Gastrointestinal peptides activate Na(+)-H+ exchanger in AR42J cells by increasing its affinity for intracellular H+.
Regulation of intracellular pH (pHi) was studied by dual wavelength fluorometry in monolayers of pancreatic AR42J cells loaded with the fluorescent probe 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. In cells superfused with N-2-hydroxyethylpiperazine-N'-2-ethanesulfonic acid-buffered solution at pH 7.40, basal pHi was determined to be 7.15 +/- 0.13. Na(+)-H+ exchange could be demonstrated in both resting cells and cells subjected to acid loading by use of transient exposure to NH4Cl. Na(+)-H+ exchange was completely blocked by 300 microM amiloride and was dependent on extracellular Na+ (apparent Km = 25 mM). When the concentration of the NH4Cl pulse was varied (0.5-25 mM), the rate of pHi recovery increased as pHi became acidic, reaching a maximum of 0.007 pH units/s at pHi of 6.4. Gastrointestinal hormones, including pentagastrin, cholecystokinin, and bombesin, increased the rate of Na(+)-H+ exchange without affecting cellular buffer capacity (21.5 +/- 1.8 mM/pH unit), thereby leading to an intracellular alkalinization. This was accompanied by a shift in the curve of Na(+)-H+ exchange as a function of pHi to more alkaline values, although the maximum rate of pH recovery was unchanged. Neither protein kinase C nor Ca2+ could be conclusively linked to activation of Na(+)-H+ exchange, raising the possibility of a more direct, receptor-controlled mechanism.
Bastié
M J
MJ
Department of Physiology, University of Michigan, Ann Arbor 48109.
Williams
J A
JA
D000584:Amiloride / Q000494:pharmacology; D000595:Amino Acid Sequence; D000818:Animals; D001839:Bombesin / Q000494:pharmacology; D002217:Carbachol / Q000494:pharmacology; D002352:Carrier Proteins / Q000378:metabolism*; D002460:Cell Line; D005768:Gastrointestinal Hormones / Q000494:pharmacology*; D006863:Hydrogen-Ion Concentration; D007700:Kinetics; D008969:Molecular Sequence Data; D010190:Pancreatic Neoplasms; D010418:Pentagastrin / Q000494:pharmacology; D051381:Rats; D012844:Sincalide / Q000494:pharmacology; D012965:Sodium Chloride / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers; D013373:Substance P / Q000494:pharmacology; D013755:Tetradecanoylphorbol Acetate / Q000494:pharmacology; D014407:Tumor Cells, Cultured / Q000187:drug effects / Q000378:metabolism*
D002352:Carrier Proteins; D005768:Gastrointestinal Hormones; D017923:Sodium-Hydrogen Exchangers; D013373:Substance P; D012965:Sodium Chloride; D000584:Amiloride; D002217:Carbachol; D010418:Pentagastrin; D012844:Sincalide; D013755:Tetradecanoylphorbol Acetate; D001839:Bombesin