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Optogenetic manipulation of neural circuits and behavior in Drosophila larvae

Abstract

Optogenetics is a powerful tool that enables the spatiotemporal control of neuronal activity and circuits in behaving animals. Here, we describe our protocol for optical activation of neurons in Drosophila larvae. As an example, we discuss the use of optogenetics to activate larval nociceptors and nociception behaviors in the third-larval instar. We have previously shown that, using spatially defined GAL4 drivers and potent UAS (upstream activation sequence)-channelrhodopsin-2YFP transgenic strains developed in our laboratory, it is possible to manipulate neuronal populations in response to illumination by blue light and to test whether the activation of defined neural circuits is sufficient to shape behaviors of interest. Although we have only used the protocol described here in larval stages, the procedure can be adapted to study neurons in adult flies—with the caveat that blue light may not sufficiently penetrate the adult cuticle to stimulate neurons deep in the brain. This procedure takes 1 week to culture optogenetic flies and 1 h per group for the behavioral assays.

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Figure 1: Workflow for the optogenetic activation of larval nociception.
Figure 2: Efficacy of behavioral response compared among UAS-ChR2 lines.
Figure 3: Setup of videorecording and illuminating devices.
Figure 4: Optogenetically triggered nociceptive responses in all-trans-retinal–fed ppk1.9-GAL4/UAS-ChR2YFP and control animals.

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Acknowledgements

This work was supported by grants from the US National Institutes of Neurological Disorders and Stroke 5R01NS054899 and 5R01GM086458 (W.D.T.). K.H. is a fellow supported by the Japan Society for the Promotion of Science postdoctoral fellowship for research abroad.

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Authors

Contributions

W.D.T. conceived of and supervised the project. R.Y.H. performed experiments and generated original data. K.H., R.Y.H. and W.D.T. designed the standardized protocol. K.H. and W.D.T. wrote the manuscript.

Corresponding author

Correspondence to William Daniel Tracey Jr.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Video 1

The movie shows a representative example of a ppk1.9-GAL4/UAS-channelrhodopsin-2::YFP larva executing the stereotyped escape locomotion in response to blue light stimulations. The movie was originally published in Hwang et al. (2007)22 (AVI 9407 kb)

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Honjo, K., Hwang, R. & Tracey, W. Optogenetic manipulation of neural circuits and behavior in Drosophila larvae. Nat Protoc 7, 1470–1478 (2012). https://doi.org/10.1038/nprot.2012.079

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