January 1998
Volume 39, Issue 1
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Articles  |   January 1998
Sodium, potassium, two chloride cotransport in corneal endothelium: characterization and possible role in volume regulation and fluid transport.
Author Affiliations
  • F P Diecke
    Department of Physiology, College of Physicians and Surgeons, Columbia University, New York, New York, USA.
  • Z Zhu
    Department of Physiology, College of Physicians and Surgeons, Columbia University, New York, New York, USA.
  • F Kang
    Department of Physiology, College of Physicians and Surgeons, Columbia University, New York, New York, USA.
  • K Kuang
    Department of Physiology, College of Physicians and Surgeons, Columbia University, New York, New York, USA.
  • J Fischbarg
    Department of Physiology, College of Physicians and Surgeons, Columbia University, New York, New York, USA.
Investigative Ophthalmology & Visual Science January 1998, Vol.39, 104-110. doi:
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      F P Diecke, Z Zhu, F Kang, K Kuang, J Fischbarg; Sodium, potassium, two chloride cotransport in corneal endothelium: characterization and possible role in volume regulation and fluid transport.. Invest. Ophthalmol. Vis. Sci. 1998;39(1):104-110.

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Abstract

PURPOSE: To search for membrane transporter proteins that could contribute to volume regulation and fluid transport by corneal endothelium. As an initial step, the authors have focused on Na+-K+-2Cl- cotransporters. METHODS: Bovine corneal endothelial cells were cultured to confluence. 86Rubidium was used as a tracer for K+ uptake determinations; uptake values were normalized per milligram of cell protein. RESULTS: Three components of K+ uptake were characterized: ouabain (1 mM) sensitive, bumetanide (0.1 mM) sensitive, and ouabain-bumetanide insensitive. Both the ouabain-sensitive and bumetanide-sensitive components increased in the presence of 26.2 mM HCO3-; 0.5 mM 4,4'-diisothiocyanato-stilbene-2,2'-disulfonic acid abolished this increase. The bumetanide-sensitive component was completely inhibited in the absence of Na+ or Cl-. This component was increased 33% by a 33% hypertonic solution and was decreased 38% by a 33% hypotonic solution. The protein kinase C activator phorbol 12-myristate 13-acetate decreased the activity of the cotransporter, whereas forskolin, in the presence of isobutylmethylxanthine, decreased it. Calyculin A (100 nM), an inhibitor of phosphatases 1 and 2a, produced a large (97%) activation of this component. CONCLUSIONS: These results provided for the first time conclusive evidence for the presence of a Na+-K+-2Cl- cotransporter in corneal endothelium and of its possible involvement in volume-regulatory processes in these cells. Given the uptake values reported here, such cotransporter could contribute significantly to electrolyte transport and hence to fluid transport across this preparation.

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