A Novel Light Damage Paradigm til Anvendelse i Retinal Regeneration Studies i Adult zebrafisk
Summary
Flere lys skader protokoller er blevet beskrevet til skade fotoreceptorer og dermed fremkalde en retinal regenerering respons hos voksne zebrafisk. Denne protokol beskriver en forbedret metode, der kan anvendes i pigmenterede dyr, og at skader størstedelen af stang og kegle fotoreceptorer tværs af hele nethinden.
Abstract
Light-induceret retinal degeneration (LIRD) er almindeligt anvendt i både gnavere og zebrafisk for skader stang og kegle fotoreceptorer. Hos voksne zebrafisk, udløser fotoreceptordegenerering Müller gliaceller at genindtræde cellecyklus og producere forbigående-forstærkende stamceller. Disse forfædre fortsætter med at formere sig, når de trækker til det beskadigede område, hvor de i sidste ende giver anledning til nye fotoreceptorer. I øjeblikket er der to udbredte LIRD paradigmer, som hver resulterer i varierende grader af fotoreceptor tab og tilsvarende forskelle i regenerering reaktion. Som mere genetiske og farmakologiske værktøjer er tilgængelige til at teste betydningen af enkelte gener af interesse under regenereringen, er der behov for at udvikle en robust LIRD paradigme. Her beskriver vi en LIRD protokol, der resulterer i udbredt og sammenhængende tab af både stang og kegle fotoreceptorer, hvor vi har kombineret brug af to tidligere anvendte LIRD teknikker. EndvidereKan denne protokol udvides til brug i pigmenterede dyr, hvilket eliminerer behovet for at opretholde transgene linier af interesse på albino baggrund for LIRD studier.
Introduction
Light-induceret retinal degeneration (LIRD) er almindeligt anvendt i både gnavere og zebrafisk for skader stang og kegle fotoreceptorer. Hos voksne zebrafisk, udløser fotoreceptordegenerering Müller gliaceller at genindtræde cellecyklus og producere forbigående-forstærkende stamceller. Disse forfædre fortsætter med at formere sig, når de trækker til det beskadigede område, hvor de i sidste ende giver anledning til nye fotoreceptorer. I øjeblikket er der to udbredte LIRD paradigmer, som hver resulterer i varierende grader af fotoreceptor tab og tilsvarende forskelle i regenerering reaktion. Som mere genetiske og farmakologiske værktøjer er tilgængelige til at teste betydningen af enkelte gener af interesse under regenereringen, er der behov for at udvikle en robust LIRD paradigme. Her beskriver vi en LIRD protokol, der resulterer i udbredt og sammenhængende tab af både stang og kegle fotoreceptorer, hvor vi har kombineret brug af to tidligere anvendte LIRD teknikker. EndvidereKan denne protokol udvides til brug i pigmenterede dyr, hvilket eliminerer behovet for at opretholde transgene linier af interesse på albino baggrund for LIRD studier.
Protocol
Representative Results
Discussion
Her viser vi, at kombinere en kort UV-eksponering med en kontinuerlig Kraftigt lys resulterer i udbredt fotoreceptor tab og en robust regenerering svar. Sammenlignet med de individuelle LIRD metoder, er dette kombineret metode også den mest effektive protokol til skade både stave og tappe i begge halvdele af nethinden. Vigtigere er det, denne behandling er effektiv i pigmenterede dyr samt albino dyr.
Selvom vi dokumentation for, at de kombinerede protokoller resulterer i mere udbr…
Disclosures
The authors have nothing to disclose.
Acknowledgements
Forfatterne vil gerne takke Xixia Luo for fremragende fisk dyrehold og teknisk support. Dette arbejde blev finansieret af National Institutes of Health tilskud R21EY019401 (RT) og P30EY04068 (RT), og opstart midler til RT, herunder en ubegrænset bevilling fra forskning til at forebygge blindhed til Wayne State University, Department of Ophthalmology. JT blev understøttet af en Thomas C. Rumble Fellowship leveres af Wayne State University Graduate School.
Materials
| UV light source | Leica | EL600 |
| Glass Petri dish (150 x 20 mm) | Sigma-Aldrich/Pyrex | CLS3160152BO |
| 250 ml glass beaker | Sigma-Aldrich/Pyrex | CLS1000250 |
| 4 L glass beaker | Sigma-Aldrich/Pyrex | CLS10004L |
| Aluminum foil | Fisher | 01-213-105 |
| 250 W halogen lamps | Workforce | 265-669 |
| 1.8 L clear acrylic tanks | Aquaneering | ZT180T |
| 1.8 L clear acrylic tank lids | Aquaneering | ZT180LCL |
| Fan | Honeywell | HT-900 |
| Aerator | Tetra | 77853-900 |
| Thermometer | Cole-Parmer | YO-08008-58 |
| Bent forceps (5/45) | World Precision Instruments | 504155 |
References
- Bernardos, R. L., Barthel, L. K., Meyers, J. R., Raymond, P. A. Late-stage neuronal progenitors in the retina are radial Muller glia that function as retinal stem cells. J. Neurosci. 27, 7028-7040 (2007).
- Fausett, B. V., Goldman, D. A role for alpha1 tubulin-expressing Muller glia in regeneration of the injured zebrafish retina. J. Neurosci. 26, 6303-6313 (2006).
- Fimbel, S. M., Montgomery, J. E., Burket, C. T., Hyde, D. R. Regeneration of inner retinal neurons after intravitreal injection of ouabain in zebrafish. J. Neurosci. 27, 1712-1724 (2007).
- Sherpa, T., et al. Ganglion cell regeneration following whole-retina destruction in zebrafish. Dev. Neurobiol. 68, 166-181 (2008).
- Thummel, R., et al. Pax6a and Pax6b are required at different points in neuronal progenitor cell proliferation during zebrafish photoreceptor regeneration. Exp. Eye Res. 90, 572-582 (2010).
- Vihtelic, T. S., Hyde, D. R. Light-induced rod and cone cell death and regeneration in the adult albino zebrafish (Danio rerio) retina. J. Neurobiol. 44, 289-307 (2000).
- Vihtelic, T. S., Soverly, J. E., Kassen, S. C., Hyde, D. R. Retinal regional differences in photoreceptor cell death and regeneration in light-lesioned albino zebrafish. Exp. Eye Res. 82, 558-575 (2006).
- Yurco, P., Cameron, D. A. Responses of Muller glia to retinal injury in adult zebrafish. Vision Res. 45, 991-1002 (2005).
- Kassen, S. C., et al. Time course analysis of gene expression during light-induced photoreceptor cell death and regeneration in albino zebrafish. Dev. Neurobiol. 67, 1009-1031 (2007).
- Raymond, P. A., Barthel, L. K., Bernardos, R. L., Perkowski, J. J. Molecular characterization of retinal stem cells and their niches in adult zebrafish. BMC Dev. Biol. 6, 36 (2006).
- Thummel, R., et al. Characterization of Muller glia and neuronal progenitors during adult zebrafish retinal regeneration. Exp. Eye Res. 87, 433-444 (2008).
- Thummel, R., Kassen, S. C., Montgomery, J. E., Enright, J. M., Hyde, D. R. Inhibition of Muller glial cell division blocks regeneration of the light-damaged zebrafish retina. Dev. Neurobiol. 68, 392-408 (2008).
- Montgomery, J. E., Parsons, M. J., Hyde, D. R. A novel model of retinal ablation demonstrates that the extent of rod cell death regulates the origin of the regenerated zebrafish rod photoreceptors. J. Comp. Neurol. 518, 800-814 (2010).
- Morris, A. C., Scholz, T. L., Brockerhoff, S. E., Fadool, J. M. Genetic dissection reveals two separate pathways for rod and cone regeneration in the teleost retina. Dev. Neurobiol. 68, 605-619 (2008).
- Bignami, A., Dahl, D. The radial glia of Muller in the rat retina and their response to injury. An immunofluorescence study with antibodies to the glial fibrillary acidic (GFA) protein. Exp. Eye Res. 28, 63-69 (1979).
- Bringmann, A., et al. Muller cells in the healthy and diseased retina. Prog. Retin. Eye Res. 25, 397-424 (2006).
- Bringmann, A., Wiedemann, P. Muller glial cells in retinal disease. Ophthalmologica. 227, 1-19 (2012).
- Eisenfeld, A. J., Bunt-Milam, A. H., Sarthy, P. V. Muller cell expression of glial fibrillary acidic protein after genetic and experimental photoreceptor degeneration in the rat retina. Invest. Ophthalmol. Vis. Sci. 25, 1321-1328 (1984).
- Anderson, D. H., Guerin, C. J., Erickson, P. A., Stern, W. H., Fisher, S. K. Morphological recovery in the reattached retina. Invest. Ophthalmol. Vis. Sci. 27, 168-183 (1986).
- Bruce, A. E., Oates, A. C., Prince, V. E., Ho, R. K. Additional hox clusters in the zebrafish: divergent expression patterns belie equivalent activities of duplicate hoxB5 genes. Evol. Dev. 3, 127-144 (2001).
- Fawcett, J. W., Asher, R. A. The glial scar and central nervous system repair. Brain Res. Bull. 49, 377-391 (1999).
- Lewis, G. P., Fisher, S. K. Muller cell outgrowth after retinal detachment: association with cone photoreceptors. Invest. Ophthalmol. Vis. Sci. 41, 1542-1545 (2000).
- Fischer, A. J., Bongini, R. Turning Muller glia into neural progenitors in the retina. Mol. Neurobiol. 42, 199-209 (2010).
- Karl, M. O., Reh, T. A. Regenerative medicine for retinal diseases: activating endogenous repair mechanisms. Trends Mol. Med. 16, 193-202 (2010).
- Close, J. L., Liu, J., Gumuscu, B., Reh, T. A. Epidermal growth factor receptor expression regulates proliferation in the postnatal rat retina. Glia. 54, 94-104 (2006).
- Das, A. V., et al. Neural stem cell properties of Muller glia in the mammalian retina: regulation by Notch and Wnt signaling. Dev. Biol. 299, 283-302 (2006).
- Ikeda, T., Puro, D. G. Regulation of retinal glial cell proliferation by antiproliferative molecules. Exp. Eye Res. 60, 435-443 (1995).
- Liu, B., et al. Wnt signaling promotes muller cell proliferation and survival after injury. Invest. Ophthalmol. Vis. Sci. 54, 444-453 (2013).
- Osakada, F., et al. Wnt signaling promotes regeneration in the retina of adult mammals. J. Neurosci. 27, 4210-4219 (2007).
- Wan, J., et al. Preferential regeneration of photoreceptor from Muller glia after retinal degeneration in adult rat. Vision Res. 48, 223-234 (2008).
- Wan, J., Zheng, H., Xiao, H. L., She, Z. J., Zhou, G. M. Sonic hedgehog promotes stem-cell potential of Muller glia in the mammalian retina. Biochem. Biophys. Res. Commun. 363, 347-354 (2007).
- Roesch, K., et al. The transcriptome of retinal Muller glial cells. J. Comp. Neurol. 509, 225-238 (1002).
- Chang, G. Q., Hao, Y., Wong, F. Apoptosis: final common pathway of photoreceptor death in rd, rds, and rhodopsin mutant mice. Neuron. 11, 595-605 (1993).
- Marc, R. E., et al. Extreme retinal remodeling triggered by light damage: implications for age related macular degeneration. Mol. Vis. 14, 782-806 (2008).
- Portera-Cailliau, C., Sung, C. H., Nathans, J., Adler, R. Apoptotic photoreceptor cell death in mouse models of retinitis pigmentosa. Proc. Natl. Acad. Sci. U.S.A. 91, 974-978 (1994).
- Noell, W. K., Walker, V. S., Kang, B. S., Berman, S. Retinal damage by light in rats. Invest. Ophthalmol. 5, 450-473 (1966).
- Wenzel, A., Grimm, C., Samardzija, M., Reme, C. E. Molecular mechanisms of light-induced photoreceptor apoptosis and neuroprotection for retinal degeneration. Prog. Retin. Eye Res. 24, 275-306 (2005).
- Cicerone, C. M. Cones survive rods in the light-damaged eye of the albino rat. Science. 194, 1183-1185 (1976).
- La Vail, M. M. Survival of some photoreceptor cells in albino rats following long-term exposure to continuous light. Invest. Ophthalmol. 15, 64-70 (1976).
- Thomas, J. L., Nelson, C. M., Luo, X., Hyde, D. R., Thummel, R. Characterization of multiple light damage paradigms reveals regional differences in photoreceptor loss. Exp. Eye Res. 97, 105-116 (2012).
- Qin, Z., et al. FGF signaling regulates rod photoreceptor cell maintenance and regeneration in zebrafish. Exp. Eye Res. 93, 726-734 (2011).
- Craig, S. E., et al. The zebrafish galectin Drgal1-l2 is expressed by proliferating Muller glia and photoreceptor progenitors and regulates the regeneration of rod photoreceptors. Invest. Ophthalmol. Vis. Sci. 51, 3244-3252 (2010).
- Morris, A. C., Schroeter, E. H., Bilotta, J., Wong, R. O., Fadool, J. M. Cone survival despite rod degeneration in XOPS-mCFP transgenic zebrafish. Invest. Ophthalmol. Vis. Sci. 46, 4762-4771 (2005).
- Nelson, C. M., Hyde, D. R. Muller glia as a source of neuronal progenitor cells to regenerate the damaged zebrafish retina. Adv. Exp. Med. Biol. 723, 425-430 (2012).
- Powell, C., Elsaeidi, F., Goldman, D. Injury-dependent Muller glia and ganglion cell reprogramming during tissue regeneration requires Apobec2a and Apobec2b. J. Neurosci. 32, 1096-1109 (2012).
- Ramachandran, R., Reifler, A., Wan, J., Goldman, D. Application of Cre-loxP recombination for lineage tracing of adult zebrafish retinal stem cells. Methods Mol. Biol. 884, 129-140 (2012).
- Ramachandran, R., Zhao, X. F., Goldman, D. Ascl1a/Dkk/beta-catenin signaling pathway is necessary and glycogen synthase kinase-3beta inhibition is sufficient for zebrafish retina regeneration. Proc. Natl. Acad. Sci. U.S.A. 108, 15858-15863 (2011).
- Ramachandran, R., Zhao, X. F., Goldman, D. Insm1a-mediated gene repression is essential for the formation and differentiation of Muller glia-derived progenitors in the injured retina. Nat. Cell Biol. 14, 1013-1023 (2012).
- Wan, J., Ramachandran, R., Goldman, D. HB-EGF is necessary and sufficient for Muller glia dedifferentiation and retina regeneration. Dev. Cell. 22, 334-347 (2012).
- Morris, A. C., Forbes-Osborne, M. A., Pillai, L. S., Fadool, J. M. Microarray analysis of XOPS-mCFP zebrafish retina identifies genes associated with rod photoreceptor degeneration and regeneration. Invest. Ophthalmol. Vis. Sci. 52, 2255-2266 (2011).
- Thummel, R., Bailey, T. J., Hyde, D. R. In vivo electroporation of morpholinos into the adult zebrafish retina. J. Vis. Exp. (58), e3603 (2011).
