Skip to main content
SpringerLink
Log in

Detection of cGMP in the Degenerating Retina

  • Protocol
  • First Online:
Guanylate Cyclase and Cyclic GMP

Part of the book series: Methods in Molecular Biology ((MIMB,volume 1020))

Abstract

Cyclic guanosine 3′-5′-monophosphate (cGMP) plays a key role in the physiological process of light detection in photoreceptor cells of the retina. However, there is also growing evidence that cGMP may be critically involved in some pathophysiological processes of the retina since degenerating photoreceptors in mouse models of retinitis pigmentosa and achromatopsia accumulate high levels of cGMP. Here, we describe methods that allow the detection, subcellular localization, and quantification of cGMP in the retina and propose that cGMP accumulation can be used as a biomarker for photoreceptor degeneration.

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

Access this chapter

Log in via an institution
Protocol
USD 49.95
Price excludes VAT (USA)
  • Available as PDF
  • Read on any device
  • Instant download
  • Own it forever
eBook
USD 84.99
Price excludes VAT (USA)
  • Available as EPUB and PDF
  • Read on any device
  • Instant download
  • Own it forever
Softcover Book
USD 139.00
Price excludes VAT (USA)
  • Compact, lightweight edition
  • Dispatched in 3 to 5 business days
  • Free shipping worldwide - see info
Hardcover Book
USD 109.99
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

References

  1. Arshavsky VY, Burns ME (2012) Photoreceptor signaling: supporting vision across a wide range of light intensities. J Biol Chem 287(3):1620–1626

    Article  PubMed  CAS  Google Scholar 

  2. Luo DG, Xue T, Yau KW (2008) How vision begins: an odyssey. Proc Natl Acad Sci USA 105(29):9855–9862

    Article  PubMed  CAS  Google Scholar 

  3. Biel M, Michalakis S (2009) Cyclic nucleotide-gated channels. Handb Exp Pharmacol 191:111–136

    Article  PubMed  CAS  Google Scholar 

  4. Karan S, Frederick JM, Baehr W (2010) Novel functions of photoreceptor guanylate cyclases revealed by targeted deletion. Mol Cell Biochem 334(1–2):141–155

    Article  PubMed  CAS  Google Scholar 

  5. Kuhn M (2009) Function and dysfunction of mammalian membrane guanylyl cyclase receptors: lessons from genetic mouse models and implications for human diseases. Handb Exp Pharmacol 191:47–69

    Article  PubMed  CAS  Google Scholar 

  6. Zhang X, Cote RH (2005) cGMP signaling in vertebrate retinal photoreceptor cells. Front Biosci 10:1191–1204

    Article  PubMed  CAS  Google Scholar 

  7. Yau KW, Nakatani K (1985) Light-induced reduction of cytoplasmic free calcium in retinal rod outer segment. Nature 313(6003):579–582

    Article  PubMed  CAS  Google Scholar 

  8. Baehr W, Palczewski K (2009) Focus on molecules: guanylate cyclase-activating proteins (GCAPs). Exp Eye Res 89(1):2–3

    Article  PubMed  CAS  Google Scholar 

  9. Biel M, Michalakis S (2007) Function and dysfunction of CNG channels: insights from channelopathies and mouse models. Mol Neurobiol 35(3):266–277

    Article  PubMed  CAS  Google Scholar 

  10. Ionita MA, Pittler SJ (2007) Focus on molecules: rod cGMP phosphodiesterase type 6. Exp Eye Res 84(1):1–2

    Article  PubMed  CAS  Google Scholar 

  11. Bowes C, Li T, Danciger M, Baxter LC, Applebury ML, Farber DB (1990) Retinal degeneration in the rd mouse is caused by a defect in the beta subunit of rod cGMP-phosphodiesterase. Nature 347(6294):677–680

    Article  PubMed  CAS  Google Scholar 

  12. Farber DB, Lolley RN (1974) Cyclic guanosine monophosphate: elevation in degenerating photoreceptor cells of the C3H mouse retina. Science 186(4162):449–451

    Article  PubMed  CAS  Google Scholar 

  13. Paquet-Durand F, Hauck SM, van Veen T, Ueffing M, Ekstrom P (2009) PKG activity causes photoreceptor cell death in two retinitis pigmentosa models. J Neurochem 108(3):796–810

    Article  PubMed  CAS  Google Scholar 

  14. Trifunovic D, Dengler K, Michalakis S, Zrenner E, Wissinger B, Paquet-Durand F (2010) cGMP-dependent cone photoreceptor degeneration in the cpfl1 mouse retina. J Comp Neurol 518(17):3604–3617

    Article  PubMed  CAS  Google Scholar 

  15. Hüttl S, Michalakis S, Seeliger M, Luo DG, Acar N, Geiger H, Hudl K, Mader R, Haverkamp S, Moser M, Pfeifer A, Gerstner A, Yau KW, Biel M (2005) Impaired channel targeting and retinal degeneration in mice lacking the cyclic nucleotide-gated channel subunit CNGB1. J Neurosci 25(1):130–138

    Article  PubMed  Google Scholar 

  16. Biel M, Seeliger M, Pfeifer A, Kohler K, Gerstner A, Ludwig A, Jaissle G, Fauser S, Zrenner E, Hofmann F (1999) Selective loss of cone function in mice lacking the cyclic nucleotide-gated channel CNG3. Proc Natl Acad Sci USA 96(13):7553–7557

    Article  PubMed  CAS  Google Scholar 

  17. Koch S, Sothilingam V, Garcia Garrido M, Tanimoto N, Becirovic E, Koch F, Seide C, Beck SC, Seeliger MW, Biel M, Mühlfriedel R, Michalakis S (2012) Gene therapy restores vision and delays degeneration in the CNGB1−/− mouse model of retinitis pigmentosa. Hum Mol Genet 21(20):4486–4496

    Article  PubMed  CAS  Google Scholar 

  18. Michalakis S, Mühlfriedel R, Tanimoto N, Krishnamoorthy V, Koch S, Fischer MD, Becirovic E, Bai L, Huber G, Beck SC, Fahl E, Buning H, Paquet-Durand F, Zong X, Gollisch T, Biel M, Seeliger MW (2010) Restoration of cone vision in the CNGA3−/− mouse model of congenital complete lack of cone photoreceptor function. Mol Ther 18(12):2057–2063

    Article  PubMed  CAS  Google Scholar 

  19. Thomas MG, Kumar A, Kohl S, Proudlock FA, Gottlob I (2011) High-resolution in vivo imaging in achromatopsia. Ophthalmology 118(5):882–887. doi:10.1016/j.ophtha.2010.08.053.

    Google Scholar 

  20. Thiadens AA, Somervuo V, van den Born LI, Roosing S, van Schooneveld MJ, Kuijpers RW, van Moll-Ramirez N, Cremers FP, Hoyng CB, Klaver CC (2010) Progressive loss of cones in achromatopsia: an imaging study using spectral-domain optical coherence tomography. Invest Ophthalmol Vis Sci 51(11):5952–5957

    Article  PubMed  Google Scholar 

  21. Sahel J, Bonnel S, Mrejen S, Paques M (2010) Retinitis pigmentosa and other dystrophies. Dev Ophthalmol 47:160–167

    Article  PubMed  Google Scholar 

  22. Hartong DT, Berson EL, Dryja TP (2006) Retinitis pigmentosa. Lancet 368(9549):1795–1809

    Article  PubMed  CAS  Google Scholar 

  23. Fiscus RR (2002) Involvement of cyclic GMP and protein kinase G in the regulation of apoptosis and survival in neural cells. Neuro-Signals 11(4):175–190

    Article  PubMed  CAS  Google Scholar 

  24. Sharma AK, Rohrer B (2007) Sustained elevation of intracellular cGMP causes oxidative stress triggering calpain-mediated apoptosis in photoreceptor degeneration. Curr Eye Res 32(3):259–269

    Article  PubMed  CAS  Google Scholar 

  25. Ulshafer RJ, Garcia CA, Hollyfield JG (1980) Sensitivity of photoreceptors to elevated levels of cGMP in the human retina. Invest Ophthalmol Vis Sci 19(10):1236–1241

    PubMed  CAS  Google Scholar 

  26. Olshevskaya EV, Calvert PD, Woodruff ML, Peshenko IV, Savchenko AB, Makino CL, Ho YS, Fain GL, Dizhoor AM (2004) The Y99C mutation in guanylyl cyclase-activating protein 1 increases intracellular Ca2+ and causes photoreceptor degeneration in transgenic mice. J Neurosci 24(27):6078–6085

    Article  PubMed  CAS  Google Scholar 

  27. Ramamurthy V, Niemi GA, Reh TA, Hurley JB (2004) Leber congenital amaurosis linked to AIPL1: a mouse model reveals destabilization of cGMP phosphodiesterase. Proc Natl Acad Sci USA 101(38):13897–13902

    Article  PubMed  CAS  Google Scholar 

  28. Sancho-Pelluz J, Arango-Gonzalez B, Kustermann S, Romero FJ, van Veen T, Zrenner E, Ekstrom P, Paquet-Durand F (2008) Photoreceptor cell death mechanisms in inherited retinal degeneration. Mol Neurobiol 38(3):253–269

    Article  PubMed  CAS  Google Scholar 

  29. Thapa A, Morris L, Xu J, Ma H, Michalakis S, Biel M, Ding XQ (2012) Endoplasmic reticulum stress-associated cone photoreceptor degeneration in cyclic nucleotide-gated channel deficiency. J Biol Chem 287(22):18018–18029

    Article  PubMed  CAS  Google Scholar 

  30. Paquet-Durand F, Beck S, Michalakis S, Goldmann T, Huber G, Muhlfriedel R, Trifunovic D, Fischer MD, Fahl E, Duetsch G, Becirovic E, Wolfrum U, van Veen T, Biel M, Tanimoto N, Seeliger MW (2011) A key role for cyclic nucleotide gated (CNG) channels in cGMP-related retinitis pigmentosa. Hum Mol Genet 20(5):941–947

    Article  PubMed  CAS  Google Scholar 

  31. Feil S, Zimmermann P, Knorn A, Brummer S, Schlossmann J, Hofmann F, Feil R (2005) Distribution of cGMP-dependent protein kinase type I and its isoforms in the mouse brain and retina. Neuroscience 135(3):863–868

    Article  PubMed  CAS  Google Scholar 

  32. Gamm DM, Barthel LK, Raymond PA, Uhler MD (2000) Localization of cGMP-dependent protein kinase isoforms in mouse eye. Invest Ophthalmol Vis Sci 41(9):2766–2773

    PubMed  CAS  Google Scholar 

  33. Tanaka J, Markerink-van Ittersum M, Steinbusch HW, De Vente J (1997) Nitric oxide-mediated cGMP synthesis in oligodendrocytes in the developing rat brain. Glia 19(4):286–297

    Article  PubMed  CAS  Google Scholar 

  34. Michalakis S, Geiger H, Haverkamp S, Hofmann F, Gerstner A, Biel M (2005) Impaired opsin targeting and cone photoreceptor migration in the retina of mice lacking the cyclic nucleotide-gated channel CNGA3. Invest Ophthalmol Vis Sci 46(4):1516–1524

    Article  PubMed  Google Scholar 

Download references

Acknowledgments

We thank Dr. Jan de Vente (University of Maastricht, Netherlands) for the gift of anti-cGMP antibody. This work was supported by the Deutsche Forschungsgemeinschaft (Bi484/4-1, Mi1238/1-2) and the National Eye Institute (P30EY12190 and R01EY019490).

Author information

Authors and Affiliations

  1. Center for Integrated Protein Science Munich (CIPSM), Ludwig-Maximilians-Universität, Munich, Germany

    Stylianos Michalakis & Martin Biel

  2. Department of Pharmacy, Center for Drug Research, Ludwig-Maximilians-Universität, Munich, Germany

    Stylianos Michalakis & Martin Biel

  3. Department of Cell Biology, University of Oklahoma Health Sciences Center, Oklahoma City, USA

    Jianhua Xu & Xi-Qin Ding

Authors
  1. Stylianos Michalakis
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Jianhua Xu
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Martin Biel
    View author publications

    You can also search for this author in PubMed Google Scholar

  4. Xi-Qin Ding
    View author publications

    You can also search for this author in PubMed Google Scholar

Editor information

Editors and Affiliations

  1. Addenbrooke’s Hospital, Department of Medicine, University of Cambridge, Hills Road, Cambridge, CB2 2QQ, United Kingdom

    Thomas Krieg

  2. Toxicology and Clinical Pharmacy, Department of Pharmacology, Universität Tübingen, Auf der Morgenstelle 8, Tübingen, 72076, Germany

    Robert Lukowski

Rights and permissions

Reprints and permissions

Copyright information

© 2013 Springer Science+Business Media, LLC

About this protocol

Cite this protocol

Michalakis, S., Xu, J., Biel, M., Ding, XQ. (2013). Detection of cGMP in the Degenerating Retina. In: Krieg, T., Lukowski, R. (eds) Guanylate Cyclase and Cyclic GMP. Methods in Molecular Biology, vol 1020. Humana Press, Totowa, NJ. https://doi.org/10.1007/978-1-62703-459-3_16

Download citation

  • DOI: https://doi.org/10.1007/978-1-62703-459-3_16

  • Published:

  • Publisher Name: Humana Press, Totowa, NJ

  • Print ISBN: 978-1-62703-458-6

  • Online ISBN: 978-1-62703-459-3

  • eBook Packages: Springer Protocols

Publish with us

Policies and ethics

Search

Navigation