The distribution of filipin-sterol complexes in photoreceptor synaptic membranes

The polyene antibiotic filipin, which binds to membrane sterols, has been used to investigate the relative distribution of cholesterol at photoreceptor synaptic junctions in the chick retina. Following anesthesia and aldehyde perfusion fixation, the retina is removed and immersed in fixative solution containing the filipin for 36-48 hours. The retinas are then processed for freeze-fracture. Electron microscopy of freeze-fracture replicas demonstrates that the filipin-sterol complexes are not evident between intramembrane particles of the presynaptic and postsynaptic particle arrays that are present at ribbon and basal junctions. In contrast, the synaptic vesicle fusion zone of ribbon junctions contains large numbers of filipin-sterol complexes which are observed merging with the free margins of the presynaptic particle array. There is a scarcity of such complexes, however, around the free margins of basal junction presynaptic particle arrays. These latter sites do not contain a vesicle fusion zone. Particle-poor areas of membrane that surround postsynaptic particles arrays of ribbon and basal junctions also do not contain filipin binding sites. The nonsynaptic membrane of photoreceptor terminals contains large numbers of filipin-sterol complexes, less tightly packed than in the synaptic vesicle fusion zone. Coated vesicle fusion sites in the presynaptic membrane contain groups of intramembrane particles but the filipin-sterol complexes are excluded from these sites. The observations suggest that synaptic membrane domains which interact with cytoskeletal components, such as clathrin and pre- and postsynaptic densities contain less cholesterol than other domains such as the synaptic vesicle fusion zones.