Abstract

Optogenetics is a powerful research tool because it enables high-resolution optical control of neuronal activity. However, current optogenetic approaches are limited to transgenic systems expressing microbial opsins and other exogenous photoreceptors. Here, we identify optovin, a small molecule that enables repeated photoactivation of motor behaviors in wild-type zebrafish and mice. To our surprise, optovin's behavioral effects are not visually mediated. Rather, photodetection is performed by sensory neurons expressing the cation channel TRPA1. TRPA1 is both necessary and sufficient for the optovin response. Optovin activates human TRPA1 via structure-dependent photochemical reactions with redox-sensitive cysteine residues. In animals with severed spinal cords, optovin treatment enables control of motor activity in the paralyzed extremities by localized illumination. These studies identify a light-based strategy for controlling endogenous TRPA1 receptors in vivo, with potential clinical and research applications in nontransgenic animals, including humans.

Access optionsAccess options

Rent or Buy article

Get time limited or full article access on ReadCube.

from$8.99

All prices are NET prices.

References

  1. 1.

    et al. Remote control of neuronal activity in transgenic mice expressing evolved G protein–coupled receptors. Neuron 63, 27–39 (2009).

  2. 2.

    , , , & Evolving the lock to fit the key to create a family of G protein–coupled receptors potently activated by an inert ligand. Proc. Natl. Acad. Sci. USA 104, 5163–5168 (2007).

  3. 3.

    , , , & Light-activated ion channels for remote control of neuronal firing. Nat. Neurosci. 7, 1381–1386 (2004).

  4. 4.

    , , , & Millisecond-timescale, genetically targeted optical control of neural activity. Nat. Neurosci. 8, 1263–1268 (2005).

  5. 5.

    Optogenetics. Nat. Methods 8, 26–29 (2011).

  6. 6.

    et al. Transient neuronal inhibition reveals opposing roles of indirect and direct pathways in sensitization. Nat. Neurosci. 14, 22–24 (2011).

  7. 7.

    et al. Remote control of neuronal activity with a light-gated glutamate receptor. Neuron 54, 535–545 (2007).

  8. 8.

    , , , & A light-gated, potassium-selective glutamate receptor for the optical inhibition of neuronal firing. Nat. Neurosci. 13, 1027–1032 (2010).

  9. 9.

    et al. Reversibly caged glutamate: a photochromic agonist of ionotropic glutamate receptors. J. Am. Chem. Soc. 129, 260–261 (2007).

  10. 10.

    et al. All optical interface for parallel, remote, and spatiotemporal control of neuronal activity. Nano Lett. 7, 3859–3863 (2007).

  11. 11.

    et al. TRPA1 contributes to cold, mechanical, and chemical nociception but is not essential for hair-cell transduction. Neuron 50, 277–289 (2006).

  12. 12.

    , & New photochemical tools for controlling neuronal activity. Curr. Opin. Neurobiol. 19, 544–552 (2009).

  13. 13.

    & Photostimulation using caged glutamate reveals functional circuitry in living brain slices. Proc. Natl. Acad. Sci. USA 90, 7661–7665 (1993).

  14. 14.

    & Rearrangements of synaptic connections in visual cortex revealed by laser photostimulation. Science 265, 255–258 (1994).

  15. 15.

    et al. Photochemical control of endogenous ion channels and cellular excitability. Nat. Methods 5, 331–338 10.1038/nmeth.1187 (2008).

  16. 16.

    et al. Optogenetic photochemical control of designer K+ channels in mammalian neurons. J. Neurophysiol. 106, 488–496 (2011).

  17. 17.

    et al. In vivo two-photon uncaging of glutamate revealing the structure-function relationships of dendritic spines in the neocortex of adult mice. J. Physiol. (Lond.) 589, 2447–2457 (2011).

  18. 18.

    et al. Allosteric control of an ionotropic glutamate receptor with an optical switch. Nat. Chem. Biol. 2, 47–52 (2006).

  19. 19.

    et al. Photolabile precursors of glutamate: synthesis, photochemical properties, and activation of glutamate receptors on a microsecond time scale. Proc. Natl. Acad. Sci. USA 91, 8752–8756 (1994).

  20. 20.

    et al. Rapid optical control of nociception with an ion-channel photoswitch. Nat. Methods 9, 396–402 (2012).

  21. 21.

    TRPV1 and TRPA1 channels in inflammatory pain: elucidating mechanisms. Ann. NY Acad. Sci. 1245, 36–37 (2011).

  22. 22.

    & TRP channels in disease. Subcell. Biochem. 45, 253–271 (2007).

  23. 23.

    et al. A gain-of-function mutation in TRPA1 causes familial episodic pain syndrome. Neuron 66, 671–680 (2010).

