Research is proposed to investigate the hypothesis that specific cell surface marker molecules are involved in the process of retinal rod outer segment shedding and phagocytosis. Rod outer segments (ROS) shed packets of discs into the retinal pigment epithelium (RPE) on a daily basis. Yet only a certain, relatively constant, number of ROS shed each day. What mechanism exists to determine which ROS tips will be ingested by the RPE and which will be ignored? A possible explanation is that there are changes in specific molecules or groups of molecules at the retina-RPE interface that mark the tips that are to be ingested. Preliminary studies in our laboratory using an in vitro encubation method that supports normal ROS shedding in the eyecup of the frog, Rana pipiens, have shown that certain sugars inhibit shedding when added to the incubation medium, an effect possibly due to a disturbance in the mechanism by which the RPE recognizes phagosomes as """"""""not self"""""""".
The first aim of the proposed project is to continue and to extend these studies in order to determine the molecular basis for sugar inhibition of shedding. Secondly, using a neural retina-free preparation and an artificial substrate, gold colloid, surface binding and uptake of the substrate by the RPE cells will be examined by both transmission and scanning electron microscopy. By altering the surface features of the gold colloid through treating it with macromolecules we will determine the molecular requirements for attachment and phagocytosis by the RPE. Finally, using lectin-gold colloid preparations we will examine the surface sugar distribution of the ROS and RPE in tissues maintained for long periods in either constant light or constant darkness, treatments that are known to profoundly disturb the normal ROS shedding process. The daily ROS shedding phenomenon appears to be a vital process in the maintenance of healthy retinas in both laboratory animals and man. Yet we still do not know how this process is controlled with the precision which has been demonstrated in many experimental studies. The proposed study will produce considerable advances in our knowledge of the relationship between the retina and the RPE and will open a new area of understanding of the basis of both health and disease in these tissues.
