Insect-machine Hybrid System: Remote Radio Control of a Freely Flying Beetle (Mercynorrhina torquata)

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

The rise of radio-enabled digital electronic devices has prompted the use of small wireless neuromuscular recorders and stimulators for studying in-flight insect behavior. This technology enables the development of an insect-machine hybrid system using a living insect platform described in this protocol. Moreover, this protocol presents the system configuration and free flight experimental procedures for evaluating the function of the flight muscles in an untethered insect. For demonstration, we targeted the third axillary sclerite (3Ax) muscle to control and achieve left or right turning of a flying beetle. A thin silver wire electrode was implanted on the 3Ax muscle on each side of the beetle. These were connected to the outputs of a wireless backpack (i.e., a neuromuscular electrical stimulator) mounted on the pronotum of the beetle. The muscle was stimulated in free flight by alternating the stimulation side (left or right) or varying the stimulation frequency. The beetle turned to the ipsilateral side when the muscle was stimulated and exhibited a graded response to an increasing frequency. The implantation process and volume calibration of the 3 dimensional motion capture camera system need to be carried out with care to avoid damaging the muscle and losing track of the marker, respectively. This method is highly beneficial to study insect flight, as it helps to reveal the functions of the flight muscle of interest in free flight.

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 components^1-3. The development of system-on-a-chip technolo....

Protocol

1. Study Animal

1. Rear individual Mecynorrhina torquata beetles (6 cm, 8 g) in separate plastic containers with wood pellet bedding.
2. Feed each beetle a cup of sugar jelly (12 ml) every 3 days.
3. Keep the temperature and humidity of the rearing room at 25 °C and 60%, respectively.
4. Test the flight capability of each beetle before implanting thin wire electrodes.
   1. Gently throw a beetle into the air. If the beetle can fly for longer than 10 sec for 5 consecutive trials, conclude that the beetle has regular flight capabilities and employ it for subsequent flight experiments. To recapture the beetle, switch off all the l....

Results

The electrode implantation procedure is presented in Figure 2. Thin silver wire electrodes were implanted into the 3Ax muscle of the beetle through small holes pierced on the soft cuticle on the muscle (Figures 2d-e). This soft cuticle is found just above the apodema of the basalar muscle after removing the anterior part of the metepisternum (Figures 2d-c). The electrodes were then secured using beeswax (.......

Discussion

The implantation process is important, as it affects the reliability of the experiment. The electrodes should be inserted into the muscle at a depth of 3 mm or less depending on the size of the beetle (avoiding contact with nearby muscles). If the electrodes touch the nearby muscles, undesirable motor actions and behaviors may occur owing to the contraction of nearby muscles. The two electrodes should be well aligned to ensure that no short circuits occur. When melting and reflowing beeswax using a soldering iron, the ex.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
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 m x 8 m 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 mm 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; 0.3 mm Rod diameter; 0.03 mm tip width; 38 mm Length  Make electrode guiding holes on cuticle
Tweezers	DUMONT	RS-5015	Pattern #5; .05 mm x .01 mm Tip Size; 110 mm Length Dissecting and implantation
Scissors	ROBOZ	RS-5620	Vannas Micro Dissecting Spring Scissors; Straight; 3mm Cutting Edge; 0.1 mm 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