Abstract
pH i recovery in acid-loaded Ehrlich ascites tumor cells and pH i maintenance at steady-state were studied using the fluorescent probe BCECF.
Both in nominally HCO −3 -free media and at 25 mm HCO −3 , the measured pH i (7.26 and 7.82, respectively) was significantly more alkaline than the pH i . value calculated assuming the transmembrane HCO −3 gradient to be equal to the Cl− gradient. Thus, pH i in these cells is not determined by the Cl− gradient and by Cl−/HCO −3 exchange.
pH i recovery following acid loading by propionate exposure, NH +4 withdrawal, or CO2 exposure is mediated by amiloride-sensitive Na+/H+ exchange in HCO3 − free media, and in the presence of HCO −3 (25 mm) by DIDS-sensitive, Na+-dependent Cl−/HCO −3 exchange. A significant residual pH i recovery in the presence of both amiloride and DIDS suggests an additional role for a primary active H+ pump in pH i regulation. pH i maintenance at steady-state involves both Na+/H+ exchange and Na+-dependent Cl−/HCO −3 exchange.
Acute removal of external Cl− induces a DIDS-sensitive, Na+-dependent alkalinization, taken to represent HCO −3 influx in exchange for cellular Cl−. Measurements of 36Cl− efflux into Cl−-free gluconate media with and without Na+ and/or HCO −3 (10 mm) directly demonstrate a DIDS-sensitive, Na+ dependent Cl−/HCO −3 exchange operating at slightly acidic pH i (pHo 6.8), and a DIDS-sensitive, Na+-independent Cl−/HCO −3 exchange operating at alkaline pH i (pH o 8.2).
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The excellent technical assistance of Marianne Schiødt and Birgit B. Jørgensen is gratefully acknowledged. The work was supported by the Carlsberg Foundation (B.K.) and by a grant from the Danish Natural Science Foundation (E.K.H. and L.O.S.).
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Kramhøft, B., Hoffmann, E.K. & Simonsen, L.O. pHi regulation in Ehrlich mouse ascites tumor cells: Role of sodium-dependent and sodium-independent chloride-bicarbonate exchange. J. Membarin Biol. 138, 121–132 (1994). https://doi.org/10.1007/BF00232640
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DOI: https://doi.org/10.1007/BF00232640