Permeability of rod outer segment disk membranes as probed by ficoll density gradient centrifugation and by turbidimetry

Abstract

Osmotic volume changes of isolated disks prepared from bovine rod outer segments were investigated utilizing the capacity of osmotically intact disks to float on 2·5% ( w v ) Ficoll under hypotonic conditions. The amount of floating disks decreases steadily and approximately linearly when the osmolarity is increased stepwise with an osmotically active solute. The slope of the decrease becomes smaller for partially permeable substances and no decrease is observed for fully permeable solutes. All ions assayed by this method as well as sucrose were osmotically fully active to disk membranes. The buffer molecules Tris and imidazole partially penetrated the disk membranes at pH 7, whereas the zwitterionic buffer Hepes was impermeable. Ethanol showed absolutely no osmotic activity to disk membranes and, therefore, can easily penetrate these membranes. Concurrent turbidimetric measurements confirmed these results. Upon prolonged exposure of disk membranes to 5% ( w v ) Ficoll, a considerable modification of the osmotic properties of disk membranes is observed, indicating a significant interaction of 5% ( w v ) Ficoll with disk membranes. No influence of 2·5% ( w v ) Ficoll on the osmotic properties of disk membranes was detected. The restrictions of the use of Ficoll for disk membrane preparative purposes will be discussed.