Na(+)-dependent and -independent amino acid transport systems in immortalized human kidney epithelial cells derived from the proximal tubule.
In the proximal tubule Na(+)-dependent (SDAT) and Na(+)-independent (SIAT) amino acid (AA) transporters are present. The effects of neutral, basic, and acidic AA on membrane voltage (Vm) of immortalized human kidney epithelial (IHKE-1) cells derived from the proximal tubule were examined using the slow whole-cell patch-clamp technique. In the presence of Na+ AA depolarized Vm in a concentration-dependent manner (0.05-5 mM) with Asp = Arg = Glu = 2Cys < Pro = Leu < Phe = AIB = Ala = Pro = Asn < Gly. In the absence of extracellular Na+ a decreased depolarization was seen with most neutral AA (Ala, Pro, Asn, Gly, Phe, and Leu), and the depolarization was increased with Asp, Glu, Arg, and 2Cys (1 mM each). In the absence of Na+ and a reduction in Cl- (5 mM) the depolarization by Arg was reduced. Unlike that predicted for transport by system b0,+ which exchanges neutral against dibasic amino acids, Leu does not hyperpolarize but depolarize Vm of IHKE-1 cells in the absence of extracellular Na+. After removal of Na+ (0 mM) and a reduction in Cl- (5 mM) in the extracellular solution, Leu or Glu hyperpolarized Vm, indicating that IHKE-1 cells possess two different SIAT systems, one Cl(-)-dependent and similar to system b0,+ and one novel Cl(-)-dependent system, which might be a Cl-/AA exchanger and can be blocked by the Cl(-)-channel blockers 5-nitro-2-(3-phenylpropylamino)-benzoate (10 microM) and 4,4'-diisothiocyanostibene-2,2'-disulfonic acid (50 microM). B system-related AA transporters might be responsible for the C(-)-independent SIAT, since we were able to detect its signal by Northern blot analysis.
Hirsch
J R
JR
Medizinische Poliklinik, Westfälische Wilhelms-Universität Münster, Germany. hirschi@uni-muensta.de
Gonska
T
T
Waldegger
S
S
Lang
F
F
Schlatter
E
E
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000345:Affinity Labels / Q000494:pharmacology; D000596:Amino Acids / Q000378:metabolism*; D001692:Biological Transport; D015152:Blotting, Northern; D002478:Cells, Cultured; D018118:Chloride Channels / Q000037:antagonists & inhibitors / Q000378:metabolism; D004848:Epithelium / Q000378:metabolism; D006801:Humans; D007668:Kidney / Q000196:embryology / Q000378:metabolism*; D007687:Kidney Tubules, Proximal / Q000196:embryology / Q000378:metabolism*; D008564:Membrane Potentials; D009579:Nitrobenzoates / Q000494:pharmacology; D018408:Patch-Clamp Techniques; D012313:RNA / Q000032:analysis; D012964:Sodium / Q000378:metabolism*
D000345:Affinity Labels; D000596:Amino Acids; D018118:Chloride Channels; D009579:Nitrobenzoates; C058176:5-nitro-2-(3-phenylpropylamino)benzoic acid; C018564:dihydro-DIDS; D012313:RNA; D012964:Sodium; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid