Skip to main content

Thank you for visiting You are using a browser version with limited support for CSS. To obtain the best experience, we recommend you use a more up to date browser (or turn off compatibility mode in Internet Explorer). In the meantime, to ensure continued support, we are displaying the site without styles and JavaScript.

Two-photon optogenetics of dendritic spines and neural circuits


We demonstrate a two-photon optogenetic method that generates action potentials in neurons with single-cell precision, using the red-shifted opsin C1V1T. We applied the method to optically map synaptic circuits in mouse neocortical brain slices and to activate small dendritic regions and individual spines. Using a spatial light modulator, we split the laser beam onto several neurons and performed simultaneous optogenetic activation of selected neurons in three dimensions.

This is a preview of subscription content, access via your institution

Relevant articles

Open Access articles citing this article.

Access options

Rent or buy this article

Get just this article for as long as you need it


Prices may be subject to local taxes which are calculated during checkout

Figure 1: Two-photon activation of individual neurons with C1V1T in mouse brain slices.
Figure 2: Two-photon stimulation of individual dendrites and spines and optical mapping of connected neurons.
Figure 3: Two-photon 3D stimulation of two individual neurons with SLMs.


  1. Yizhar, O., Fenno, L.E., Davidson, T.J., Mogri, M. & Deisseroth, K. Neuron 71, 9–34 (2011).

    Article  CAS  Google Scholar 

  2. Kätzel, D., Zemelman, B.V., Buetfering, C., Wölfel, M. & Miesenbock, G. Nat. Neurosci. 14, 100–107 (2011).

    Article  Google Scholar 

  3. Nikolenko, V., Poskanzer, K.E. & Yuste, R. Nat. Methods 4, 943–950 (2007).

    Article  CAS  Google Scholar 

  4. Rickgauer, J.P. & Tank, D.W. Proc. Natl. Acad. Sci. USA 106, 15025–15030 (2009).

    Article  CAS  Google Scholar 

  5. Andrasfalvy, B.K., Zemelman, B.V., Tang, J. & Vaziri, A. Proc. Natl. Acad. Sci. USA 107, 11981–11986 (2010).

    Article  CAS  Google Scholar 

  6. Papagiakoumou, E. et al. Nat. Methods 7, 848–854 (2010).

    Article  CAS  Google Scholar 

  7. Oron, D., Papagiakoumou, E., Anselmi, F. & Emiliani, V. Prog. Brain Res. 196, 119–143 (2012).

    Article  CAS  Google Scholar 

  8. Feldbauer, K. et al. Proc. Natl. Acad. Sci. USA 106, 12317–12322 (2009).

    Article  CAS  Google Scholar 

  9. Zhu, P. et al. Front. Neural Circuits 3, 21 (2009).

    Article  Google Scholar 

  10. Mattis, J. et al. Nat. Methods 9, 159–172 (2012).

    Article  CAS  Google Scholar 

  11. Nikolenko, V. et al. Front. Neural Circuits 2, 5 (2008).

    Article  Google Scholar 

  12. Lutz, C. et al. Nat. Methods 5, 821–827 (2008).

    Article  CAS  Google Scholar 

  13. Lefort, S., Tomm, C., Floyd Sarria, J.-C. & Petersen, C.C.H. Neuron 61, 301–316 (2009).

    Article  CAS  Google Scholar 

  14. Schüz, A. & Palm, G. J. Comp. Neurol. 286, 442–455 (1989).

    Article  Google Scholar 

  15. Nikolenko, V., Peterka, D.S. & Yuste, R. J. Neural Eng. 7, 045001 (2010).

    Article  Google Scholar 

  16. Yang, S. et al. J. Neural Eng. 8, 046002 (2011).

    Article  Google Scholar 

  17. Maurer, C., Khan, S., Fassl, S., Bernet, S. & Ritsch-Marte, M. Opt. Express 18, 3023–3034 (2010).

    Article  Google Scholar 

  18. Daria, V.R., Stricker, C., Bowman, R., Redman, S. & Bachor, H.-A. Appl. Phys. Lett. 95, 093701 (2009).

    Article  Google Scholar 

  19. Piestun, R. & Shamir, J. Proc. IEEE 90, 222–224 (2002).

    Article  Google Scholar 

  20. Grewe, B.F., Voigt, F.F., van't Hoff, M. & Helmchen, F. Biomed. Opt. Express 2, 2035–2046 (2011).

    Article  CAS  Google Scholar 

Download references


We thank M. Agetsuma and Y. Shin for assistance with viral injections and surgeries, and other members of the laboratory for help and comments. R.Y., A.M.P., J.J.H. and D.S.P. are supported by the HHMI, Kavli Institute for Brain Science, National Eye Institute, Keck Foundation, Deutsche Forschungsgemeinschaft (DFG grant HI 1728/1-1 to J.J.H.) and Department of Defense Multidisciplinary University Research Initiative Program. R.P. is supported by the US National Institute of Mental Health. K.D. is supported by the HHMI, US National Institutes of Health, California Institute for Regenerative Medicine, Gatsby Foundation and Defense Advanced Research Projects Agency Reorganization and Plasticity to Accelerate Injury Recovery Program.

Author information

Authors and Affiliations



A.M.P., D.S.P. and R.Y. designed and built the microscope software and hardware. A.M.P., D.S.P. and J.J.H. performed experiments, data analysis and quantification. R.P. and K.D. provided viral constructs, technical assistance, advice and opsin characterization. A.M.P., D.S.P., J.J.H. and R.Y. contributed to the writing of the manuscript.

Corresponding author

Correspondence to Rafael Yuste.

Ethics declarations

Competing interests

The authors declare no competing financial interests.

Supplementary information

Supplementary Text and Figures

Supplementary Figures 1–10 and Supplementary Table 1 (PDF 4963 kb)

Rights and permissions

Reprints and Permissions

About this article

Cite this article

Packer, A., Peterka, D., Hirtz, J. et al. Two-photon optogenetics of dendritic spines and neural circuits. Nat Methods 9, 1202–1205 (2012).

Download citation

  • Received:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI:

This article is cited by


Quick links

Nature Briefing

Sign up for the Nature Briefing newsletter — what matters in science, free to your inbox daily.

Get the most important science stories of the day, free in your inbox. Sign up for Nature Briefing