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Retinal Cell Density In Chicks After Deprivation Myopia Measured With Adaptive-Optics Multiphoton Microscopy


Purpose: : Abnormal eye growth during deprivation myopia might modify the morphology of the retinal structures. We have used adaptive-optics multiphoton microscopy to image the retina of chicks with induced myopia. Cell density in those retinas was compared with those in a normal reference group.

Methods: : A custom adaptive-optics multiphoton microscope operating in backscattering-mode (Bueno et al., J. Biomed. Opt. 2010) was used to study the structure of chick retina. The instrument combines a Ti:Sapphire femtosecond laser for illumination and a pair of galvanometric mirrors as scanning unit. A photon-counting unit recorded the signal corresponding to two-photon excitation fluorescence (TPEF) originated from the chick retinal structures. A motorized stage allowed optical sectioning across the entire retina. The multiphoton images of the photoreceptors (PR) and ganglion cells’ (GC) layers were analyzed. Different parameters such as cell density, inter-cone distance, individual GC size and resolving power were obtained as a function of retinal eccentricity. Flat-mounted unstained chick retinas of both a control (N=4, near emmetropic) group and a group with induced myopia (N=3, mean refraction=-10.4 D) were imaged at varying retinal locations. Myopia was induced by placing an occluder in one of the eyes of every chick for about one week.

Results: : Single cells at the PR and GC layers provided enough TPEF signal for counting and analysis. For all samples, the local average intercenter PR distance increased with eccentricity. We did not find significant differences in PR density between the control eyes and those exhibiting deprivation myopia. GC density decreased with eccentricity, although this was on average 13% lower for myopic eyes. The size of the GCs increased approximately two-fold when comparing the area centralis and the peripheral retina. For each retinal location, the GC size for myopic retinas was larger than controls’ one (83% on average).

Conclusions: : Adaptive-optics multiphoton microscopy provided a clear visualization of single cells of the chick retina. The distribution of PRs and GCs depended on retinal location. For PR density, no significant differences were found when comparing control retinas and those from eyes with deprivation myopia. However, for every retinal location GCs showed a larger size in myopic eyes, which agrees with a decrease in the density. These results may help to better understand the impact of deprivation myopia in the chick retina.

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