INTRACELLULAR ALKALINIZATION MOBILIZES CALCIUM FROM AGONIST-SENSITIVEPOOLS IN RAT LACRIMAL ACINAR-CELLS

1. We have investigated interactions between intracellular pH (pH(i)) and the intracellular free calcium concentration ([Ca2+](i)) in collag enase-isolated rat lacrimal acinar cells. The fluorescent dyes fura-2 and 2',7'-bis(carboxyethyl)-5-carboxyfluorescein (BCECF) were used to measure [Ca2+](i) and pH(i), respectively. 2. Application of the weak base NH4Cl alkalinized the cytosol and caused a dose-dependent increas e in [Ca2+](i). Trimethylamine (TMA) also alkalinized the cytosol and increased [Ca2+](i). The increase in [Ca2+](i) evoked by NH4Cl or TMA mras much smaller than that evoked by the secretory agonist acetylchol ine (ACh). 3. Application of NH4Cl also increased [Ca2+](i) in cells b athed in Ca2+-free medium, indicating that NH4Cl released Ca2+ from an intracellular pool. 4. Ammonium chloride had no effect on [Ca2+](i) i n cells bathed in Ca2+-free medium if agonist-sensitive intracellular Ca2+ pools had been depleted with either ACh or the microsomal Ca2+-AT Pase inhibitor 2,5-di(tert-butyl)hydroquinone. Treatment of cells with NH4Cl in Ca2+-free medium reduced the amount of Ca2+ released by ACh. These results suggest that NH4Cl released Ca2+ from the same intracel lular pool released by ACh. 5. Calcium release from the agonist-sensit ive pool was also triggered when the cytosol was alkalinized by removi ng the weak acid acetate. 6. Ammonium chloride caused a modest increas e in inositol phosphate production, suggesting that NH4Cl may have rel eased stored Ca2+ via an increase in the intracellular inositol 1,4,5- trisphosphate concentration. 7. The increase in [Ca2+](i) evoked by NH 4Cl was not sustained even in the presence of extracellular Ca2+. In c ontrast, when a low dose of ACh was used to evoke intracellular Ca2+ r elease of similar magnitude, sustained Ca2+ entry was observed. 8. Alk alinizing the cytosol appeared to partially inhibit Ca2+ entry trigger ed by thapsigargin. or by ACh. 9. We suggest that alkalinizing the cyt oplasm in unstimulated lacrimal acinar cells can release Ca2+ from the intracellular agonist-sensitive Ca2+ pool. However, releasing stored Ca2+ via alkalinization does not appear to trigger significant Ca2+ en try, perhaps because intracellular alkalinization inhibits either the Ca2+ entry pathway or the mechanism which couples the entry pathway to store depletion.