Skip to main content
SpringerLink
Log in

Selecting Channelrhodopsin Constructs for Optimal Visual Restoration in Differing Light Conditions

  • Protocol
  • First Online:
Channelrhodopsin

Part of the book series: Methods in Molecular Biology ((MIMB,volume 2191))

Abstract

Channelrhodopsin (ChR)-based optogenetics is one promising approach to restore vision in photoreceptor degenerative diseases such as retinitis pigmentosa (RP) and age-related macular degeneration (AMD). Currently, a large number of ChRs from different alga species as well as engineered variants are available. They vary with their light response properties like peak sensitive wavelength (λmax), current amplitude, and kinetics. Therefore, it is important to choose an appropriate ChR for practical applications, such as vision restoration. Here we describe a standard laboratory protocol for characterizing properties of ChRs in in vitro in human embryonic kidney (HEK) cells. Based on such characterization, we also discuss the criteria for selecting optimal ChRs for optogenetic vision restoration.

This is a preview of subscription content, log in via an institution to check access.

Access this chapter

Log in via an institution

Subscribe and save

Springer+ Basic
$34.99 /Month
  • Get 10 units per month
  • Download Article/Chapter or eBook
  • 1 Unit = 1 Article or 1 Chapter
  • Cancel anytime
Subscribe now

Buy Now

Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 89.00
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 119.99
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 169.99
Price excludes VAT (USA)
  • Durable hardcover edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info

Tax calculation will be finalised at checkout

Purchases are for personal use only

Institutional subscriptions

Similar content being viewed by others

References

  1. Bi A, Cui J, Ma YP, Olshevskaya E, Pu M, Dizhoor AM, Pan ZH (2006) Ectopic expression of a microbial-type rhodopsin restores visual responses in mice with photoreceptor degeneration. Neuron 50(1):23–33. https://doi.org/10.1016/j.neuron.2006.02.026

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  2. Tomita H, Sugano E, Isago H, Hiroi T, Wang Z, Ohta E, Tamai M (2010) Channelrhodopsin-2 gene transduced into retinal ganglion cells restores functional vision in genetically blind rats. Exp Eye Res 90(3):429–436. https://doi.org/10.1016/j.exer.2009.12.006

    Article  CAS  PubMed  Google Scholar 

  3. Tomita H, Sugano E, Yawo H, Ishizuka T, Isago H, Narikawa S, Kugler S, Tamai M (2007) Restoration of visual response in aged dystrophic RCS rats using AAV-mediated channelopsin-2 gene transfer. Invest Ophthalmol Vis Sci 48(8):3821–3826. https://doi.org/10.1167/iovs.06-1501

    Article  PubMed  Google Scholar 

  4. Lin B, Koizumi A, Tanaka N, Panda S, Masland RH (2008) Restoration of visual function in retinal degeneration mice by ectopic expression of melanopsin. Proc Natl Acad Sci U S A 105(41):16009–16014. https://doi.org/10.1073/pnas.0806114105

    Article  PubMed  PubMed Central  Google Scholar 

  5. Lagali PS, Balya D, Awatramani GB, Munch TA, Kim DS, Busskamp V, Cepko CL, Roska B (2008) Light-activated channels targeted to ON bipolar cells restore visual function in retinal degeneration. Nat Neurosci 11(6):667–675. https://doi.org/10.1038/nn.2117

    Article  CAS  PubMed  Google Scholar 

  6. Zhang Y, Ivanova E, Bi A, Pan ZH (2009) Ectopic expression of multiple microbial rhodopsins restores ON and OFF light responses in retinas with photoreceptor degeneration. J Neurosci 29(29):9186–9196. https://doi.org/10.1523/jneurosci.0184-09.2009

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  7. Busskamp V, Duebel J, Balya D, Fradot M, Viney TJ, Siegert S, Groner AC, Cabuy E, Forster V, Seeliger M, Biel M, Humphries P, Paques M, Mohand-Said S, Trono D, Deisseroth K, Sahel JA, Picaud S, Roska B (2010) Genetic reactivation of cone photoreceptors restores visual responses in retinitis pigmentosa. Science (New York, NY) 329(5990):413–417. https://doi.org/10.1126/science.1190897

    Article  CAS  Google Scholar 

  8. Pan ZH, Ganjawala TH, Lu Q, Ivanova E, Zhang Z (2014) ChR2 mutants at L132 and T159 with improved operational light sensitivity for vision restoration. PLoS One 9(6):e98924. https://doi.org/10.1371/journal.pone.0098924

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  9. Pan Z-H, Lu Q, Bi A, Dizhoor AM, Abrams GW (2015) Optogenetic approaches to restoring vision. Annu Rev Vis Sci 1(1):185–210. https://doi.org/10.1146/annurev-vision-082114-035532

    Article  PubMed  Google Scholar 

  10. Ganjawala TH, Lu Q, Fenner MD, Abrams GW, Pan ZH (2019) Improved CoChR variants restore visual acuity and contrast sensitivity in a mouse model of blindness under ambient light conditions. Mol Ther. https://doi.org/10.1016/j.ymthe.2019.04.002

  11. Nagel G, Ollig D, Fuhrmann M, Kateriya S, Musti AM, Bamberg E, Hegemann P (2002) Channelrhodopsin-1: a light-gated proton channel in green algae. Science (New York, NY) 296(5577):2395–2398. https://doi.org/10.1126/science.1072068

    Article  CAS  Google Scholar 

  12. Nagel G, Szellas T, Huhn W, Kateriya S, Adeishvili N, Berthold P, Ollig D, Hegemann P, Bamberg E (2003) Channelrhodopsin-2, a directly light-gated cation-selective membrane channel. Proc Natl Acad Sci U S A 100(24):13940–13945. https://doi.org/10.1073/pnas.1936192100

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  13. Ernst OP, Lodowski DT, Elstner M, Hegemann P, Brown LS, Kandori H (2014) Microbial and animal rhodopsins: structures, functions, and molecular mechanisms. Chem Rev 114(1):126–163. https://doi.org/10.1021/cr4003769

    Article  CAS  PubMed  Google Scholar 

  14. Govorunova EG, Sineshchekov OA, Li H, Janz R, Spudich JL (2013) Characterization of a highly efficient blue-shifted channelrhodopsin from the marine alga Platymonas subcordiformis. J Biol Chem 288(41):29911–29922. https://doi.org/10.1074/jbc.M113.505495

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  15. Kleinlogel S, Feldbauer K, Dempski RE, Fotis H, Wood PG, Bamann C, Bamberg E (2011) Ultra light-sensitive and fast neuronal activation with the Ca(2)+−permeable channelrhodopsin CatCh. Nat Neurosci 14(4):513–518. https://doi.org/10.1038/nn.2776

    Article  CAS  PubMed  Google Scholar 

  16. Mattis J, Tye KM, Ferenczi EA, Ramakrishnan C, O'Shea DJ, Prakash R, Gunaydin LA, Hyun M, Fenno LE, Gradinaru V, Yizhar O, Deisseroth K (2012) Principles for applying optogenetic tools derived from direct comparative analysis of microbial opsins. Nat Methods 9(2):159–172. https://doi.org/10.1038/nmeth.1808

    Article  CAS  Google Scholar 

  17. Klapoetke NC, Murata Y, Kim SS, Pulver SR, Birdsey-Benson A, Cho YK, Morimoto TK, Chuong AS, Carpenter EJ, Tian Z, Wang J, Xie Y, Yan Z, Zhang Y, Chow BY, Surek B, Melkonian M, Jayaraman V, Constantine-Paton M, Wong GK, Boyden ES (2014) Independent optical excitation of distinct neural populations. Nat Methods 11(3):338–346. https://doi.org/10.1038/nmeth.2836

    Article  CAS  PubMed  PubMed Central  Google Scholar 

  18. Govorunova EG, Sineshchekov OA, Janz R, Liu X, Spudich JL (2015) NEUROSCIENCE. Natural light-gated anion channels: A family of microbial rhodopsins for advanced optogenetics. Science (New York, NY) 349(6248):647–650. https://doi.org/10.1126/science.aaa7484

    Article  CAS  Google Scholar 

  19. Wietek J, Prigge M (2016) Enhancing channelrhodopsins: an overview. Methods Mol Biol 1408:141–165. https://doi.org/10.1007/978-1-4939-3512-3_10

    Article  CAS  PubMed  Google Scholar 

Download references

Author information

Authors and Affiliations

  1. Department of Ophthalmology, Visual and Anatomical Sciences (OVAS), Wayne State University School of Medicine, Detroit, MI, USA

    Tushar H. Ganjawala & Zhuo-Hua Pan

Authors
  1. Tushar H. Ganjawala
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Zhuo-Hua Pan
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Department of Chemistry and Biochemistry, Worcester Polytechnic Institute, Worcester, MA, USA

    Robert E. Dempski

Rights and permissions

Reprints and permissions

Copyright information

© 2021 Springer Science+Business Media, LLC, part of Springer Nature

About this protocol

Check for updates. Verify currency and authenticity via CrossMark

Cite this protocol

Ganjawala, T.H., Pan, ZH. (2021). Selecting Channelrhodopsin Constructs for Optimal Visual Restoration in Differing Light Conditions. In: Dempski, R. (eds) Channelrhodopsin. Methods in Molecular Biology, vol 2191. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0830-2_12

Download citation

  • DOI: https://doi.org/10.1007/978-1-0716-0830-2_12

  • Published:

  • Publisher Name: Humana, New York, NY

  • Print ISBN: 978-1-0716-0829-6

  • Online ISBN: 978-1-0716-0830-2

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation