Journal
INVESTIGATIVE OPHTHALMOLOGY & VISUAL SCIENCE
Volume 46, Issue 7, Pages 2522-2530Publisher
ASSOC RESEARCH VISION OPHTHALMOLOGY INC
DOI: 10.1167/iovs.04-1175
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PURPOSE. L-type amino acid transporters (LATs) prefer branched-chain and aromatic amino acids, including neurotransmitter precursors. The objective of this study was to clarify the expression and function of LAT at the inner blood retinal barrier (BRB). METHODS. [H-3]L-Leucine transport at the inner BRB was characterized by using in vivo integration plot analysis and a conditionally immortalized rat retinal capillary endothelial cell line (TR-iBRB2). The expression of the LAT1 was demonstrated by quantitative real-time RT-PCR, immunoblot, and immunohistochemical analyses. RESULTS. The apparent influx permeability clearance of [H-3]L-leucine in the rat retina was found to be 203 mu L/(min . g retina), supporting a carrier-mediated influx transport of L-leucine at the BRB. [H-3]L-Leucine uptake by TR-iBRB2 cells was an Na+-independent and concentration-dependent process with a K-m of 14.1 mu M. This process was more potently cis inhibited by substrates of LAT1, D-leucine, D-phenylalanine, and D-methionine, than those of LAT2, L-alanine, and L-glutamine. [H-3]L-Leucine efflux from TR-iBRB2 cells was trans-stimulated by substrates of LAT1. The expression of LAT1 mRNA was 100- and 15-fold greater than that of LAT2 in TR-iBRB2 and magnetically isolated rat retinal vascular endothelial cells, respectively. The expression of LAT1 protein was observed in TR-iBRB2 and primary cultured human retinal endothelial cells and immuno-staining of LAT1 was observed along the rat retinal capillaries. CONCLUSIONS. LAT1 is expressed at the inner BRB and mediates blood-to-retina L-leucine transport. This transport system plays a key role in maintaining large neutral amino acids as well as neurotransmitters in the neural retina.
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