Study of diffusion characters of macromolecules among different intraretinal layers

Objective To investigate the diffusion characters of macromolecules among different intraretinal layers.Methods The fresh separated pig retina was spread on the diffusion apparatus which was designed by us.Carboxyfluorescein(Mr 376) and FITC-dextrans of various molecular weights(Mr 4 400,9 300,19 600,38 900,71 200 and 150 000) were dissolved in RPMI 1640 solutions which were applying to inner or outer surface of retina.After 1 hour,the retina were observed under a fluorescence microscope and then stained with hematoxylin-eosin.Results The distribution of Carboxyfluorescein and dextrans with different molecular weight in the normal pig retina varied.No matter Carboxyfluorescein was applying to inner or outer surface,the inner nuclear layer (INL) and outer nuclear layer (ONL) were the brightest;FITC-dextrans applying to inner retinal surface,Mr 4 400 dextran were largely arrested at INL.The INL of the Mr 9 300~71 200 dextran diffusion retina section became dark and the nerve fiber layer (NFL) and inner plexiform layer got brighter.As for Mr 150 000 dextran,the NFL was bright and the other layers were dark.FITC-dextrans applying to outer retinal surface,most dextrans were blocked before ONL.Conclusion ONL and INL may act as bottle-neck barriers to diffusion of macromolecules.According to the upper limit of the molecular size of macromolecule which can diffuse across the layers,the layers were ranked as follows (from small to large):ONLINLinner and outer plexiform layerNFL and inner limiting membrane.