  24. 24.

    S. et al. Synergistic role of TRPV1 and TRPA1 in pancreatic pain and inflammation. Gastroenterology 140, 1283–1291 (2011).

  25. 25.

    & Zebrafish-based small molecule discovery. Chem. Biol. 10, 901–908 (2003).

  26. 26.

    & In vivo drug discovery in the zebrafish. Nat. Rev. Drug Discov. 4, 35–44 (2005).

  27. 27.

    et al. Rapid behavior-based identification of neuroactive small molecules in the zebrafish. Nat. Chem. Biol. 6, 231–237 (2010).

  28. 28.

    & Early retinal development in the zebrafish, Danio rerio: light and electron microscopic analyses. J. Comp. Neurol. 404, 515–536 (1999).

  29. 29.

    et al. Zebrafish behavioral profiling links drugs to biological targets and rest/wake regulation. Science 327, 348–351 (2010).

  30. 30.

    , & TRP ion channels and temperature sensation. Annu. Rev. Neurosci. 29, 135–161 (2006).

  31. 31.

    et al. Noxious cold ion channel TRPA1 is activated by pungent compounds and bradykinin. Neuron 41, 849–857 (2004).

  32. 32.

    , & TRP ion channels in the nervous system. Curr. Opin. Neurobiol. 14, 362–369 (2004).

  33. 33.

    , , , & TRPA1 mediates the noxious effects of natural sesquiterpene deterrents. J. Biol. Chem. 283, 24136–24144 (2008).

  34. 34.

    et al. Zebrafish TRPA1 channels are required for chemosensation but not for thermosensation or mechanosensory hair cell function. J. Neurosci. 28, 10102–10110 (2008).

  35. 35.

    et al. HC-030031, a TRPA1 selective antagonist, attenuates inflammatory- and neuropathy-induced mechanical hypersensitivity. Mol. Pain 4, 48 (2008).

  36. 36.

    Molecular mechanisms of photosensitization. Biochimie 68, 771–778 (1986).

  37. 37.

    & Quenching of singlet oxygen by tertiary aliphatic amines. Effect of DABCO (1,4-diazabicyclo [2.2.2]octane). J. Am. Chem. Soc. 90, 6527–6528 (1968).

  38. 38.

    et al. Noxious compounds activate TRPA1 ion channels through covalent modification of cysteines. Nature 445, 541–545 (2007).

  39. 39.

    et al. TRPA1 underlies a sensing mechanism for O2. Nat. Chem. Biol. 7, 701–711 (2011).

  40. 40.

    , , & TRP channel activation by reversible covalent modification. Proc. Natl. Acad. Sci. USA 103, 19564–19568 (2006).

  41. 41.

    & Photoinduced stereospecific formation of substituted hydantoin from hexobarbital. J. Photochem. Photobiol. A Chem. 54, 187–196 (1990).

  42. 42.

    et al. Rhodanine dyes for dye-sensitized solar cells: spectroscopy, energy levels and photovoltaic performance. Phys. Chem. Chem. Phys. 11, 133–141 (2009).

  43. 43.

    et al. Identification of diaryl ether–based ligands for estrogen-related receptor alpha as potential antidiabetic agents. J. Med. Chem. 54, 788–808 (2011).

  44. 44.

    et al. Reversible targeting of noncatalytic cysteines with chemically tuned electrophiles. Nat. Chem. Biol. 8, 471–476 (2012).

  45. 45.

    & Function, structure and mechanism of bacterial photosensory LOV proteins. Nat. Rev. Microbiol. 9, 713–723 (2011).

  46. 46.

    et al. Protease-stable polycationic photosensitizer conjugates between polyethyleneimine and chlorin(e6) for broad-spectrum antimicrobial photoinactivation. Antimicrob. Agents Chemother. 50, 1402–1410 (2006).

  47. 47.

    , & A method for assessing the effects of drugs on the central actions of 5-hydroxytryptamine. Br. J. Pharmacol. Chemother. 20, 106–120 (1963).

Download references

Acknowledgements

We thank K. Kwan and D. Corey (Harvard Medical School), T. Miyamoto and A. Patapoutian (Scripps Research Institute), and T. Numata and Y. Mori (Kyoto University) for human TrpA1 constructs. A. Schier, D. Prober and D. Robson (Harvard University) generously provided TrpA1 mutant zebrafish. We thank R. Gaudet, A. Vakkasglu, B. Shoichet, T. Dunn, M. Ahrens, F. Engert, R. Mazitschek and members of our research groups for helpful advice. This work was supported by US National Institutes of Health (NIH) grants K01MH091449 (D.K.), MH086867 and MH085205 (R.T.P.), P01 NS072040 (C.J.W.), R01 AI050875 (L.H. and M.R.H.); the Charles and Ann Sanders Massachusetts General Hospital Research Scholar award (R.T.P.); and the Michael Hooker Chair and the NIMH PDSP (B.L.R.). M.J.Z. was supported by grants from National Institute of Neurological Disorders and Stroke (NINDS; R01NS060725, R01NS067688), J.C.-B. was supported by a National Research Service Award training grant from NINDS (F31NS068038) and D.J.M. was supported by NIH grants HL109004 and DA026982.

Author information

Affiliations

  1. Department of Medicine, Cardiovascular Research Center and Division of Cardiology, Massachusetts General Hospital, Harvard Medical School, Charlestown, Massachusetts, USA.

    • David Kokel
    • , Chung Yan J Cheung
    • , Robert Mills
    • , Shan Jin
    • , Youngnam N Jin
    • , Giancarlo Bruni
    • , David J Milan
    •  & Randall T Peterson
  2. Broad Institute, Cambridge, Massachusetts, USA.

    • David Kokel
    • , Chung Yan J Cheung
    • , Shan Jin
    • , Youngnam N Jin
    • , Giancarlo Bruni
    •  & Randall T Peterson
  3. University of North Carolina at Chapel Hill Neuroscience Center, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.

    • Jaeda Coutinho-Budd
    •  & Mark J Zylka
  4. Department of Cell and Molecular Physiology, University of North Carolina at Chapel Hill, Chapel Hill, North Carolina, USA.

    • Mark J Zylka
  5. Wellman Center for Photomedicine, Massachusetts General Hospital, Boston, Massachusetts, USA.

    • Liyi Huang
    •  & Michael R Hamblin
  6. Department of Dermatology, Harvard Medical School, Boston, Massachusetts, USA.

    • Liyi Huang
    •  & Michael R Hamblin
  7. Department of Infectious Diseases, First Affiliated College and Hospital, Guangxi Medical University, Nanning, China.

    • Liyi Huang
  8. Department of Pharmacology and National Institute of Mental Health Psychoactive Drug Screening Program, University of North Carolina Chapel Hill Medical School, Chapel Hill, North Carolina.

    • Vincent Setola
    • , Xi-Ping Huang
    •  & Bryan L Roth
  9. Neurobiology Department, Children's Hospital Boston, Harvard Medical School, Boston, Massachusetts, USA.

    • Jared Sprague
    •  & Clifford J Woolf
  10. Harvard School of Dental Medicine, Boston, Massachusetts, USA.

    • Jared Sprague
  11. Harvard-Massachusetts Institute of Technology Division of Health Sciences and Technology, Cambridge, Massachusetts, USA.

    • Michael R Hamblin

Authors

  1. Search for David Kokel in:

  2. Search for Chung Yan J Cheung in:

  3. Search for Robert Mills in:

  4. Search for Jaeda Coutinho-Budd in:

  5. Search for Liyi Huang in:

  6. Search for Vincent Setola in:

  7. Search for Jared Sprague in:

  8. Search for Shan Jin in:

  9. Search for Youngnam N Jin in:

  10. Search for Xi-Ping Huang in:

  11. Search for Giancarlo Bruni in:

  12. Search for Clifford J Woolf in:

  13. Search for Bryan L Roth in:

  14. Search for Michael R Hamblin in:

  15. Search for Mark J Zylka in:

  16. Search for David J Milan in:

  17. Search for Randall T Peterson in:

Contributions

D.K. designed and performed the research, analyzed the data and wrote the manuscript with R.T.P. C.Y.J.C., R.M., J.C.-B., L.H., V.S., J.S., S.J., Y.N.J., G.B. and X.-P.H. designed and performed the experiments and interpreted data. C.J.W., B.L.R., M.R.H., M.J.Z. and D.J.M. analyzed and interpreted the data. All authors contributed to data interpretation and commented on the manuscript.

Competing interests

The authors declare no competing financial interests.

Corresponding authors

Correspondence to David Kokel or Randall T Peterson.

Supplementary information

PDF files

  1. 1.

    Supplementary Text and Figures

    Supplementary Results

Videos

  1. 1.

    Supplementary Movie 1

    Supplemenary Video 1

  2. 2.

    Supplementary Movie 2

    Supplemenary Video 2

  3. 3.

    Supplementary Movie 3

    Supplemenary Video 3

  4. 4.

    Supplementary Movie 4

    Supplemenary Video 4

  5. 5.

    Supplementary Movie 5

    Supplemenary Video 5

  6. 6.

    Supplementary Movie 6

    Supplemenary Video 6

About this article

Publication history

Received

Accepted

Published

DOI

https://doi.org/10.1038/nchembio.1183