Modulation of calcium signals by intracellular pH in isolated rat pancreatic acinar cells.
1. We have investigated the interactions between intracellular pH (pH1) and the intracellular free calcium concentration ([Ca2+]i) in isolated rat pancreatic acinar cells. The fluorescent dyes fura-2 and BCECF were used to measure [Ca2+]i and pHi, respectively. 2. Sodium acetate and ammonium chloride (NH4Cl) were used to acidify and alkalinize pHi, respectively. Cytosolic acidification had no effect on [Ca2+]i in resting pancreatic acinar cells, whereas cytosolic alkalinization released Ca2+ from intracellular stores. 3. Cytosolic acidification using either acetate or a CO2-HCO3(-)-buffered medium enhanced Ca2+ signals evoked by acetylcholine (ACh) and cholecystokinin (CCK). In contrast, both NH4Cl and trimethylamine (TMA) inhibited Ca2+ signals during stimulation with either ACh or CCK. This inhibitory effect was also observed in the absence of extracellular Ca2+, and was therefore not due to changes in Ca2+ entry. 4. Calcium oscillations evoked by physiological concentrations of CCK were enhanced by cytosolic acidification and inhibited by cytosolic alkalinization. 5. In order to determine the effects of pHi upon Ca2+ handling by intracellular Ca2+ stores, intraorganellar [Ca2+] was monitored using the low affinity Ca2+ indicator mag-fura-2 in permeabilized cells. Addition of NH4Cl, which is expected to alkalinize intraorganellar pH, did not alter intraorganellar [Ca2+] in permeabilized cells, suggesting that changing intraorganellar pH does not release Ca2+ from intracellular stores. Addition of NH4Cl or acetate also did not affect the rate of Ca2+ release induced by inositol 1,4,5-trisphosphate (InsP3). 6. Modification of extraorganellar ('cytosolic') pH did not affect the rate of ATP-dependent Ca2+ uptake into stores, but did modify the rate of Ca2+ release evoked by submaximal concentrations of InsP3. The rate of Ca2+ release was increased at more alkaline extraorganellar pHs. These results would suggest that manipulation of intraorganellar pH does not affect Ca2+ handling by the intracellular stores. In contrast, extraorganellar ('cytosolic') pH does affect InsP3-induced Ca2+ release from the stores. 7. In conclusion, changes in intracellular pH in pancreatic acinar cells can profoundly alter cytosolic [Ca2+]. This may shed light on earlier observations whereby cell-permeant weak acids and bases can modulate fluid secretion in epithelia.
Speake
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Cell Physiology Group, School of Biological Sciences (G.38), University of Manchester, UK.
Elliott
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AC
D000085:Acetates / Q000494:pharmacology; D000109:Acetylcholine / Q000819:agonists / Q000494:pharmacology; D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000819:agonists / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002766:Cholecystokinin / Q000819:agonists / Q000494:pharmacology; D005071:Evoked Potentials / Q000187:drug effects / Q000502:physiology; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D015544:Inositol 1,4,5-Trisphosphate / Q000494:pharmacology; D007424:Intracellular Fluid / Q000737:chemistry; D015388:Organelles / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D010179:Pancreas / Q000166:cytology* / Q000187:drug effects / Q000502:physiology; D051381:Rats; D017207:Rats, Sprague-Dawley; D015398:Signal Transduction / Q000187:drug effects / Q000502:physiology*
D000085:Acetates; D000643:Ammonium Chloride; D015544:Inositol 1,4,5-Trisphosphate; D002766:Cholecystokinin; D000109:Acetylcholine; D002118:Calcium