Using anterograde tracing with HRPand antibodies (ABs) against neurofilaments, we show that regrowth of retinal ganglion cell (RGC) axons in the lizard Gallotia galloti commences only 2 months after optic nerve transection (ONS) and continues over at least 9 months. This is unusually long when compared to RGC axon regeneration in fish or amphibians. Following ONS, lizard RGCs up-regulate the immediate early gene C-JUN for 9 months or longer, indicating their reactive state. In keeping with the in vivo data, axon outgrowth from lizard retinal explants is increased above control levels from 6 weeks, reaches its maximum as late as 3 months, and remains elevated for at least 1 year after ONS. By means of BrdU incorporation assays and antiproliferating cell nuclear antigen immunohistochemistry, we show that the late axon outgrowth is not derived from new RGCs that might have arisen in reaction to ONS: no labeled cells were detected in lizard retinas at 0.5, 1, 1.5, 3, 6, and 12 months after ONS. Conversely, numbers of RGCs undergoing apoptosis were too low to be detectable in TUNEL assays at any time after ONS. These results demonstrate that retinal axon regeneration in G. galloti is due to axon regrowth from the resident population of RGCs, which remain in a reactive state over an extended time interval. Neurogenesis does not appear to be involved in RGC axon regrowth in G. galloti.
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