Sodium-bicarbonate cotransporter NBCn1 in the kidney medullary thick ascending limb cell line is upregulated under acidic conditions and enhances ammonium transport

Exp Physiol. 2010 Sep;95(9):926-37. doi: 10.1113/expphysiol.2010.053967. Epub 2010 Jun 30.


In this study, we examined the effect of bicarbonate transporters on ammonium/ammonia uptake in the medullary thick ascending limb cell line ST-1. Cells were treated with 1 mm ouabain and 0.2 mM bumetanide to minimize carrier-mediated NH(4)(+) transport, and the intracellular accumulation of (14)C-methylammonium/methylammonia ((14)C-MA) was determined. In CO(2)/HCO(3)(-)-free solution, cells at normal pH briefly accumulated (14)C-MA over 7 min and reached a plateau. In CO(2)/HCO(3)(-) solution, however, cells markedly accumulated (14)C-MA over the experimental period of 30 min. This CO(2)/HCO(3)(-)-dependent accumulation was reduced by the bicarbonate transporter blocker, 4,4-diisothiocyanatostilbene-2,2-disulfonate (DIDS; 0.5 mM). Replacing Cl(-) with gluconate reduced the accumulation, but the reduction was more substantial in the presence of DIDS. Incubation of cells at pH 6.8 (adjusted with NaHCO(3) in 5% CO(2)) for 24 h lowered the mean steady-state intracellular pH to 6.96, significantly lower than 7.28 for control cells. The presence of DIDS reduced (14)C-MA accumulation in control conditions but had no effect after acidic incubation. Immunoblotting showed that NBCn1 was upregulated after acidic incubation and in NH(4)Cl-containing media. The Cl(-)-HCO(3)(-) exchanger AE2 was present, but its expression remained unaffected by acidic incubation. Expressed in Xenopus oocytes, NBCn1 increased carrier-mediated (14)C-MA transport, which was abolished by replacing Na(+). Two-electrode voltage clamp of oocytes exhibited negligible current after NH(4)Cl application. These results suggest that DIDS-sensitive HCO(3)(-) extrusion normally governs NH(4)(+)/NH(3) uptake in the medullary thick ascending limb cells. We propose that, in acidic conditions, DIDS-sensitive HCO(3)(-) extrusion is inactivated, while NBCn1 is upregulated to stimulate NH(4)(+) transport.

Publication types

  • Research Support, N.I.H., Extramural
  • Research Support, Non-U.S. Gov't

MeSH terms

  • 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / pharmacology
  • Ammonia / metabolism
  • Animals
  • Anion Transport Proteins / metabolism
  • Antiporters / metabolism
  • Bicarbonates / metabolism*
  • Biological Transport
  • Cell Line
  • Chlorides / metabolism
  • Gluconates / metabolism
  • Hydrogen-Ion Concentration
  • Kidney Medulla / drug effects
  • Kidney Medulla / metabolism*
  • Kinetics
  • Loop of Henle / drug effects
  • Loop of Henle / metabolism*
  • Membrane Potentials
  • Methylamines / metabolism
  • Patch-Clamp Techniques
  • Quaternary Ammonium Compounds / metabolism*
  • Rats
  • SLC4A Proteins
  • Sodium / metabolism*
  • Sodium-Bicarbonate Symporters / antagonists & inhibitors
  • Sodium-Bicarbonate Symporters / genetics
  • Sodium-Bicarbonate Symporters / metabolism*
  • Up-Regulation
  • Xenopus


  • Anion Transport Proteins
  • Antiporters
  • Bicarbonates
  • Chlorides
  • Gluconates
  • Methylamines
  • Quaternary Ammonium Compounds
  • SLC4A Proteins
  • Slc4a7 protein, rat
  • Sodium-Bicarbonate Symporters
  • Ammonia
  • Sodium
  • methylamine
  • 4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid
  • gluconic acid