ALTERATIONS IN THE LIGHT TRANSMISSION THROUGH SINGLE LENS FIBERS DURING CALCIUM-MEDIATED DISINTEGRATIVE GLOBULIZATION

Purpose. The purpose of this study was to examine changes in the light transmission through single cortical fibers isolated from the rat len s during the process of disintegrative globulization. Methods. Single cortical fibers were isolated from adult rat lens by treatment with tr ypsin in a solution containing 10 mM HEPES, 10 mM EDTA, and 280 mM suc rose (pH 7.4, 300 to 310 mOsm) HEPES-EDTA-sucrose (HES) solution. The isolated fibers were illuminated by a white light source, and tile rig ht transmission through the fibers was collected by a charge-coupled d evice camera and quantified by digital image analysis. In some experim ents, thin sections of fixed lens cells were examined using transmissi on electron microscopy. Results. Enzymatic dissociation of the lens yi elded elongated fibers, which, in the presence of Ringer's solution (c ontaining 2 mM Ca2+), underwent disintegrative globulization. Measurem ents of light transmission through elongated fibers suspended in HES s olution showed maximal transmission at the center of the fiber. Exposu re of the cortical fibers to Ringer's solution led to biphasic changes in the intensity of the transmitted light. Within 5 to 10 minutes of exposure to Ringer's solution, a general decrease in tile light transm ission across;; the long axis of the fiber was observed. Extended supe rfusion led to a local, apparent increase in light transmission corres ponding to the formation of membrane blebs and globules. Images of dis integrated globules focused above their equator showed bright halos wi th dark central zones. In electron micrographs, tile single fibers sho wed uniform electron density. No significant inhomogeneities or precip itation of intracellular crystallins was observed in globules generate d from fiber cells exposed to Ringer's solution; in addition, no high molecular weight protein aggregates were found in the globules. Conclu sions. Exposure to calcium alters the light-transmitting properties of isolated cortical fibers. The initial decrease in the average light t ransmittance of the fiber appears to be secondary to cell swelling and may relate to protein-based opacification. An apparent increase in li ght transmission through calcium-generated globules is Likely because of the Becke line generated by a mismatch between the refractive index of the medium and the globule cytoplasm and accentuated by the transi tion from rod-shaped to spheroidal morphology.