Hyperosmolality inhibits bicarbonate absorption in rat medullary thick ascending limb via a protein-tyrosine kinase-dependent pathway

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In the rat medullary thick ascending limb (MTAL), hyperosmolality inhibits transepithelial HCO3/- absorption (JHCO3/-) by inhibiting apical membrane Na+/H+ exchange. To examine signaling mechanisms involved in this regulatory response, MTALs were isolated and perfused in vitro with 25 mH HCO3/- solutions (290 mosmol/kg H2O). Osmolality was increased in lumen and bath solutions by addition of 300 mM mannitol or 75 mM NaCl. Addition of mannitol reduced JHCO3/- by 60% and addition of NaCl reduced JHCO3/- by 50%. With the protein tyrosine kinase (PTK) inhibitor genistein (7 μM) or herbimycin A (1 μM) in the bath, addition of mannitol reduced JHCO3/- only by 11% and addition of NaCl reduced JHCO3/- only by 15%. Staurosporine (10-7 M) or forskolin (10-6 M) in the bath had no effect on inhibition of JHCO3/- by hypertonic NaCl. Genistein had no effect on inhibition of JHCO3/- by vasopressin (a cyclic AMP-dependent process) or stimulation of JHCO3/- by prostaglandin E2 (a protein kinase C-dependent process). Under isosmotic conditions, addition of genistein or herbimycin A to the bath increased JHCO3/- by 30% through stimulation of apical membrane Na+/H+ exchange. Addition of the tyrosine phosphatase inhibitor molybdate (50 μM) to the bath reproduced the inhibition of JHCO3/- observed with hyperosmolality. These data indicate that 1) the effect of hyperosmolality to inhibit MTAL HCO3/- absorption through inhibition of apical membrane Na+/H+ exchange is mediated via a PTK-dependent pathway that functions independent of regulation by cyclic AMP and protein kinase C, and 2) a constitutive PTK activity inhibits apical membrane Na+/H+ exchange and HCO3/absorption under isosmotic conditions. Our results suggest that tyrosine phosphorylation is a critical step in inhibition of the apical Na+/H+ exchanger isoform NHE-3 by hyperosmolality.

Original languageEnglish (US)
Pages (from-to)9883-9889
Number of pages7
JournalJournal of Biological Chemistry
Issue number17
StatePublished - Jan 1 1995


ASJC Scopus subject areas

  • Biochemistry
  • Molecular Biology
  • Cell Biology

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