Activation of the Na+-K+ (NH4+)-2Cl--cotransporter from rat submandibular glands in response to VIP

N. Chaïb, E. Kabré, M. Métioui, E. Alzola, H. Amsallem, A. Marino, A. Moran, J. P. Dehaye

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Abstract

A cellular suspension from rat submandibular glands was prepared with collagenase. The intracellular pH (pH(i)) was estimated with 2',7'-bis-(2- carboxy-ethyl)-5(6)-carboxyfluorescein (BCECF). After exposure to NH4Cl, the pH(i) transiently increased (diffusion of NH3) and then dropped (influx of NH4+). Isoproterenol increased 2.5-fold the rate of NH4+ influx; bumetanide, an inhibitor of the Na+-K+-2Cl--cotransporter blocked the response to isoproterenol, confirming that the beta-adrenergic agonist stimulated the cotransporter. Forskolin (1 μmol/L) mimicked the response to isoproterenol. VIP (1 nmol/L-1 μmol/L) also increased the activity of the cotransporter. Cyclic AMP rather than calcium was the mediator of this activation since 1) carbachol which increased the [Ca2+](i) fivefold increased the uptake of NH4+ by only 50%; 2) only high concentrations of VIP significantly increased the [Ca2+](i);3) incubation in the presence of EGTA had no effect on the response to VIP; 4) low concentrations (nmol/L) of the neuropeptide increased the intracellular level of cAMP; and 5) the stimulation of the cotransporter by VIP, forskolin, and isoproterenol was inhibited by H8, an inhibitor of cAMP-dependent protein kinase. It is concluded that the Na+-K+-2Cl--cotransporter of rat submandibular glands is activated by isoproterenol, forskolin, and neuropeptides of the VIP family by a mechanism involving cAMP-dependent processes. The activation of the cotransporter by VIP could partly, explain the potentiating effect of VIP on the response to sialagogues like substance P or muscarinic agonists.

Original languageEnglish
Pages (from-to)1759-1770
Number of pages12
JournalPeptides
Volume19
Issue number10
DOIs
StatePublished - 1 Dec 1998

Keywords

  • Ammonium chloride
  • BCECF
  • Carbachol
  • Intracellular calcium
  • Salivary glands
  • Secretion

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