Role of HCO3- in regulation of cytoplasmic pH in ciliary epithelial cells.
Cytoplasmic pH (pHi) was monitored using the pH-sensitive absorbance of 5(6)carboxy-4',5'-dimethylfluorescein in monolayers of a cell clone derived from bovine pigmented ciliary epithelium (PE) transformed with the simian virus 40. 1) Changing extracellular media from a nominally HCO3(-)-free solution to a solution containing 28 mM HCO3(-)-5% CO2 at constant extracellular pH (7.4) resulted in a delayed alkalinization of pHi, which was 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) sensitive and was inhibited in Na+-free medium and in Cl(-)-depleted cells. 2) DIDS pretreatment acidified pHi in HCO3(-)-containing media. 3) Replacing extracellular Cl- resulted in a DIDS-sensitive, HCO3(-)-dependent, and Na+-independent alkalinization. 4) Replacing extracellular Na+ in HCO3(-)-containing media led to a partly DIDS-sensitive intracellular acidification. 5) Recovery of pHi after an alkali load (acetate prepulse) had a HCO3(-)-dependent and DIDS-sensitive component. 6) Two Na+-dependent components participated in pHi regulation after an acid load (NH4+ prepulse) in HCO3(-)-containing solution. One was amiloride sensitive, the other was DIDS sensitive and was inhibited in HCO3(-)-free media and after Cl- depletion. We conclude that in cultured PE, in addition to Na+-H+ exchange, two HCO3-transporters participate in pHi regulation. Cl(-)-dependent Na+-HCO3-symport regulates pHi during steady state and after an acid load, and Na+-independent Cl(-)-HCO3-exchange is involved in pHi recovery after an alkali load.
Helbig
H
H
Institut für Klinische Physiologie, Klinikum Steglitz der Freien Universität Berlin, Federal Republic of Germany.
Korbmacher
C
C
Stumpff
F
F
Coca-Prados
M
M
Wiederholt
M
M
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000494:pharmacology; D000584:Amiloride / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D002712:Chlorides / Q000494:pharmacology; D002924:Ciliary Body / Q000502:physiology*; D003593:Cytoplasm / Q000187:drug effects / Q000502:physiology; D004848:Epithelium / Q000187:drug effects / Q000502:physiology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D012964:Sodium / Q000494:pharmacology
D001639:Bicarbonates; D002712:Chlorides; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D000584:Amiloride; D012964:Sodium; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid