Addition of human melanopsin renders mammalian cells photoresponsive

Abstract

A small number of mammalian retinal ganglion cells act as photoreceptors for regulating certain non-image forming photoresponses1,2,3,4,5,6,7,8,9,10. These intrinsically photosensitive retinal ganglion cells express the putative photopigment melanopsin11,12,13. Ablation of the melanopsin gene renders these cells insensitive to light14; however, the precise role of melanopsin in supporting cellular photosensitivity is unconfirmed. Here we show that heterologous expression of human melanopsin in a mouse paraneuronal cell line (Neuro-2a) is sufficient to render these cells photoreceptive. Under such conditions, melanopsin acts as a sensory photopigment, coupled to a native ion channel via a G-protein signalling cascade, to drive physiological light detection. The melanopsin photoresponse relies on the presence of cis-isoforms of retinaldehyde and is selectively sensitive to short-wavelength light. We also present evidence to show that melanopsin functions as a bistable pigment in this system, having an intrinsic photoisomerase regeneration function that is chromatically shifted to longer wavelengths.

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Figure 1: Expression of human melanopsin renders Neuro-2a cells light-sensitive.
Figure 2: Melanopsin uses cis-isoforms of retinaldehyde.
Figure 3: Spectral sensitivity.
Figure 4: The phototransduction cascade in melanopsin-expressing cells loaded with 9-cis-retinaldehyde.

References

  1. 1

    Berson, D. M., Dunn, F. A. & Takao, M. Phototransduction by retinal ganglion cells that set the circadian clock. Science 295, 1070–1073 (2002)

    ADS  CAS  Article  Google Scholar 

  2. 2

    Hattar, S. et al. Melanopsin and rod–cone photoreceptive systems account for all major accessory visual functions in mice. Nature 424, 75–81 (2003)

    ADS  Article  Google Scholar 

  3. 3

    Panda, S. et al. Melanopsin is required for non-image-forming photic responses in blind mice. Science 301, 525–527 (2003)

    ADS  CAS  Article  Google Scholar 

  4. 4

    Sekaran, S., Foster, R. G., Lucas, R. J. & Hankins, M. W. Calcium imaging reveals a network of intrinsically light-sensitive inner-retinal neurons. Curr. Biol. 13, 1290–1298 (2003)

    CAS  Article  Google Scholar 

  5. 5

    Warren, E. J., Allen, C. N., Brown, R. L. & Robinson, D. W. Intrinsic light responses of retinal ganglion cells projecting to the circadian system. Eur. J. Neurosci. 17, 1727–1735 (2003)

    Article  Google Scholar 

  6. 6

    Lucas, R., Douglas, R. & Foster, R. Characterization of an ocular photopigment capable of driving pupillary constriction in mice. Nature Neurosci. 4, 621–626 (2001)

    CAS  Article  Google Scholar 

  7. 7

    Lucas, R. J. et al. Identifying the photoreceptive inputs to the mammalian circadian system using transgenic and retinally degenerate mice. Behav. Brain Res. 125, 97–102 (2001)

    CAS  Article  Google Scholar 

  8. 8

    Czeisler, C. A. et al. Suppression of melatonin secretion in some blind patients by exposure to bright light. N. Engl. J. Med. 332, 6–11 (1995)

    CAS  Article  Google Scholar 

  9. 9

    Mrosovsky, N., Lucas, R. & Foster, R. Persistence of masking responses to light in mice lacking rods and cones. J. Biol. Rhythms 16, 585–587 (2001)

    CAS  Article  Google Scholar 

  10. 10

    Lockley, S. W. et al. Relationship between melatonin rhythms and visual loss in the blind. J. Clin. Endocrinol. Metab. 82, 3763–3770 (1997)

    CAS  PubMed  Google Scholar 

  11. 11

    Hattar, S., Liao, H. W., Takao, M., Berson, D. M. & Yau, K. W. Melanopsin-containing retinal ganglion cells: architecture, projections, and intrinsic photosensitivity. Science 295, 1065–1070 (2002)

    ADS  CAS  Article  Google Scholar 

  12. 12

    Gooley, J. J., Lu, J., Chou, T. C., Scammell, T. E. & Saper, C. B. Melanopsin in cells of origin of the retinohypothalamic tract. Nature Neurosci. 4, 1165 (2001)

    CAS  Article  Google Scholar 

  13. 13

    Hannibal, J., Hindersson, P., Knudsen, S. M., Georg, B. & Fahrenkrug, J. The photopigment melanopsin is exclusively present in pituitary adenylate cyclase-activating polypeptide-containing retinal ganglion cells of the retinohypothalamic tract. J. Neurosci. 22, RC191 (2002)

    Article  Google Scholar 

  14. 14

    Lucas, R. J. et al. Diminished pupillary light reflex at high irradiances in melanopsin-knockout mice. Science 299, 245–247 (2003)

    ADS  CAS  Article  Google Scholar 

  15. 15

    Provencio, I., Jiang, G., De Grip, W. J., Hayes, W. P. & Rollag, M. D. Melanopsin: An opsin in melanophores, brain, and eye. Proc. Natl Acad. Sci. USA 95, 340–345 (1998)

    ADS  CAS  Article  Google Scholar 

  16. 16

    Hubbard, R. & St. George, R. The rhodopsin system of the squid. J. Gen. Physiol. 41, 501–528 (1958)

    CAS  Article  Google Scholar 

  17. 17

    Liu, R. S., Crescitelli, F., Denny, M., Matsumoto, H. & Asato, A. E. Photosensitivity of 10-substituted visual pigment analogues: detection of a specific secondary opsin-retinal interaction. Biochemistry 25, 7026–7030 (1986)

    CAS  Article  Google Scholar 

  18. 18

    Hao, W. & Fong, H. The endogenous chromophore of retinal G protein-coupled receptor opsin from the pigment epithelium. J. Biol. Chem. 274, 6085–6090 (1999)

    CAS  Article  Google Scholar 

  19. 19

    Koyanagi, M. et al. Bistable UV pigment in the lamprey pineal. Proc. Natl Acad. Sci. USA 101, 6687–6691 (2004)

    ADS  CAS  Article  Google Scholar 

  20. 20

    Brainard, G. et al. Action spectrum for melatonin regulation in humans: evidence for a novel circadian photoreceptor. J. Neurosci. 21, 6405–6412 (2001)

    CAS  Article  Google Scholar 

  21. 21

    Thapan, K., Arendt, J. & Skene, D. An action spectrum for melatonin suppression: evidence for a novel non-rod, non-cone photoreceptor system in humans. J. Physiol. (Lond.) 535, 261–267 (2001)

    CAS  Article  Google Scholar 

  22. 22

    Hankins, M. W. & Lucas, R. J. The primary visual pathway in humans is regulated according to long-term light exposure through the action of a nonclassical photopigment. Curr. Biol. 12, 191–198 (2002)

    CAS  Article  Google Scholar 

  23. 23

    Newman, L. A., Walker, M. T., Brown, R. L., Cronin, T. W. & Robinson, P. R. Melanopsin forms a functional short-wavelength photopigment. Biochemistry 42, 12734–12738 (2003)

    CAS  Article  Google Scholar 

  24. 24

    Provencio, I. et al. A novel human opsin in the inner retina. J. Neurosci. 20, 600–605 (2000)

    CAS  Article  Google Scholar 

  25. 25

    Bellingham, J., Whitmore, D., Philp, A. R., Wells, D. J. & Foster, R. G. Zebrafish melanopsin: isolation, tissue localisation and phylogenetic position. Brain Res. Mol. Brain Res. 107, 128–136 (2002)

    CAS  Article  Google Scholar 

  26. 26

    Beindl, W. et al. Inhibition of receptor/G protein coupling by suramin analogues. Mol. Pharmacol. 50, 415–423 (1996)

    CAS  PubMed  Google Scholar 

  27. 27

    Gilman, A. G proteins and dual control of adenylate cyclase. Cell 36, 577–579 (1984)

    CAS  Article  Google Scholar 

  28. 28

    Asano, T. & Ogasawara, N. Uncoupling of gamma-aminobutyric acid B receptors from GTP-binding proteins by N-ethylmaleimide: effect of N-ethylmaleimide on purified GTP-binding proteins. Mol. Pharmacol. 29, 244–249 (1986)

    CAS  PubMed  Google Scholar 

  29. 29

    Zemelman, B. V., Lee, G. A., Ng, M. & Miesenbock, G. Selective photostimulation of genetically chARGed neurons. Neuron 33, 15–22 (2002)

    CAS  Article  Google Scholar 

  30. 30

    Leszkiewicz, D., Kandler, K. & Aizenman, E. Enhancement of NMDA receptor-mediated currents by light in rat neurones in vitro . J. Physiol. (Lond.) 524, 365–374 (2000)

    CAS  Article  Google Scholar 

  31. 31

    Qiu, X. et al. Induction of photosensitivity by heterologous expression of melanopsin. Nature doi:10.1038/nature03345 (this issue)

  32. 32

    Panda, S. et al. Illumination of the melanopsin signaling pathway. Science 307, 600–604 (2005)

    ADS  CAS  Article  Google Scholar 

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Acknowledgements

This work was supported by a Wellcome Trust Showcase Award (M.W.H. and R.J.L.) and in part by the BBSRC (R.J.L.) and NSBRI through NASA NCC 9-58 (R.J.L. and R. Foster). We are grateful to R.K. Crouch for the gift of 11-cis retinaldehyde, to R. Douglas for scientific discussions and comments on the manuscript, and to K. Wells for help and advice with cell culture.

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Correspondence to R. J. Lucas or M. W. Hankins.

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Supplementary information

Supplementary Figure 1

Opsin gene expression in Neuro-2a cells by RT-PCR. (PDF 260 kb)

Supplementary Figure 2

Detection of melanopsin and EGFP proteins in Neuro-2a cells by western blot. (PDF 596 kb)

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Melyan, Z., Tarttelin, E., Bellingham, J. et al. Addition of human melanopsin renders mammalian cells photoresponsive. Nature 433, 741–745 (2005). https://doi.org/10.1038/nature03344

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