title: Involvement of system A in the retina-to-blood transport of l-proline across the inner blood-retinal barrier.
abstract: The purpose of the present study was to elucidate the mechanisms of retina-to-blood transport of l-proline across the blood-retinal barrier (BRB) in vivo and in vitro, and to identify the responsible transporter(s). The vitreous humor/retina-to-blood transport of [(3)H]l-proline across the BRB was evaluated by microdialysis. Transport mechanisms of [(3)H]l-proline were investigated by cellular uptake using an in vitro model of the inner BRB (TR-iBRB2 cells). The mRNA level of system A was determined by quantitative real-time PCR analysis with specific primers. [(3)H]l-Proline and [(14)C]d-mannitol, which is a bulk flow marker, were bi-exponentially eliminated from the vitreous humor after vitreous bolus injection. The elimination rate constant of [(3)H]l-proline during the terminal phase was 1.6-fold greater than that of [(14)C]d-mannitol. The terminal elimination rate constant difference between [(3)H]l-proline and [(14)C]d-mannitol was reduced in the retinal presence of 3 mM l-proline and 5 mM alpha-methylaminoisobutyric acid, suggesting that l-proline is transported via a carrier-mediated retina-to-blood transport process across the BRB. [(3)H]l-Proline uptake by TR-iBRB2 cells appeared to be mediated through a saturable and Na(+)-dependent process. The corresponding Michaelis-Menten constant was 392 muM. This process was reduced by substrates for system A, suggesting that system A is involved in l-proline uptake. Of the isoforms of system A, ATA1, ATA2, and ATA3, ATA2 mRNA is predominantly expressed in TR-iBRB2 cells and isolated rat retinal endothelial cells. In conclusion, system A, most likely ATA2, is responsible for the retina-to-blood transport of l-proline across the inner BRB and may play a role in maintaining the concentration of small neutral amino acids in the retina.
authors.affiliation: Department of Pharmaceutics, Graduate School of Medicine and Pharmaceutical Sciences, University of Toyama, 2630 Sugitani, Toyama 930-0194, Japan.
authors.forename: Daisuke
authors.initials: D
authors.lastname: Yoneyama
authors.forename: Yumiko
authors.initials: Y
authors.lastname: Shinozaki
authors.forename: Wan-Liang
authors.initials: WL
authors.lastname: Lu
authors.forename: Masatoshi
authors.initials: M
authors.lastname: Tomi
authors.forename: Masanori
authors.initials: M
authors.lastname: Tachikawa
authors.forename: Ken-Ichi
authors.initials: K
authors.lastname: Hosoya
mesh_terms: D026924:Amino Acid Transport System A / Q000235:genetics / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport; D001769:Blood / Q000378:metabolism*; D001813:Blood-Retinal Barrier / Q000378:metabolism*; D002196:Capillaries; D018929:Cell Culture Techniques; D004730:Endothelium, Vascular / Q000378:metabolism*; D008297:Male; D017551:Microdialysis; D011392:Proline / Q000378:metabolism*; D012333:RNA, Messenger / Q000378:metabolism; D051381:Rats; D055647:Rats, Transgenic; D017208:Rats, Wistar; D012160:Retina / Q000378:metabolism*; D012171:Retinal Vessels / Q000166:cytology; D020133:Reverse Transcriptase Polymerase Chain Reaction; D013997:Time Factors; D014822:Vitreous Body / Q000378:metabolism
chemical_list: D026924:Amino Acid Transport System A; D012333:RNA, Messenger; D011392:Proline