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BioNanoScience

, Volume 6, Issue 4, pp 329–331 | Cite as

Expression of Channelrhodopsin-2 Using in Suspension Electroporation for Studying the Monosynaptic Transmission in Neuronal Culture

  • Natalia Bal
  • Aleksey Malyshev
  • Ivan Smirnov
  • Pavel Balaban
Article
  • 94 Downloads

Abstract

Understanding the mechanism of synaptic transmission and its plasticity is one of the central goals of modern neuroscience. Neuronal culture is a very convenient model for studying mechanisms of synaptic transmission. However, investigation of two or more synaptically connected neurons in culture by conventional methods is a difficult task. In this study, we describe new protocol for studying the synaptic transmission between cultured neurons using in suspension electroporation for the expression of channelrhodopsin-2 (ChR2) which was applied to only certain subpopulation of cultured cells, leaving the majority of neurons completely untreated. We show that this technique allows reliable long-lasting (hours) recording of monosynaptic excitatory postsynaptic potentials (EPSPs) in cultured hippocampal neurons using a repeated light stimulation of neighboring ChR2-expressing neurons.

Keywords

Electroporation Neuronal culture Channelrhodopsin Optogenetics Synaptic transmission 

Notes

Acknowledgments

This research was funded by the Russian Science Foundation grant 14-25-00072 (neuronal cell culture and development of the electroporation protocol) and by RFBR grant no. 15-04-06286 (whole cell recording with optical stimulation).

Compliance with Ethical Standards

All experimental procedures were conducted in accordance with the European Communities Council Directive of 24 November 1986 (86/609/ EEC) on the protection of animals used for scientific purposes. The study protocol was approved by the Ethics Committee of the Institute of Higher Nervous Activity and Neurophysiology of RAS.

References

  1. 1.
    Molnár, E. (2011). Long-term potentiation in cultured hippocampal neurons. Seminars in Cell & Developmental Biology, 22, 506–513. doi: 10.1016/j.semcdb.2011.07.017.CrossRefGoogle Scholar
  2. 2.
    Sombati, S., & Delorenzo, R. J. (1995). Recurrent spontaneous seizure activity in hippocampal neuronal networks in culture. Journal of Neurophysiology, 73, 1706–1711.Google Scholar
  3. 3.
    Bi, G. Q., & Poo, M. M. (1998). Synaptic modifications in cultured hippocampal neurons: dependence on spike timing, synaptic strength, and postsynaptic cell type. Journal of Neuroscience, 18, 10464–10472.Google Scholar
  4. 4.
    Boyden, E. S., Zhang, F., Bamberg, E., Nagel, G., Deisseroth, K. (2005). Millisecond-timescale, genetically targeted optical control of neural activity. Nature Neuroscience, 8, 1263–1268. doi: 10.1038/nn1525.CrossRefGoogle Scholar
  5. 5.
    Hayakawa, K., & Yasuhiko, H. (2013). Foreign gene transfer method by electroporation technique. Patent US 2013/0122592 A1.Google Scholar
  6. 6.
    Malyshev, A., Goz, R., LoTurco, J. J., Volgushev, M. (2015). Advantages and limitations of the use of optogenetic approach in studying fast-scale spike encoding. PLoS ONE, 10(4), e0122286. doi: 10.1371/journal.pone.0122286.CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media New York 2016

Authors and Affiliations

  1. 1.Institute of Higher Nervous Activity and Neurophysiology of RASMoscowRussia

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