Physiological response of bovine subcommissural organ to endothelin 1 and bradykinin |
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Authors: | S Schöniger T Caprile C R Yulis Q Zhang E M Rodríguez F Nürnberger |
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Institution: | (1) Department of Cellular Biology and Anatomy, Medical College of Georgia, Augusta, GA 30912, USA;(2) Department of Biochemistry and Molecular Biology, Medical College of Georgia, Augusta, Ga., USA;(3) Department of Ophthalmology, Medical College of Georgia, Augusta, Ga., USA |
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Abstract: | Elevated glutamate levels have been reported in humans with diabetic retinopathy. Retinal Müller glial cells regulate glutamate
levels via the GLAST transporter and system xc
− (cystine-glutamate exchanger). We have investigated whether transporter function and gene and/or protein expression are altered
in mouse Müller cells cultured under conditions of hyperglycemia or oxidative stress (two factors implicated in diabetic retinopathy).
Cells were subjected to hyperglycemic conditions (35 mM glucose) over an 8-day period or to oxidative stress conditions (induced
by exposure to various concentrations of xanthine:xanthine oxidase) for 6 h. The Na+-dependent and –independent uptake of 3H] glutamate was assessed as a measure of GLAST and system xc
− function, respectively. Hyperglycemia did not alter the uptake of 3H] glutamate by GLAST or system xc
−; neither gene nor protein expression decreased. Oxidative stress (70:14 or 100:20 μM xanthine:mU/ml xanthine oxidase) decreased
GLAST activity by ~10% but increased system xc
− activity by 43% and 89%, respectively. Kinetic analysis showed an oxidative-stress-induced change in Vmax, but not Km. Oxidative stress caused a 2.4-fold increase in mRNA encoding xCT, the unique component of system xc
−. Of the two isoforms of xCT (40 and 50 kDa), oxidative stress induced a 3.6-fold increase in the 40-kDa form localized to
the plasma membrane. This is the first report of the differential expression and localization of xCT isoforms as caused by
cellular stress. Increased system xc
− activity in Müller cells subjected to conditions associated with diabetic retinopathy may be beneficial, as this exchanger
is important for the synthesis of the antioxidant glutathione.
This work was supported by NIH R01 EY014560. |
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Keywords: | Cystine-glutamate exchanger Cell culture Transporter kinetics Mouse (C57BL/6) |
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