Basolateral Na(+)-independent Cl(-)-HCO3- exchange in primary cultures of rat IMCD cells.
The role of anion exchange in the regulation of intracellular pH (pHi) under base load and steady-state conditions was investigated in confluent monolayers of rat inner medullary collecting duct (IMCD) cells in primary culture using the pH-sensitive fluoroprobe 2,7-bis(carboxyethyl)-5(6')-carboxyfluorescein (BCECF). Recovery of pHi after imposition of a base load induced either by replacement of HCO3-/CO2 by N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid (HEPES) at the same extracellular pH (pHo) or deletion of Cl- from a HCO3-/CO2-buffered solution had an absolute requirement for Cl-, was Na+ independent, and was inhibited approximately 90% by 50 microM 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS). When pHo was decreased by lowering HCO3- concentration in the constant presence of 5% CO2, the rate of decrement in pHi was significantly blunted in the absence of Cl-. Imposition of a positive or negative diffusion potential of equal but opposite magnitude did not modify the anion exchange rate, confirming the electroneutrality of the process. Under steady-state conditions, pHi of cells bathed in a HCO3-/CO2-buffered solution was 7.33 +/- 0.06, significantly lower than that of cells bathed in a nominally HCO3-/CO2-free buffer (7.50 +/- 0.04), indicating that under physiological conditions the pathway functions as a base extruder. In studies performed on cells grown on permeable supports, the anion exchange pathway was found to be confined exclusively to the basolateral-equivalent cell surface. In summary, confluent monolayers of rat IMCD cells in primary culture possess a Na(+)-independent, DIDS-inhibitable electroneutral Cl(-)-HCO3- exchange pathway that is confined to the basolateral cell surface. The transporter is an important determinant of steady-state pHi and is the predominant mechanism whereby the cell recovers from imposed elevations in pHi.
Kraut
J A
JA
Medical Service, West Los Angeles Veterans Affairs Medical Center, California 90073.
Hart
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Nord
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D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001485:Basement Membrane / Q000378:metabolism; D001639:Bicarbonates / Q000494:pharmacology; D002245:Carbon Dioxide / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002478:Cells, Cultured; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000494:pharmacology; D006706:Homeostasis; D006863:Hydrogen-Ion Concentration; D007425:Intracellular Membranes / Q000378:metabolism; D007474:Ion Exchange; D007679:Kidney Medulla; D007685:Kidney Tubules, Collecting / Q000166:cytology / Q000378:metabolism*; D051381:Rats; D012964:Sodium / Q000494:pharmacology*
D001639:Bicarbonates; D002352:Carrier Proteins; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D002245:Carbon Dioxide; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D012964:Sodium; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid