Glial, vascular, and neuronal cytogenesis in whole-mounted cat retina

A method was developed for detecting cytogenesis in retinal whole-mount preparations by bromodeoxyuridine (BrdU) immunohistochemistry. Because BrdU is a nonspecific marker that labels all cells in the S phase of the cell cycle, it is ideally combined with other cell-specific markers to study the cytogenesis of specific cell types. Double-label protocols to visualize mitotically active astrocytes and cells associated with the forming vasculature have been developed and applied to the retina. This approach revealed that, during normal development of the kitten retina, vascular mitogenesis occurs predominantly in the ganglion cell and nerve fiber layers, where the inner retinal plexus is formed by a process involving transformation of mesenchymal precursor cells and division of vascular endothelial cells. The peak density of vascular mitogenesis moved in a central-to-peripheral manner and was associated with the leading edge of the forming capillary plexus. A small number of dividing vascular endothelial cells was also associated with angiogenesis, the process responsible for the formation of the outer retinal plexus, vessels at the area centralis, and the radial peripapillary capillaries. Cytogenesis associated with astrocytes occurred in the ganglion cell and nerve fiber layers but was apparent predominantly at or close to the optic nerve head. Confirming earlier studies, neuronal mitogenesis was shown to occur predominantly at the ventricular zone, first at the area centralis and spreading peripherally with increasing maturity. A second region of neuronal cytogenesis, at the subventricular zone, was also apparent. Tissue hyperoxia decreased the rate of vasculogenic cell division but had no apparent effect on neurogenic or astrocytic cell division. Four distinct zones of cell generation were therefore identified within the retina, each associated with either glial, vascular, or neuronal cytogenesis. Thus, BrdU immunohistochemistry in whole-mounted retinal preparations offers a fast and reliable alternative to [3H]thymidine autoradiography for the study of the topography of cytogenesis during development.