Alternatively spliced isoform of apical Na(+)-K(+)-Cl(-) cotransporter gene encodes a furosemide-sensitive Na(+)-Cl(-)cotransporter

Am J Physiol Renal Physiol. 2001 Apr;280(4):F574-82. doi: 10.1152/ajprenal.2001.280.4.F574.


In the absence of vasopressin, medullary thick ascending limb cells express a K(+)-independent, furosemide-sensitive Na(+)-Cl(-) cotransporter that is inhibited by hypertonicity. The murine renal specific Na(+)-K(+)-2 Cl(-) cotransporter gene (SLC12A1) gives rise to six alternatively spliced isoforms. Three feature a long COOH-terminal domain that encodes the butmetanide-sensitive Na(+)-K(+)-2 Cl(-) cotransporter (BSC1-9/NKCC2), and three with a short COOH-terminal domain, known as mBSC1-A4, B4, or F4 (19). Here we have determined the functional characteristics of mBSC1-A4, as expressed in Xenopus laevis oocytes. When incubated at normal oocyte osmolarity (approximately 200 mosmol/kgH(2)O), mBSC1-4-injected oocytes do not express significant Na(+) uptake over H(2)O-injected controls, and immunohistochemical analysis shows that the majority of mBSC1-4 protein is in the oocyte cytoplasm and not at the plasma membrane. In contrast, when mBSC1-4 oocytes are exposed to hypotonicity (approximately 100 mosmol/kgH(2)O), a significant increase in Na(+) uptake but not in (86)Rb(+) uptake is observed. The increased Na(+) uptake is Cl(-) dependent, furosemide sensitive, and cAMP sensitive but K(+) independent. Sodium uptake increases with decreasing osmolarity between 120 and 70 mosmol/kgH(2)O (r = 0.95, P < 0.01). Immunohistochemical analysis shows that in hypotonic conditions mBSC1-A4 protein is expressed in the plasma membrane. These studies indicate that the mBSC1-A4 isoform of the SLC12A1 gene encodes a hypotonically activated, cAMP- and furosemide-sensitive Na(+)-Cl(-) cotransporter. Thus it is possible that alternative splicing of the BSC1 gene could provide the molecular mechanism enabling the Na(+)-Cl(-)-to-Na(+)-K(+)-2Cl(-) switching in thick ascending limb cells.

Publication types

  • Research Support, Non-U.S. Gov't
  • Research Support, U.S. Gov't, P.H.S.

MeSH terms

  • 1-Methyl-3-isobutylxanthine / pharmacology
  • Alternative Splicing / physiology*
  • Animals
  • Biological Transport / drug effects
  • Biological Transport / physiology
  • Bumetanide / pharmacology
  • Carrier Proteins / chemistry
  • Carrier Proteins / genetics*
  • Carrier Proteins / metabolism*
  • Cell Membrane
  • Cyclic AMP / metabolism
  • Cyclic AMP-Dependent Protein Kinases / antagonists & inhibitors
  • Cyclic AMP-Dependent Protein Kinases / metabolism
  • Cytosol / metabolism
  • Diuretics / pharmacology*
  • Enzyme Inhibitors / pharmacology
  • Female
  • Furosemide / pharmacology*
  • Gene Expression / physiology
  • Hypotonic Solutions / pharmacology
  • Isomerism
  • Isoquinolines / pharmacology
  • Loop of Henle / enzymology
  • Mammals
  • Oocytes / physiology
  • Osmolar Concentration
  • Phosphodiesterase Inhibitors / pharmacology
  • Sodium Radioisotopes / pharmacokinetics
  • Sodium-Potassium-Chloride Symporters
  • Sulfonamides*
  • Tritium
  • Xenopus laevis


  • Carrier Proteins
  • Diuretics
  • Enzyme Inhibitors
  • Hypotonic Solutions
  • Isoquinolines
  • Phosphodiesterase Inhibitors
  • Sodium Radioisotopes
  • Sodium-Potassium-Chloride Symporters
  • Sulfonamides
  • Bumetanide
  • Tritium
  • Furosemide
  • Cyclic AMP
  • Cyclic AMP-Dependent Protein Kinases
  • N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide
  • 1-Methyl-3-isobutylxanthine