Confocal laser scanning microscopy imaging of dynamic TMREmovement in the mitochondria of epithelial and superficial corticalfiber cells of bovine lenses

[摘要] Purpose: Recent confocal laser scanning microscopy studies of themitochondria of vertebrate lenses show a striking difference in thedistribution and morphology of the mitochondria of lens epithelial andsuperficial cortical cells. This study, using confocal microscopy, wasundertaken to image the movement of the mitochondria specific dyetetramethylrhodamine ethyl ester (TMRE) in the epithelium andsuperficial cortex of whole live bovine lens.Methods: Cultured bovine lenses were loaded with 5 μg/ml TMRE for15 min at room temperature. TMRE fluorescence was acquired with a Zeiss510 (configuration META 18) confocal laser scanning microscope for 10 to15 min using 488 nm Argon laser excitation and 505 nm long pass emissionfilter settings. The uncoupler of the electron transport chainpotential, carbonyl cyanide m-chlorophenylhydrazone (CCCP, 32.5 μM), was used to demonstrate the fluorescent specificity of TMRE.Results: Multidirectional dynamic movement of TMRE was observed inepithelial cells and bidirectional dynamic movement was seen in thesuperficial cortical fiber cells of live bovine lenses. In theepithelium, the movement of TMRE fluorescence was up to 5 μm/minwhereas in the superficial cortex the observed movement was up to 18.5μm/min. The movement of TMRE fluorescence was abolished withtreatment with the uncoupler, CCCP.Conclusions: The observed dynamics of TMRE fluorescence movement mayrepresent actual mitochondrial movement, indicating the dynamic state ofthe mitochondria in both lens epithelium and superficial cortex. Thatthis activity is found not only in the epithelium but also in thesuperficial cortex indicates that the superficial cortical fiber cellsplay a much more active role in lens metabolism than previouslysuspected. Alternatively, the observed movement of TMRE across amitochondrial network could represent change in the distribution ofpotential across the inner membrane, presumably allowing energytransmission across the cell from regions of low to regions of high ATPdemand.

[学科分类] 生物化学/生物物理