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Sally A McFadden, Guang Zeng, Laura Walker, Alexandra P Metse, Christine Frances Wildsoet; Changes in Cell Density in the Retinal Ganglion Cell Layer After Optic Nerve Section in Young Guinea Pigs. Invest. Ophthalmol. Vis. Sci. 2015;56(7 ):3623.
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© ARVO (1962-2015); The Authors (2016-present)
Optic nerve section (ONS) induces degeneration of retinal ganglion cells (RGCs). We studied the early effects of ONS on cell density in the RGC layer in young guinea pigs.
The optic nerve (ON) was sectioned in 2 day-old anaesthetised guinea pigs. The fibres of the ON were cut leaving the sheaf intact. Control animals received sham surgery. At 3, 5, 7, 14 or 63 days after surgery, retinas were extracted and whole mounts made which were stained to count cells in the RGL layer. Cell nuclei were counter stained using DAPI (n=33); displaced amacrine cells (DACs) were detected using Anti-Choline Acetyltransferase (ChAT) and doubled stained with DAPI (n= 10); and microglia were detected using antibody against Macroglia-specific Protein (Iba1, n= 10). Counts were made along each of 8 radii at 1 mm intervals (64 samples/retina). Additionally, whole mounts were stained with Cresyl Violet to determine the frequency of different soma sizes in the RGC layer (n=8). Gaussian fits were made to cell distributions and used to discriminate RGC cells from DACs and microglia based on soma size.
In control eyes, nuclei density reduced by 11% between 5-18 days of age (retinal average 2881 to 2776 nucleui/mm2, p < 0.001). Of these, 49% were RGCs, and 23% were DACs positive for ChAT. Relative to the control eyes, a significant reduction in total nuclei density occurred 14 days after ONS (p = 0.003, -15%), but no significant decline was detected at 3, 5, or 7 days after ONS (-5%,+1%,-3% respectively). ChAT positive DAC density relatively increased 7 and 14 days after ONS, and microglia increased in the fibre layer 7 days after ONS. After accounting for these increases, the remaining RGC density reduced 14 days after ONS, but not earlier. However, the average soma size of the RGC frequency distribution was smaller 3 days after ONS (mean radius; Control , 4.6±0.85 µm; ONS, 4.1±0.85 µm; p = 0.004), and reduced further by 7 days (ONS, 4.0±0.5 µm) and by 14 days after ONS, nearly all large cells had disappeared. In contrast, the frequency of smaller cells classified as DACs and microglia increased 7 days after ONS.
RGC loss occurs 14 days after ONS, but reductions occur in soma size within 3 days of surgery suggesting RGC integrity may be compromised within days of ONS.
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