Library

Notes: Summary Retinae from two day old rats were used in this study and the cultures were handled according to standard methods used in this laboratory. In the first few days of cultivation an abundant outgrowth of nerve fibers into the cell-free medium was observed. These fibers later degenerated and by the beginning of the second week they had completely disappeared. In the living cultures, differentiating ganglion cells, bipolar and horizontal neurons could be seen in the main explant in association with various types of glial cells. Rod cells became arranged as epithelial sheets or as clusters of cells which often formed rosettes. The nuclei of these sensory cells possessed a characteristic chromatin pattern by which they always could be differentiated from other cells in the cultures. Cytoplasmic extensions that developed from the free surface of the sensory rod cells were observed within a week following explantation. A “limiting membrane” separated these extensions from the nucleated part of the rod cells. Morphologic details of the different neuronal cell types could be demonstrated in cultures by Bodian's silver impregnation technique. With the electron microscope, retinal development in culture was observed and compared to the development of the retina of the intact eye. Cilia developed from processes extending from the rod cell free surface. These processes were the rod cell inner segments in which many mitochondria were seen. At the bases of these segments terminal bars developed forming the “outer limiting membrane”. In the area of the terminal bars microvillous extensions projected between the rod cell inner segments. After twelve days in vitro a bulb-like enlargement containing a lamellar membrane system developed at the end of the cilium. This bulb-like enlargement was a beginning of the rod cell outer segment. The lamellar system did not acquire the symmetry or precise organization during cultivation that was observed in the retina of the intact eye. The distinguishing characteristics of individual neuronal cell types seen in cultivated retinae were the same as those described for their counterparts in the retina in situ, but regular plexiform layers failed to develop. Likewise, there were no indications of typical synapses in the neuropils of the cultures. There were many processes containing vesicles similar to those in presynaptic endings and mitochondria but membrane thickenings were not apparent. The results indicate that the retina cultivated in vitro does not behave as an organized entity. The component cells dissociated more and more with time, and developmental differentiation was observed only at the cellular level.