Optogenetic manipulation of neural activity in freely moving Caenorhabditis elegans

We present an optogenetic illumination system capable of real-time light delivery with high spatial resolution to specified targets in freely moving Caenorhabditis elegans. A tracking microscope records the motion of an unrestrained worm expressing channelrhodopsin-2 or halorhodopsin in specific cell types. Image processing software analyzes the worm's position in each video frame, rapidly estimates the locations of targeted cells and instructs a digital micromirror device to illuminate targeted cells with laser light of the appropriate wavelengths to stimulate or inhibit activity. Because each cell in an unrestrained worm is a rapidly moving target, our system operates at high speed (∼50 frames per second) to provide high spatial resolution (∼30 μm). To test the accuracy, flexibility and utility of our system, we performed optogenetic analyses of the worm motor circuit, egg-laying circuit and mechanosensory circuits that have not been possible with previous methods.

[1]  S. Brenner,et al.  The structure of the ventral nerve cord of Caenorhabditis elegans. , 1976, Philosophical transactions of the Royal Society of London. Series B, Biological sciences.

[2]  S. Brenner,et al.  The neural circuit for touch sensitivity in Caenorhabditis elegans , 1985, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[3]  R. Edwards,et al.  Molecular cloning of a putative vesicular transporter for acetylcholine. , 1994, Proceedings of the National Academy of Sciences of the United States of America.

[4]  E. Marder,et al.  Principles of rhythmic motor pattern generation. , 1996, Physiological reviews.

[5]  S. R. Wicks,et al.  A Dynamic Network Simulation of the Nematode Tap Withdrawal Circuit: Predictions Concerning Synaptic Function Using Behavioral Criteria , 1996, The Journal of Neuroscience.

[6]  Contribution of neurons to habituation to mechanical stimulation in Caenorhabditis elegans. , 2001, Journal of neurobiology.

[7]  A. Miyawaki,et al.  An optical marker based on the UV-induced green-to-red photoconversion of a fluorescent protein , 2002, Proceedings of the National Academy of Sciences of the United States of America.

[8]  Caenorhabditis Elegans Martinchalfieandjohnsulston.  Developmental Genetics of the Mechanosensory Neurons of Caenorhabditis elegans , 2003 .

[9]  E. Bamberg,et al.  Channelrhodopsin-2, a directly light-gated cation-selective membrane channel , 2003, Proceedings of the National Academy of Sciences of the United States of America.

[10]  K. Deisseroth,et al.  Millisecond-timescale, genetically targeted optical control of neural activity , 2005, Nature Neuroscience.

[11]  E. Bamberg,et al.  Light Activation of Channelrhodopsin-2 in Excitable Cells of Caenorhabditis elegans Triggers Rapid Behavioral Responses , 2005, Current Biology.

[12]  Qiang Liu,et al.  Low Conductance Gap Junctions Mediate Specific Electrical Coupling in Body-wall Muscle Cells of Caenorhabditis elegans* , 2006, Journal of Biological Chemistry.

[13]  Stephen E Von Stetina,et al.  The motor circuit. , 2006, International review of neurobiology.

[14]  D. Chklovskii,et al.  Wiring optimization can relate neuronal structure and function. , 2006, Proceedings of the National Academy of Sciences of the United States of America.

[15]  Feng Zhang,et al.  Channelrhodopsin-2 and optical control of excitable cells , 2006, Nature Methods.

[16]  Paul W. Sternberg,et al.  Systems level circuit model of C. elegans undulatory locomotion: mathematical modeling and molecular genetics , 2007, Journal of Computational Neuroscience.

[17]  Herwig Baier,et al.  Remote Control of Neuronal Activity with a Light-Gated Glutamate Receptor , 2007, Neuron.

[18]  Feng Zhang,et al.  Nociceptive Neurons Protect Drosophila Larvae from Parasitoid Wasps , 2007, Current Biology.

[19]  R. K. Herman.  Mosaic analysis in the nematode Caenorhabditis elegans. , 2007, Journal of neurogenetics.

[20]  E. Boyden,et al.  Multiple-Color Optical Activation, Silencing, and Desynchronization of Neural Activity, with Single-Spike Temporal Resolution , 2007, PloS one.

[21]  Feng Zhang,et al.  Multimodal fast optical interrogation of neural circuitry , 2007, Nature.

[22]  H. Horvitz,et al.  FMRFamide neuropeptides and acetylcholine synergistically inhibit egg-laying by C. elegans , 2008, Nature Neuroscience.

[23]  F. Engert,et al.  Escape Behavior Elicited by Single, Channelrhodopsin-2-Evoked Spikes in Zebrafish Somatosensory Neurons , 2008, Current Biology.

[24]  M. Zhen,et al.  Optogenetic analysis of synaptic function , 2008, Nature Methods.

[25]  John Bryden,et al.  Neural control of Caenorhabditis elegans forward locomotion: the role of sensory feedback , 2008, Biological Cybernetics.

[26]  Ethan K. Scott,et al.  Optogenetic dissection of a behavioral module in the vertebrate spinal cord , 2009, Nature.

[27]  Stefan R. Pulver,et al.  Temporal dynamics of neuronal activation by Channelrhodopsin-2 and TRPA1 determine behavioral output in Drosophila larvae. , 2009, Journal of neurophysiology.

[28]  Sharad Ramanathan,et al.  Optical interrogation of neural circuits in Caenorhabditis elegans , 2009, Nature Methods.

[29]  S. Cooper,et al.  Remote Control , 2002, Nursing standard (Royal College of Nursing (Great Britain) : 1987).

[30]  Michael J. O'Donovan,et al.  Motoneurons Dedicated to Either Forward or Backward Locomotion in the Nematode Caenorhabditis elegans , 2010, The Journal of Neuroscience.

[31]  Jeffrey N. Stirman,et al.  High-throughput study of synaptic transmission at the neuromuscular junction enabled by optogenetics and microfluidics , 2010, Journal of Neuroscience Methods.

[32]  Michael A. Henninger,et al.  High-Performance Genetically Targetable Optical Neural Silencing via Light-Driven Proton Pumps , 2010 .