Sodium induces simultaneous changes in cytosolic calcium and pH in salt-tolerant quince protoplasts

Previous experiments with salt-resistant quince BA29 (Cydonia oblonga cv. Mill.) have shown that this cultivar takes up sodium transiently into the cytosol of shoot protoplasts only in the absence of calcium chloride, or at <1 mM calcium chloride. Addition of NaCl ≥100 mM to single protoplasts from in vitro-cultivated quince in the presence of 1.0 mM calcium induced instant changes in the cytosolic concentrations of calcium and protons. These changes were investigated by use of tetra [acetoxymethyl] esters of the fluorescent stilbene chromophores Fura 2 and bis-carboxyethyl-carboxyfluorescein (BCECF), respectively. The cytosolic Ca2+ dynamics in the protoplasts were dependent on the concentration of NaCl added. The changes in calcium differed in amplitude and final concentration and were correlated in time mainly with changes in pH. Addition of 100–400 mM NaCl to the protoplasts caused an oscillating increase in the cytosolic level of calcium, and then a decrease. Addition of mannitol, of equiosmolar concentration to NaCl, did not increase the cytosolic calcium concentration. Moreover, there was no increase in cytosolic calcium when NaCl was added in the presence of calcium binding ethylene glycol-bis(beta-aminoethylether)-N,N,N′,N′-tetra acetic acid (EGTA), or lantan or verapamil, two inhibitors of plasma membrane calcium channels. Therefore, we conclude that, in salt-resistant quince, sodium induces an influx of calcium into the cytosol by plasma membrane calcium channels, and a simultaneous increase in cytosolic pH. Because these changes were obtained in the presence of 1 mM calcium in the medium, they were not due to sodium uptake into the cytosol.