Regulation of Cl/HCO3 exchange in gastric parietal cells.
Microspectrofluorimetry of the fluorescent indicators 2',7'-bis-(2-carboxyethyl)-5(and-6)carboxyfluorescein and 6-methoxy-N-(3-sulfopropyl)-quinolinium was used to measure intracellular pH (pHi), intracellular Cl (Cli), and transmembrane fluxes of HCO3 and Cl in single parietal cells (PC) in isolated rabbit gastric glands incubated in HCO3/CO2-buffered solutions. Steady-state pHi was 7.2 in both resting (50 microM cimetidine) and stimulated (100 microM histamine) PCs. Transmembrane anion (HCO3 or Cl) flux rates during Cl removal from or readdition to the perfusate were the same in resting and stimulated PCs. These rates increased at alkaline pHi, though this pHi dependence was small in the physiological range. Maximum velocity (Vmax) for Cl influx or HCO3 efflux was 80-110 mM/min at pHi 7.6-7.8, and the Km for extracellular concentrations of Cl (Clo) was 25 mM; in the physiological range (pHi 7.1-7.3), Vmax for anion fluxes was approximately 50 mM/min. Steady-state Cli in the unstimulated PC was 62 +/- 5 mM, but on histamine stimulation, Cli decreased rapidly to 25 mM and then increased back to a steady-state level of 44 mM. HCO3 fluxes due to Cl removal or readdition were completely blocked by 0.5 mM 4,4'-diisothiocyanatodihydrostilbene-2,2'-disulfonic acid (H2DIDS), but Cl fluxes were only inhibited by 80%. H2DIDS did not inhibit the decrease in Cli that occurred with histamine treatment. Diphenylamine carboxylate (0.5 mM) inhibited Cl flux by only 50% and caused no additional inhibition of Cl flux when used in conjunction with H2DIDS. Transmembrane anion fluxes during solution Cl removal or readdition occurred 80% through the anion exchanger at the basal membrane and 20% through other pathway(s), presumably the Cl channel in the apical membrane. We conclude that the increase in transport activity via the Cl/HCO3 exchanger that occurs during histamine-induced increases in HCl secretion is due mostly to the decrease in Cli. In the resting cell with Cli = 62 mM, Clo = 120 mM, pHi = 7.2, and extracellular pH = 7.4, the anion exchanger is poised near its thermodynamic equilibrium. During histamine stimulation Cli drops from 62 mM to 44 mM, the thermodynamic equilibrium of the anion exchanger at the basolateral membrane is disturbed, and the anion exchanger then exchanges cellular HCO3 for extracellular Cl. Cli serves a crucial regulatory role in stimulus-secretion coupling in the PC.
Thomas
H A
HA
Department of Molecular and Cell Biology, University of California, Berkeley.
Machen
T E
TE
D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D002021:Buffers; D002352:Carrier Proteins / Q000378:metabolism*; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides / Q000378:metabolism*; D005452:Fluoresceins / Q000737:chemistry; D006632:Histamine / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D010295:Parietal Cells, Gastric / Q000378:metabolism*; D011806:Quinolinium Compounds / Q000737:chemistry; D011817:Rabbits; D013050:Spectrometry, Fluorescence; D062367:ortho-Aminobenzoates / Q000494:pharmacology
D001639:Bicarbonates; D002021:Buffers; D002352:Carrier Proteins; D027963:Chloride-Bicarbonate Antiporters; D002712:Chlorides; D005452:Fluoresceins; D011806:Quinolinium Compounds; D062367:ortho-Aminobenzoates; D012856:4-Acetamido-4'-isothiocyanatostilbene-2,2'-disulfonic Acid; D006632:Histamine; C052335:6-methoxy-N-(3-sulfopropyl)quinolinium; C043829:2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein; C001209:fenamic acid; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid