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M. O. Karl, S. Hayes, B. R. Nelson, T. A. Reh; Müller Glia Cell Proliferation After Neurotoxic Injury in the Murine Retina. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3550.
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Regeneration of retinal neurons following ocular injury in bird and fish is evident. After retinal damage, some Müller cells spontaneously dedifferentiate, enter the cell cycle and regenerate the retina. In fish this regeneration is complete, in the bird regeneration is limited. Data suggesting a similar process might occur in mammals is inconclusive.
Intraocular injections were performed with pulled capillary glass in adult mice (postnatal day 30). The left eye was injected with 2 µL of 0.1 M NMDA on day 1 and with various factors and BrdU on day 3. Additional BrdU injections were delivered intra-peritoneally at a dose of 50 mg/kg when indicated. The retinas were flatmounted or sectioned and analysed by immunhistochemistry.
Although Müller glia start to migrate extensively after NMDA dependent neurotoxic injury, they do not proliferate. We have confirmed that Müller glia can be induced to proliferate in a damaged adult murine retina (P30) in vivo. Application of proliferative factors after NMDA induced damage induces many cells across the retina to enter S-phase in vivo. As early as 4 hours after EGF application, 1168 ±157 sem cell nuclei /mm2 (n=4) incorporated BrdU (mean of 5 stacks on each flatmounted retina; n=4). Most of the BrdU labeled cells are also labeled with Sox2 antibodies and are located in the Müller glia layer. One day after the EGF injection, retinas contained more than 4500 BrdU labeled nuclei (653 ±35 sem BrdU labeled nuclei /mm2 (n=11)). At this time-point, proliferating cells - some of them labeled for Muller cell markers Sox2 or cyclin D3 - are spread across all retinal layers.Experiments in a Nestin-GFP reporter mouse showed GFP and BrdU double-labeled cells. Five to seven days after EGF injection, BrdU labeled nuclei accumulate in the ONL and RGC layer; however, the total number of BrdU labeled nuclei has declined. Application of recombinant protein Wnt3a, a small molecule Wnt pathway activator, FGF and sonic hedgehog also induce proliferation of Sox2 labeled cells, at levels similar to that observed with EGF.
Whether or not Müller glia can de-differentiate to become retinal progenitor cells, with the potential to regenerate retinal neurons, is currently under investigation.
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