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Regulation of the Na + ‐K + (NH + 4 )‐2Cl − cotransporter of rat submandibular glands
Author(s) -
Chaïb N.,
Kabré E.,
Métioui M.,
Cabrita Franco M.,
Dehaye J.P.
Publication year - 1999
Publication title -
journal of cellular physiology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.529
H-Index - 174
eISSN - 1097-4652
pISSN - 0021-9541
DOI - 10.1002/(sici)1097-4652(199909)180:3<422::aid-jcp13>3.0.co;2-t
Subject(s) - submandibular gland , cotransporter , chemistry , medicine , endocrinology , microbiology and biotechnology , biochemistry , biology , sodium , organic chemistry
A cellular suspension from rat submandibular glands was exposed to different concentrations of NH 4 Cl, and the variations of the intracellular concentration of calcium ([Ca 2+ ] i ) and the intracellular pH (pH i ) were measured using fura‐2 and 2′,7′‐bis‐(2‐carboxy‐ethyl)‐5(6)‐carboxyfluorescein. More than 5 mmol/l NH 4 Cl significantly increased the [Ca 2+ ] i without affecting the response to 100 µmol/l carbachol. When exposed to 1 and 5 mmol/l NH 4 Cl, the cells acidified immediately. At 30 mmol/l, NH 4 Cl first alkalinized the cells and the pH i subsequently dropped. This drop reflects the uptake of NH + 4ions that dissociate to NH 3 and H + in the cytosol. These protons are exchanged for extracellular sodium by the Na + /H + exchanger because the presence of an inhibitor of the exchanger in the medium increased the acidification induced by 1 mmol/l NH 4 Cl. Ouabain partly blocked the uptake of NH + 4 . In the combined presence of ouabain and bumetanide (an inhibitor of the Na + ‐K + ‐2Cl − cotransporter), 1 mmol/l NH 4 Cl alkalinized the cells. The contribution of the Na/K ATPase and the Na + ‐K + ‐2Cl − cotransporter in the uptake of NH + 4was independent of the presence of calcium in the medium. Isoproterenol increased the uptake of NH + 4by the cotransporter. Conversely, 1 mmol/l extracellular ATP blocked the basal uptake of NH + 4by the cotransporter. This inhibition was reversed by extracellular magnesium or Coomassie Blue. It was mimicked by benzoyl‐ATP but not by CTP, GTP, UTP, ADP, or ADPβS. ATP only slightly inhibited the increase of cyclic AMP (−22%) by isoproterenol but fully blocked the stimulation of the cotransporter by the β‐adrenergic agonist. ATP increased the release of 3 H‐arachidonic acid from prelabeled cells but SK&F 96365, an imidazole‐based cytochrome P450 inhibitor, did not affect the inhibition by ATP. It is concluded that the activation of a purinoceptor inhibits the basal and the cyclic AMP‐stimulated activity of the Na + ‐K + ‐2Cl − cotransporter. J. Cell. Physiol. 180:422–430, 1999. © 1999 Wiley‐Liss, Inc.