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Optogenetic inhibition of behavior with anion channelrhodopsins


Optogenetics uses light exposure to manipulate physiology in genetically modified organisms. Abundant tools for optogenetic excitation are available, but the limitations of current optogenetic inhibitors present an obstacle to demonstrating the necessity of neuronal circuits. Here we show that anion channelrhodopsins can be used to specifically and rapidly inhibit neural systems involved in Drosophila locomotion, wing expansion, memory retrieval and gustation, thus demonstrating their broad utility in the circuit analysis of behavior.

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Figure 1: G. theta anion channelrhodopsins are inhibitors of motor function and neuronal spiking.
Figure 2: Flies avoid optogenetic suppression of sweet-taste receptors.

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  1. Sweeney, S.T., Broadie, K., Keane, J., Niemann, H. & O'Kane, C.J. Neuron 14, 341–351 (1995).

    Article  CAS  Google Scholar 

  2. Kitamoto, T. J. Neurobiol. 47, 81–92 (2001).

    Article  CAS  Google Scholar 

  3. Zemelman, B.V., Lee, G.A., Ng, M. & Miesenböck, G. Neuron 33, 15–22 (2002).

    Article  CAS  Google Scholar 

  4. Boyden, E.S., Zhang, F., Bamberg, E., Nagel, G. & Deisseroth, K. Nat. Neurosci. 8, 1263–1268 (2005).

    Article  CAS  Google Scholar 

  5. Hamada, F.N. et al. Nature 454, 217–220 (2008).

    Article  CAS  Google Scholar 

  6. Tye, K.M. & Deisseroth, K. Nat. Rev. Neurosci. 13, 251–266 (2012).

    Article  CAS  Google Scholar 

  7. Lima, S.Q. & Miesenböck, G. Cell 121, 141–152 (2005).

    Article  CAS  Google Scholar 

  8. Zhang, F. et al. Nature 446, 633–639 (2007).

    Article  CAS  Google Scholar 

  9. Chow, B.Y. et al. Nature 463, 98–102 (2010).

    Article  CAS  Google Scholar 

  10. Wietek, J. et al. Science 344, 409–412 (2014).

    Article  CAS  Google Scholar 

  11. Berndt, A., Lee, S.Y., Ramakrishnan, C. & Deisseroth, K. Science 344, 420–424 (2014).

    Article  CAS  Google Scholar 

  12. Govorunova, E.G., Sineshchekov, O.A., Janz, R., Liu, X. & Spudich, J.L. Science 349, 647–650 (2015).

    Article  CAS  Google Scholar 

  13. Thoma, V. et al. Nat. Commun. 7, 10678 (2016).

    Article  CAS  Google Scholar 

  14. Klapoetke, N.C. et al. Nat. Methods 11, 338–346 (2014).

    Article  CAS  Google Scholar 

  15. Aso, Y. et al. eLife 3, e04580 (2014).

    Article  Google Scholar 

  16. Wu, M.-C. et al. Proc. Natl. Acad. Sci. USA 111, 5367–5372 (2014).

    Article  CAS  Google Scholar 

  17. Inada, K., Kohsaka, H., Takasu, E., Matsunaga, T. & Nose, A. PLoS One 6, e29019 (2011).

    Article  CAS  Google Scholar 

  18. Mahn, M., Prigge, M., Ron, S., Levy, R. & Yizhar, O. Nat. Neurosci. 19, 554–556 (2016).

    Article  CAS  Google Scholar 

  19. Knoflach, F., Hernandez, M.-C. & Bertrand, D. Biochem. Pharmacol. 115, 10–17 (2016).

    Article  CAS  Google Scholar 

  20. Wiegert, J.S. & Oertner, T.G. Nat. Neurosci. 19, 527–528 (2016).

    Article  CAS  Google Scholar 

  21. Salvaterra, P.M. & Kitamoto, T. Brain Res. Gene Expr. Patterns 1, 73–82 (2001).

    Article  CAS  Google Scholar 

  22. Peabody, N.C. et al. J. Neurosci. 28, 14379–14391 (2008).

    Article  CAS  Google Scholar 

  23. Petersen, L.K. & Stowers, R.S. PLoS One 6, e24531 (2011).

    Article  CAS  Google Scholar 

  24. Weiss, L.A., Dahanukar, A., Kwon, J.Y., Banerjee, D. & Carlson, J.R. Neuron 69, 258–272 (2011).

    Article  CAS  Google Scholar 

  25. Connolly, J.B. et al. Science 274, 2104–2107 (1996).

    Article  CAS  Google Scholar 

  26. Freeman, M. Cell 87, 651–660 (1996).

    Article  CAS  Google Scholar 

  27. Choi, Y.-J., Lee, G. & Park, J.H. Development 133, 2223–2232 (2006).

    Article  CAS  Google Scholar 

  28. Pfeiffer, B.D. et al. Genetics 186, 735–755 (2010).

    Article  CAS  Google Scholar 

  29. Claridge-Chang, A. et al. Cell 139, 405–415 (2009).

    Article  CAS  Google Scholar 

  30. Vogt, K. et al. eLife 3, e02395 (2014).

    Article  Google Scholar 

  31. Quinn, W.G., Harris, W.A. & Benzer, S. Proc. Natl. Acad. Sci. USA 71, 708–712 (1974).

    Article  CAS  Google Scholar 

  32. Van Vactor, D.L. Jr., Cagan, R.L., Krämer, H. & Zipursky, S.L. Cell 67, 1145–1155 (1991).

    Article  Google Scholar 

  33. Park, D., Veenstra, J.A., Park, J.H., & Taghert, P.H. PLoS One 3, e1896 (2008).

  34. Parnas, D., Haghighi, A.P., Fetter, R.D., Kim, S.W. & Goodman, C.S. Neuron 32, 415–424 (2001).

  35. Verstreken, P. et al. Neuron 40, 733–748 (2003).

    Article  CAS  Google Scholar 

  36. Tracey, W.D. Jr., Wilson, R.I., Laurent, G. & Benzer, S. Cell 113, 261–273 (2003).

    Article  CAS  Google Scholar 

  37. Meliza, C.D. & Margoliash, D. J. Neurosci. 32, 15158–15168 (2012).

    Article  CAS  Google Scholar 

  38. Altman, D., Machin, D., Bryant, T. & Gardner, S. Statistics with Confidence: Confidence Interval and Statistical Guidelines (BMJ Books, 2000).

  39. Claridge-Chang, A. & Assam, P.N. Nat. Methods 13, 108–109 (2016).

    Article  CAS  Google Scholar 

  40. Cumming, G. Understanding the New Statistics Effect Sizes, Confidence Intervals, and Meta-analysis (Routledge, 2012).

  41. Gardner, M.J. & Altman, D.G. Br. Med. J. (Clin. Res. Ed.) 292, 746–750 (1986).

    Article  CAS  Google Scholar 

  42. Efron, B. Ann. Stat. 7, 1–26 (1979).

    Article  Google Scholar 

  43. DiCiccio, T.J. & Efron, B. Stat. Sci. 11, 189–212 (1996).

    Article  Google Scholar 

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We thank J. Spudich (University of Texas Health Science Center at Houston) for providing the GtACR1 and GtACR2 sequences and plasmids; J.A. Veenstra (Université de Bordeaux) for the anti-Crz antibody; G. Rubin (Howard Hughes Medical Institute) and J. Park (University of Tennessee, Knoxville) for providing materials; S. Aw (Institute of Molecular and Cell Biology) for loan of the high-speed camera; G. Augustine for reading the manuscript; L. Robinson (Insight Editing London) for editing of the manuscript; and S.Y.H. Tan for drawing the behavior rig diagrams. F.M., S.O., J.Y.C. and A.C.-C. were supported by grant MOE-2013-T2-2-054 from the Ministry of Education; J.C.S. and A.C.-C. were supported by grants 1231AFG030 and 1431AFG120 from the A*STAR Joint Council Office. J.H. was supported by the A*STAR Scientific Scholars Fund. T.-W.K. was supported by the National Research Foundation Fellowship NRF-NRFF2015-06 and a block grant from the Temasek Life Sciences Laboratory. The authors were supported by a Biomedical Research Council block grant to the Institute of Molecular and Cell Biology. F.M., S.O., K.C. and A.C.-C. received support from the Duke-NUS Medical School, including the Integrated Biology and Medicine doctoral program (to K.C.).

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Authors and Affiliations



Conceptualization, F.M. and A.C.-C.; methodology, F.M., J.C.S. and A.C.-C.; software, J.C.S. (CRITTA, LabView) and J.H. (Python); investigation, F.M. (transgenic design, genetics, falling, walking immobilization, wing expansion, eye toxicity, valence, PER and neuroanatomy), J.C.S. (falling and high-frame rate paralysis), S.O. (learning and survival), J.Y.C. (brain dissection, immunohistochemistry and microscopy), K.C. (PER) and T.-W.K. (electrophysiology); resources, J.C.S. (instrumentation); data analysis, J.H. (valence, electrophysiology), K.C. (PER), J.C.S. (paralysis, falling and walking immobilization), S.O. (STM) and F.M. (paralysis, valence and anatomy); writing original draft, F.M. and A.C.-C. with contributions from all authors; writing revision, F.M., S.O., J.C.S. and A.C.-C.; visualization, F.M., J.H., J.C.S., S.O., K.C. and A.C.-C.; supervision, A.C.-C.; project administration, A.C.-C.; funding acquisition, A.C.-C.

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Correspondence to Adam Claridge-Chang.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–10 (PDF 24927 kb)

GtACR flies fall from a vertical surface when illuminated

Flies expressing one of three optogenetic inhibitors in their cholinergic neurons (Cha-Gal4>UAS-GtACR1, Cha-Gal4>UAS-GtACR2 and ChaGal4>UAS-eNpHR) were illuminated with light from a projector. Cha>GtACR1 and Cha>GtACR2 flies fell from the vertical acrylic surface upon exposure to green or blue light respectively, and were immobilized. Cha>GtACR2 flies retained some motor activity while illuminated with blue light. Cha>eNpHR flies did not fall upon exposure to red light and remained mobile. (MP4 9806 kb)

GtACR flies are immobilized by illumination

A. Green light at 38 μW/mm2 rendered a Cha>GtACR1 fly immobile, though it regained some motor control during illumination. Green dot indicates when light was turned on. B. Illumination of a GtACR1/+ fly with 38 μW/mm2 green light had no effect. C. A Cha>GtACR2 fly was rendered completely paralyzed by illumination with 391 μW/mm2 blue light. Blue dot indicates when light was turned on. D. A GtACR2/+ fly was unaffected by illumination with 391 μW/mm2 blue light. E. While positioned 3 mm above an amber LED (approximately 1.9 mW/mm2), a Cha>eNpHR fly retained mobility, though it was paralyzed transiently when passing directly above the emitter. Light was on throughout this recording. F. A Cha>eNpHR fly was unaffected by amber illumination at 495 μW/mm2. Amber dot indicates when light was on. (MP4 25723 kb)

Supplementary Video 3

Cha>GtACR flies adopt a static pose during illumination (indicated by colored dots), but Cha>Chrimson flies have active seizures and adopt a tetanic pose with extended wings. Control animals were unaffected by projector light (green 92 μW/mm2; blue 67 μW/mm2; red 70 μW/mm2). (MP4 25469 kb)

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Mohammad, F., Stewart, J., Ott, S. et al. Optogenetic inhibition of behavior with anion channelrhodopsins. Nat Methods 14, 271–274 (2017).

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