in setting up and maintaining the research facilities. The authors thank Prof. Michel Maharbiz (U.C. Berkeley) his advice and discussion, Prof. Kris Pister and his group (U.C. Berkeley) for their support in providing the GINA used in this study.
Materials
| Mecynorrhina torquata beetle | Kingdom of Beetle Taiwan | 10 g, 8 cm, pay load capacity is 30% of the body mass Aproval of importing and using by Agri-Food and Veterinary Authority of Singapore (AVA; HS code: 01069000, product code: ALV002). |
|
| Wireless backpack stimulator | Custom | TI CC2431 micocontroler The board is custom made based on the GINA board from Prof. Kris Pister’s lab. The layout of GINA board can be found at https://openwsn.atlassian.net/wiki/display/OW/GINA |
|
| Wii Remote control | Nintendo | Bluetooth remote control to send the command to the operator laptop | |
| BeetleCommander v1.8 | Custom. Maharbiz group at UC Berkeley and Sato group at NTU | Establish the wireless communication of the backpack and the operator laptop. Configure the stimulus parameters and log the positional data. Visualize the flight data. | |
| GINA base station | Kris Pister group at UC Berkeley | TI MSP430F2618 and AT86RF231 | |
| Motion capture system | VICON | T160 | 8 cameras for a flight arena of 12.5 x 8 x 4 m |
| Motion capture system | VICON | T40s | 12 cameras for a flight arena of 12.5 x 8 x 4 m |
| Micro battery | Fullriver | 201013HS10C | 3.7V, 10 mAh |
| Retro reflective tape | Reflexite | V92-1549-010150 | V92 reflective tape, silver color |
| PFA-Insulated Silver Wire | A-M systems | 786000 | 127 µm bare, 177.8 µm coated, 3 mm bare silver flame exposed at tips |
| SMT Micro Header | SAMTEC | FTSH-110-01-L-DV | 0.3 x 6 mm, bend to make a 3 mm long slider to secure the electrode into the PCB header. |
| Beeswax | Secure the electrodes | ||
| Dental Wax | Vertex | Immobilize the beetle | |
| Insect pin | ROBOZ | RS-6082-30 | Size 00; 0.3mm Rod diameter; 0.03 mm tip width; 38 mm Length Make electrode guiding holes on cuticle |
| Tweezers | DUMONT | RS-5015 | Pattern #5; .05 X .01mm Tip Size; 110mm Length Dissecting and implantation |
| Scissors | ROBOZ | RS-5620 | Vannas Micro Dissecting Spring Scissors; Straight; 3mm Cutting Edge; 0.1mm Tip Width; 3" Overall Length Dissecting and implantation |
| Potable soldering iron | DAIYO | DS241 | Reflow beeswax |
| Hotplate | CORNING | PC-400D | Melting beeswax and dental wax |
| Flourescent lamp | Philips | TL5 14W | Light the entire flight arena with 30 panels (60 x 60 cm2). Each panel has 3 lamps. 14 W, 549 mm x 17 mm |
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