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Investigative Ophthalmology & Visual Science, Vol 37, 2269-2275, Copyright © 1996 by Association for Research in Vision and Ophthalmology

ARTICLES AND REPORTS

Identification of a novel, sodium-dependent, reduced glutathione transporter in the rat lens epithelium

R Kannan, JR Yi, D Tang, BV Zlokovic and N Kaplowitz
Division of Gastrointestinal and Liver Diseases, University of Southern California School of Medicine, Los Angeles 90033, USA.

PURPOSE: To determine whether glutathione (GSH) transporter(s) other than the previously identified rat canalicular GSH transporter (RcGshT) is present in the lens. METHODS: Poly (A) +RNA isolated from rat and guinea pig lens cortex and epithelium was injected into Xenopus laevis oocytes. The effect of sodium removal was determined by measuring cell- associated radioactivity in lenticular epithelium or cortex mRNA injected oocytes (pretreated with acivicin to inhibit gamma glutamyltranspeptidase) after 1 hour of incubation in NaCl medium or choline chloride (Na(+)-free) medium containing tracer GSH (plus unlabeled GSH). The effect of 2 mM bromosulfophthalein-GSH (BSP-GSH) on GSH uptake in the lens epithelium and cortex in NaCl medium at two GSH concentrations also was determined. The molecular form of uptake of GSH in lens epithelial mRNA-injected oocytes was examined by high- performance liquid chromatography. Western blot analysis was performed to study the presence of RcGshT in the cortex and epithelium. RESULTS: Oocytes injected with mRNA from rat and guinea pig lens epithelium and cortex compartments expressed GSH transport. High-performance liquid chromatography confirmed that epithelial uptake was as intact GSH under conditions of inhibition of GSH synthesis with dl-buthionine sulfoximine. The mean GSH uptake (nmol/oocyte per hour) in epithelial mRNA-injected oocytes was significantly reduced (P < 0.01, n = 4 oocyte preparations) under Na(+)-free conditions compared to NaCl medium at 0.05 mM and 2 mM GSH in the medium. Uptake in cortical mRNA-injected oocytes was unaffected by Na+ removal. Lens epithelial uptake exhibited a strong inhibition by BSP-GSH at 0.05 mM (55%) and 2 mM (64%), whereas cortical uptake was unaffected by BSP-GSH. Western blot analysis identified RcGshT in the cortical and epithelial regions. CONCLUSIONS: Results from the current study provide strong evidence for the presence of a hitherto unreported Na(+)-dependent, BSP-GSH inhibitable GSH transporter in the lens epithelium, which may mediate concentrative, basolateral uptake of aqueous GSH consistent with in situ eye perfusion studies. The Na(+)-independent, BSP-GSH insensitive RcGshT may function as an apical GSH effluxer in lens epithelium and in mediating concentration gradient driven inward GSH movement by uptake-efflux in the lens cortex.


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H. J. Gukasyan, V. H. L. Lee, K.-J. Kim, and R. Kannan
Net Glutathione Secretion across Primary Cultured Rabbit Conjunctival Epithelial Cell Layers
Invest. Ophthalmol. Vis. Sci., April 1, 2002; 43(4): 1154 - 1161.
[Abstract] [Full Text] [PDF]


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