Osmotic activation of a Na(+)-dependent Cl-/HCO3- exchanger.
In many systems, osmotically induced cell shrinkage activates the Na+/H+ exchanger. To assess the role of H(+)-extruding transporters in the response to osmotic shrinkage in vascular smooth muscle (VSM) and Chinese hamster ovary (CHO) cells, intracellular pH (pHi) was measured with 2',7'-bis(carboxy-ethyl)-5(6)- carboxyfluorescein-acetoxymethyl ester (BCECF-AM) after exposing cells to hypertonic medium. In nominally HCO(3-)-free medium, addition of 200 mM sucrose caused pHi to increase 0.33 pH unit on average in VSM cells but only 0.13 pH unit in CHO cells. Permeant solutes failed to increase pHi significantly. Cytochalasin B (1-20 microM), colchicine (1-10 microM), Ca2+ removal, and downregulation of protein kinase C activity did not affect osmotic activation of H+ extrusion in either cell type. Additional work was carried out to determine why osmotic activation of H+ extrusion was less in CHO than in VSM cells. In CHO cells, the osmotically induced delta pHi was only weakly sensitive to amiloride, suggesting that osmotic forces may activate an H+ transport system other than Na+/H+ exchange. In the presence of 10 mM HCO3-, osmotically induced delta pHi decreased by 60% in VSM cells but increased by 50% in CHO cells compared with the delta pHi in HCO(3-)-free medium. Lastly, removal of extracellular Cl- did not affect osmotically induced delta pHi in VSM cells but completely abolished the response in CHO cells. We conclude that in VSM cells osmotically induced changes in pHi are mediated by Na+/H+ exchange, whereas in CHO cells they are most likely mediated by a Na(+)-dependent Cl-/HCO3- exchanger.
Division of Nephrology, University of California, San Francisco 94143.
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D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D017920:Antiporters / Q000378:metabolism*; D001639:Bicarbonates / Q000494:pharmacology; D016466:CHO Cells; D002478:Cells, Cultured; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000494:pharmacology; D006224:Cricetinae; D006863:Hydrogen-Ion Concentration; D006982:Hypertonic Solutions / Q000494:pharmacology; D009131:Muscle, Smooth, Vascular / Q000166:cytology / Q000378:metabolism; D009995:Osmosis; D051381:Rats; D012964:Sodium / Q000494:pharmacology*
D017920:Antiporters; D001639:Bicarbonates; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D006982:Hypertonic Solutions; D000584:Amiloride; D012964:Sodium