Localization and regulation of the rat renal na+-k+-2cl-
cotransporter, bsc-1.
Ecelbarger, Carolyn A., James Terris, John R. Hoyer, S[stod]oren
Nielsen, J. B. Wade, and Mark A. Knepper.
Laboratory of Kidney and Electrolyte Metabolism, National Heart,
Lung and Blood Institute, National Institutes of Health, Bethesda,
MD; Department of Physiology, Uniformed Services University of the
Health Sciences, Bethesda, MD; Dept. of Pediatrics, Children's
Hospital of Philadelphia, University of Pennsylvania School of
Medicine, Philadelphia, PA; Dept. of Cell Biology, Institute of
Anatomy, University of Aarhus, DK-8000 Aarhus, Denmark; and Dept. of
Physiology, University of Maryland School of Medicine, Baltimore,
MD
APStracts 3:0094F, 1996.
To investigate the role of the thick ascending limb (TAL) Na+-K+-2Cl-
cotransporter in regulation of water excretion, we have prepared a
peptide-derived polyclonal antibody based on the cloned cDNA sequence
of the rat type-1 bumetanide-sensitive cotransporter, BSC-1 (also
termed "NKCC-2"). Immunoblots revealed a single broad 161 kDa
band in membrane fractions of rat renal outer medulla and cortex, but
not from rat colon or parotid gland. A similar protein was labeled in
mouse kidney. Immunoperoxidase immunohistochemistry in rat kidney
revealed labeling restricted to the medullary and cortical TALs.
Because long-term regulation of urinary concentrating ability may
depend on regulation of Na+-K+-2Cl- cotransporter abundance, we used
immunoblotting to evaluate the effects of several in vivo factors on
expression levels of BSC-1 protein in rat kidney outer medulla.
Chronic oral saline loading with 0.16 M NaCl markedly increased BSC-1
abundance. However, long-term vasopressin infusion or thirsting of
rats did not affect BSC-1 abundance. Chronic furosemide infusion
caused a 9 kDa upward shift in apparent molecular weight and an
apparent increase in expression level. These results support the
previous identification of BSC-1 as the TAL Na+-K+-2Cl- transporter
and demonstrate that the expression of this transporter is regulated.
Received 16 January 1996; accepted in final form 9 May 1996.
APS Manuscript Number F10-6.
Article publication pending Am. J. Physiol. (Renal Fluid Electrolyte
Physiology).
ISSN 1080-4757 Copyright 1996 The American Physiological Society.
Published in APStracts on 28 May 96