Optogenetic techniques are extremely useful for proving that the particular activity pattern of neuron(s) is the causal reason for a specific neural function. One of the major problems associated with optogenetics, however, is the requirement of a strong light for the activation. To analyze C. elegans behavior with optogenetics, most methods use a microscope with a strong light source to obtain enough light intensity and a computer-controlled motorized stage to keep the worm in the field of view. However, such methods limit the observation to one animal per assay, require multiple expensive equipments, and may not be suitable for experiments with delicate signal gradients such as odor or temperature. To overcome the limitations associated with such methods, we have established a simple and easy-to-use optogenetic system in combination with a strong LED ring and an improved channelrhodopsin (ChR). First, we developed a strong LED ring that allows exposure of an entire 9-cm plate at 0.5 mW/mm2 with 470 nm light; the intensity is comparable to the strongest light intensity obtained from a mercury lamp for GFP excitation. The LED can also be used to expose a small area of the plate at 1.2 mW/mm2. We have also developed a custom-made program with LabVIEW to control LED intensity in synchronization with capturing the images via a USB camera. Because of the high resolution of the camera (approximately 2.5 k x 2 k), the system does not need a microscope or a motorized stage, and we can record the behavior of multiple worms on a 9-cm plate simultaneously. Thus, using this system we are able to efficiently conduct optogenetic behavioral analysis for delicate signal gradients. In addition, this system is relatively inexpensive: the total cost, including the LED ring, camera, and PC is approximately $14,000. To prove that the LED can actually activate the optogenetic ion channel, we expressed an improved ChR, ChRGR (Wen et al., PLoS ONE, 2010), in the body wall muscles with a
myo-3 promoter. Upon light stimulation, the movement of transgenic animals was significantly inhibited in the presence of all-trans-retinal, indicating that the LED intensity was strong enough to activate ChRGR. We are currently trying to express ChRGR in the AWB sensory neurons to understand how the activity of AWB regulates avoidance behavior to repulsive odor 2-nonanone, which is enhanced by preexposure to the odor in a dopamine-dependent manner (Kimura et al., J. Neurosci., 2010).