C. elegans Tracking and Behavioral Measurement

Summary

We have developed a video-rate tracking microscope system that can record and quantify C. elegans behavior at high resolution and high speeds. We have also developed computational methods to reduce the dimensionality of the worm images to a fundamental set of measurements that completely describe the shape of the worm.

Abstract

We have developed instrumentation, image processing, and data analysis techniques to quantify the locomotory behavior of C. elegans as it crawls on the surface of an agar plate. For the study of the genetic, biochemical, and neuronal basis of behavior, C. elegans is an ideal organism because it is genetically tractable, amenable to microscopy, and shows a number of complex behaviors, including taxis, learning, and social interaction^1,2. Behavioral analysis based on tracking the movements of worms as they crawl on agar plates have been particularly useful in the study of sensory behavior³, locomotion⁴, and general mutational phenotyping⁵. Our system works by moving the camera and illumination system as the worms crawls on a stationary agar plate, which ensures no mechanical stimulus is transmitted to the worm. Our tracking system is easy to use and includes a semi-automatic calibration feature. A challenge of all video tracking systems is that it generates an enormous amount of data that is intrinsically high dimensional. Our image processing and data analysis programs deal with this challenge by reducing the worms shape into a set of independent components, which comprehensively reconstruct the worms behavior as a function of only 3-4 dimensions^6,7. As an example of the process we show that the worm enters and exits its reversal state in a phase specific manner.

Protocol

1. Description of Tracking Microscope

1. An agar plate is illuminated by a fiber light source and imaged with a camera. This system is mounted to an X,Y translation stage.
2. The stage is moved by standard stepper motors, which are connected to a stepper motor controller.
3. The controller and camera are connected to the computer and controlled by custom programs written in LabVIEW.
4. The camera images the surface of an agar plate and identifies dark objects on a light background.
5. The image quality is adjusted so that computer program can quantify objects in real time. The gain, brightness, and shutter speed of the ca....

Results

Example: When foraging, C. elegans transitions from forward to reverse motion, often performing a reorientation (omega turn) before returning to the forward motion state. Quantifying this transition is important in understanding the foraging patterns of movement and also in the worm's motor control. The power to reveal subtle details of locomotion behavior can be seen using our tracker device.

As an example we look at the forward to reverse and reverse to forward transition by captur.......

Discussion

The study of locomotion and natural behavior requires non-invasive tracking techniques in partner with data reduction techniques. Here we have demonstrated an easy to use tracking system that records detailed images of C. elegans behavior as it crawls on the surface of an agar plate. The amount of information contained in these images is vast and high-dimensional, and so we have also developed methods to reduce the dimensionality of the data into only four fundamental measures. These measures are comprehensive a.......

Materials

List of materials used in this article

Name	Company	Catalog Number	Comments
CCD camera	Basler	A601f
Lens	Edmund Optics	MMS series
Fiber Illumination	Dolan Jenner	DC-950H
Translation stage	Deltron	LS3-4
Stepper Motor	US digital	MS23C
Stepper motor drive	Gecko	G201
Stepper motor control	SimpleStep	SSXYZ
All programming code is available. Please send a request email to the corresponding author.