Abstract
Abstract: :
Purpose: The zebrafish (Danio rerio) is a model organism for the genetic analysis of vertebrate photoreceptor development. The zebrafish has a duplex retina containing 4 types of cone photoreceptors arranged in a highly ordered crystalline mosaic and rod photoreceptors. Throughout the life of the animal, retinal growth, including the addition of new cones, occurs at the retinal margin from a pool of mitotic progenitors. Rods, however, develop from a population of progenitors that reside in the inner nuclear layer. The goal of this project was to examine the development and organization of rod photoreceptors in relation to the cones using transgenic zebrafish. Methods: Transgenic zebrafish, expressing green fluorescent protein (GFP) under the control of the Xenopus opsin promoter, were used throughout the study. The transgene was crossed into the albino background to facilitate in vivo imaging. The spatial and temporal pattern of rod differentiation was investigated by confocal and multiphoton microscopy in live embryos or retinal whole mounts maintained in teleost saline. Cones were identified by immunolabeling of frozen sections. Results: Analysis of GFP expression in whole mount retinas by confocal microscopy enabled imaging of entire rod photoreceptors as well as analysis of the spatial arrangement of rods across large areas of the retina. These revealed a regular pattern of rods within the cone mosaic. The rod spherules were arranged in rows perpendicular to the annular growth at the retinal margin. The rod nuclei, which are vitreal to the cone nuclei, were tightly packed into 1 to 3 layers depending upon the size of the animal and the region of the retina. The myoids projected in a regular mosaic pattern around the UV-sensitive cones as demonstrated by the spacing of the projections and by immunochemistry. The rod ellipsoids, identified by the dense cluster of mitochondria that exclude the soluble GFP, formed tight groups associated with the tips of the UV cones and were arranged in rows parallel to the retinal margin. As retinal growth continued, new rods at the margin appeared to project around the UV-sensitive cones. In the central retina new rods projected in rows parallel to the cone nuclei. Conclusions: The data suggest a tightly regulated spatial arrangement of rod photoreceptors directed by the cone mosaic. This arrangement provides a model to explore genetic mechanisms underlying rod and cone interactions during development and in retinal dystrophies.
Keywords: 564 retinal development • 606 transgenics/knock-outs • 471 microscopy: confocal/tunneling