Underexpression of the Na+-dependent neutral amino acid transporter ASCT2 in the spontaneously hypertensive rat kidney.
This study examined the inward transport of l-[(14)C]alanine, an ASCT2 preferential substrate, in monolayers of immortalized renal proximal tubular epithelial (PTE) cells from Wistar-Kyoto (WKY) and spontaneously hypertensive (SHR) rats. The expression of ASCT2 in WKY and SHR PTE cells and kidney cortices from WKY and SHR was also evaluated. l-[(14)C]alanine uptake was highly dependent on extracellular Na(+). Replacement of NaCl by LiCl or choline chloride abolished transport activity in SHR and WKY PTE cells. In the presence of the system L inhibitor BCH, Na(+)-dependent l-alanine uptake in WKY and SHR PTE cells was inhibited by alanine, serine, and cysteine, which is consistent with amino acid transport through ASCT2. The saturable component of Na(+)-dependent l-alanine transport under V(max) conditions in SHR PTE cells was one-half of that in WKY PTE cells, with similar K(m) values. Differences in magnitude of Na(+)-dependent l-alanine uptake through ASCT2 between WKY and SHR PTE cells correlated positively with differences in ASCT2 protein expression, this being more abundant in WKY PTE cells. Abundance of ASCT2 transcript and protein in kidney cortices of SHR rats was also lower than that in normotensive WKY rats. In conclusion, immortalized SHR and WKY PTE cells take up l-alanine mainly through a high-affinity Na(+)-dependent amino acid transporter, with functional features of ASCT2 transport. The activity and expression of the ASCT2 transporter were considerably lower in the SHR cells.
Pinho
Maria João
MJ
Institute of Pharmacology and Therapeutics, Faculty of Medicine, 4200 Porto, Portugal.
Pinto
Vanda
V
Serrão
Maria Paula
MP
Jose
Pedro A
PA
Soares-da-Silva
Patrício
P
D000409:Alanine / Q000378:metabolism; D026962:Amino Acid Transport System ASC / Q000096:biosynthesis*; D000596:Amino Acids / Q000096:biosynthesis / Q000378:metabolism / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D015153:Blotting, Western; D002460:Cell Line; D002794:Choline / Q000494:pharmacology; D004847:Epithelial Cells / Q000187:drug effects / Q000378:metabolism; D007672:Kidney Cortex / Q000166:cytology / Q000187:drug effects / Q000378:metabolism; D007700:Kinetics; D018021:Lithium Chloride / Q000494:pharmacology; D015778:Minor Histocompatibility Antigens; D011189:Potassium Chloride / Q000494:pharmacology; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D011918:Rats, Inbred SHR; D011921:Rats, Inbred WKY; D020133:Reverse Transcriptase Polymerase Chain Reaction
D026962:Amino Acid Transport System ASC; D000596:Amino Acids; D015778:Minor Histocompatibility Antigens; D012333:RNA, Messenger; C498849:Slc1a5 protein, rat; D000643:Ammonium Chloride; D011189:Potassium Chloride; D018021:Lithium Chloride; D002794:Choline; D000409:Alanine