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  • Title: Na+-independent nucleoside transporters regulate adenosine and hypoxanthine levels in Müller cells and the inner blood-retinal barrier.
    Author: Akanuma S, Soutome T, Hisada E, Tachikawa M, Kubo Y, Hosoya K.
    Journal: Invest Ophthalmol Vis Sci; 2013 Feb 27; 54(2):1469-77. PubMed ID: 23361509.
    Abstract:
    PURPOSE: To elucidate the mechanism(s) of hypoxanthine production in Müller cells and the elimination of hypoxanthine across the inner blood-retinal barrier (BRB). METHODS: The hypoxanthine biosynthesis and adenosine transport in Müller cells were investigated using a conditionally immortalized rat Müller cell line, TR-MUL5 cells. The elimination of hypoxanthine across the inner BRB was assessed by an in vivo microdialysis method and an in vitro transport study using a conditionally immortalized rat retinal capillary endothelial cell line, TR-iBRB2 cells. RESULTS: [(3)H]Hypoxanthine was detected in TR-MUL5 cells and TR-MUL5 cell-cultured medium 3 hours after [(3)H]adenosine incubation, indicating that the hypoxanthine is produced in TR-MUL5 cells. [(3)H]Adenosine was taken up into TR-MUL5 cells, which express mRNAs of nucleoside transporters (ENT1-2 and CNT1-2), in an Na(+)-independent and concentration-dependent manner (Km = 20 μM). Moreover, 100 μM nitrobenzylmercaptopurine riboside (NBMPR) and azidothymidine, which are inhibitors of ENT2, inhibited [(3)H]adenosine uptake, suggesting that ENT2 is a major contributor to adenosine transport in Müller cells. [(3)H]Hypoxanthine was eliminated from the rat vitreous humor and this process was inhibited in the presence of NBMPR. [(3)H]Hypoxanthine uptake by TR-iBRB2 cells took place in an Na(+)-independent and concentration-dependent manner with Km values of 4.3 μM and 2.9 mM, and was inhibited by 100 μM NBMPR. CONCLUSIONS: Our findings suggest that hypoxanthine is produced from adenosine in Müller cells and ENT2 plays a major role in adenosine uptake in Müller cells. Hypoxanthine in the retina is eliminated via Na(+)-independent equilibrative nucleoside transporters.
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