UCLA

Ingestion of photoreceptor outer segments (POS/OS) by the retinal pigment epithelium (RPE) is a fundamentally important process for vision. The RPE form an actin based phagocytic cup that extends along the OS and engages in trogocytosis to break off a piece of the OS for subsequent ingestion. This process is highly dynamic, and so we have used live cell imaging with high temporal resolution to understand the kinetics of ingestion and the localization of different relevant proteins throughout the ingestion process. We have found that the phagocytic cup is initiated by mDia1 formin activity. The Arp2/3 complex creates branched actin around the rim of the cup and beneath phagosomes to hold them in their proper location for scission and ingestion. Once excised, Myosin 7a traffics the phagosome basally into the cell for subsequent degradation. When photoreceptor-RPE interaction stops, such as during a retinal detachment, there are consequences to the photoreceptor synapse. Utilizing the high resolution capabilities of electron microscope (EM) tomography, we then explored the ultrastructure of the rod photoreceptor synapse (or rod spherule, RS) after 7 days of retinal detachment. 7 days of retinal detachment results in significant degeneration to the synaptic ultrastructure at the outer plexiform layer, including a loss of rod bipolar dendrites, simplification of horizontal cell axon terminals, fragmentation of the synaptic ribbon, and a simplification of the rod spherule invagination membrane. Following up, we then explored the timescale of these degenerative changes, performing tomography on synapses in retinas detached for 30min., 1hr., 24hrs., 48hrs., and 72hrs., in order to understand how long of a detachment may be too late for full recovery. By 24hrs. of detachment, there is significant remodeling including elongated ribbons and simplified horizontal cell processes; at least one rod bipolar cell process per rod spherule retracts by 24hrs. of detachment in a majority of cases. 24hrs. is likely the latest time point for reattachment without incurring long term visual deficits due to synaptic degeneration. By better understanding the way that OS are phagocytosed, as well as the consequences to the rod synapse when they are not, we can learn significant additional information about the health and maintenance of the rod photoreceptor.