Abstract
Abstract: :
Purpose: The rho–/– mouse is a well established model of retinal degeneration. The purpose of this study was to make a detailed evaluation of the changes in thickness of all the retinal layers during the degenerative process and in particular to determine whether there is regional variation. Observed changes were correlated with patterns of retinal gene expression to better understand the underlying pathology. Methods: The thickness of the retina, individual layers and photoreceptor outer segment lengths were measured in both paraffin–embedded sections and in flatmounts stained with propidium iodide and PNA using confocal microscopy. Expression of photoreceptor marker genes including M and S opsins, cone arrestin and PDE6B was measured by qRT–PCR. Results: Retinal thicknesses measured from confocal z–series and conventional sections correlated extremely well. No significant differences in retinal thicknesses were observed between dorsal, ventral, nasal and temporal quadrants but there was a small increase in retinal thickness towards the periphery. Degeneration in the rho–/– retina was largely comparable between quadrants and from central to more periperal retina. mRNA expression of cone marker genes did not follow the known pattern of cone cell loss, and decreased to below WT levels at P30. The number of cone cells was confirmed to be unaltered at this time point, but the length of cone outer segments was reduced. Conclusions: We have demonstrated that confocal z–series of flatmounts can be used to reliably measure retinal thickness and provide an effective approach for analysing spatial variations within a single retina. Whilst cone photoreceptors remain grossly unaltered by defects in rod photoreceptors until the later stages of degeneration they do undergo molecular changes and exhibit shortened outer segment by P30.
Keywords: retinal degenerations: cell biology • photoreceptors • imaging/image analysis: non-clinical