ISSN:
1432-2013
Keywords:
BCECF
;
Na+/H+ exchanger
;
HCO 3 − /Cl− exchanger
;
Na+-dependent HCO 3 − transporter
;
DIDS
;
HOE-694
Source:
Springer Online Journal Archives 1860-2000
Topics:
Medicine
Notes:
Abstract The pH regulation in HT29 colon carcinoma cells has been investigated using the pH-sensitive fluorescent indicator 2′,7′-biscarboxyethyl-5(6)-carboxyfluorescein (BCECF). Under control conditions, intracellular pH (pHi) was 7.21±0.07 (n=22) in HCO 3 − -containing and 7.21±0.09 (n=12) in HCO 3 − -free solution. HOE-694 (10 μmol/l), a potent inhibitor of the Na+/H+ exchanger, did not affect control pHi. As a means to acidify cells we used the NH 4 + /NH3 (20 mmol/l) prepulse technique. The mean peak acidification was 0.37±0.07 pH units (n=6). In HCC 3 − -free solutions recovery from acid load was completely blocked by HOE-694 (1 μmol/l), whereas in HCO3 3 − -containing solutions a combination of HOE-694 and 4,4′-diisothiocyanatostilbene-2, 2′-disulphonate (DIDS, 0.5 mmol/l) was necessary to show the same effect. Recovery from acid load was Na+-dependent in HCO 3 − -containing and HCO 3 − -free solutions. Removal of external Cl− caused a rapid, DIDS-blockable alkalinization of 0.33±0.03 pH units (n=15) and of 0.20±0.006 pH units (n=5), when external Na+ was removed together with Cl−. This alkalinization was faster in HCO 3 − -containing than in HCO 3 − -free solutions. The present observations demonstrate three distinct mechanisms of pH regulation in HT29 cells: (a) a Na+/H+ exchanger, (b) a HCO 3 − /Cl− exchanger and (c) a Na+-dependent HCC 3 − transporter, probably the Na+-HCO 3 − /Cl− antiporter. Under HCO 3 − — free conditions the Na+/H+ exchanger fully accounts for recovery from acid load, whereas in HCO 3 − -containing solutions this is accomplished by the Na+/H+ exchanger and a Na+-dependent mechanism, which imports HCO 3 − . Recovery from alkaline load is caused by the HCO 3 − /Cl− exchanger.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00374856
