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A. K. Ball, M. L. Kisilak, L. G. Epps, E. L. Irving, M. C. W. Campbell; Rod Density Changes Inversely Correlated With Eye Size in the Chick Retina. Invest. Ophthalmol. Vis. Sci. 2008;49(13):3595. doi: https://doi.org/.
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Cones are among the first retinal cells to differentiate, while rods differentiate over a longer period extending up to 2 wks after birth in some species. The surface area of the chick retina increases by approximately 75% between days 0 and 14. If rod differentiation occurs at the same rate as the rapid increase in eye size, the packing density of rods should not change significantly during this time. In this study we compared the changes in rod density and eye size that occur between days 0 and 14.
Ocular dimensions were determined in vivo by A-scan ultrasonography at 0, 7, 9 and 14 days of age and some birds were killed at each timepoint. Retinas were isolated and fixed in 4% phosphate buffered formaldehyde for 2 hrs and rinsed in PBS. A 5 mm2 sample immediately above the pecten corresponding to the central retina was selected for analysis. Retinal samples were incubated overnight in PBS containing 20 µg WGA Alexa 594, 40 µg PNA Alexa 488 (Molecular Probes), 0.1% Triton-X 100, and 0.1% DMSO. Retinas were washed in PBS and mounted in PBS/glycerol. Retinas were examined by epi-fluorescence and confocal microscopy. Rod and cone density was determined using Image-J and nearest neighbor distances using Biotas. The density of rods was compared to the 1/axial length2 at each time point.
The linear axial length of the eye was determined to be 7.6±0.9, 8.7±0.8, 9.0±0.8, and 9.9±0.9 mm at days 0, 7, 9 and 14, respectively. Rods were labeled only by WGA and cones only by PNA. The average distance between rods was 6.2±0.4 µm and the distance between cones was 2.5±0.2 µm. Rod density was determined to be 69±12, 46±9, 37±6, and 38±6 per µm2 at days 0, 7, 9 and 14, respectively, representing an 81% decrease between 0 and 14 days. The greatest change in both axial length and rod density occurred between 0 and 7 days. Weak WGA staining of some photoreceptors at day 0 suggested that rods were still being differentiated at this time point. The rate of exponential decrease in rod density with age (53 days) was not significantly different from twice the rate of the increase in axial length (25 days). There was a significant correlation between rod density and 1/axial length2 (p<0.001).
These findings demonstrate significant changes in rod density that are consistent with a linear expansion of the retina occurring at the same rate as axial length change. This suggests that most rod differentiation is complete before hatching and that rod density decreases as the chick eye grows.
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