pH regulation in HT29 colon carcinoma cells.
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 HCO3(-)-containing and 7.21 +/- 0.09 (n = 12) in HCO3(-)-free solution. HOE-694 (10 mumol/l), a potent inhibitor of the Na+/H+ exchanger, did not affect control pHi. As a means to acidify cells we used the NH4+/NH3 (20 mmol/l) prepulse technique. The mean peak acidification was 0.37 +/- 0.07 pH units (n = 6). In HCO3(-)-free solutions recovery from acid load was completely blocked by HOE-694 (1 mumol/l), whereas in HCO3(-)-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 HCO3(-)-containing and HCO3(-)-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 HCO3(-)-containing than in HCO3(-)-free solutions. The present observations demonstrate three distinct mechanisms of pHi regulation in HT29 cells: (a) a Na+/H+ exchanger, (b) a HCO3-/Cl- exchanger and (c) a Na(+)-dependent HCO3- transporter, probably the Na(+)-HCO3-/Cl- antiporter.(ABSTRACT TRUNCATED AT 250 WORDS)
Physiologisches Institut, Albert-Ludwigs-Universität, Freiburg, Germany.
M
M
Köttgen
J
J
Leipziger
K G
KG
Fischer
R
R
Nitschke
R
R
Greger
D000136:Acid-Base Equilibrium; D017920:Antiporters / Q000378:metabolism; D001639:Bicarbonates / Q000378:metabolism*; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000378:metabolism*; D003110:Colonic Neoplasms / Q000378:metabolism*; D005453:Fluorescence; D006146:Guanidines / Q000494:pharmacology; D006801:Humans; D006863:Hydrogen-Ion Concentration; D007413:Intestinal Mucosa / Q000378:metabolism*; D017136:Ion Transport; D012964:Sodium / Q000378:metabolism; D017923:Sodium-Hydrogen Exchangers / Q000037:antagonists & inhibitors / Q000378:metabolism*; D013450:Sulfones / Q000494:pharmacology; D014407:Tumor Cells, Cultured
D017920:Antiporters; D001639:Bicarbonates; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D006146:Guanidines; D017923:Sodium-Hydrogen Exchangers; D013450:Sulfones; C074107:3-methylsulfonyl-4-piperidinobenzoyl guanidine; D012964:Sodium