Based on sequence analysis, the salt overly sensitive (SOS1) gene has been suggested to function as a Na+/H+ antiporter located at the plasma membrane of plant cells, being expressed mostly in the meristem zone of the root and in the parenchyma cells surrounding the vascular tissue of the stem. In this study, we compared net H+ and Ca2+ fluxes and intracellular pH and [Ca2+]cyt in the root meristem zone of Arabidopsis wild-type (WT) and sos mutants before and after salt stress. In addition, we studied the effect of pretreatment with amiloride (an inhibitor of Na+/H+ antiporters) on net ion fluxes, intracellular pH and intracellular Ca2+ activity ([Ca2+]cyt) in WT plants and sos1 mutants before and after salt stress. Net ion fluxes were measured using microelectrode ion flux estimation (MIFE) and intracellular pH and [Ca2+]cyt using fluorescence lifetime imaging microscopy (FLIM) techniques. During the first 15 min after NaCl application, sos1 mutants showed net H+ efflux and intracellular alkalinization in the meristem zone, whereas sos2 and sos3 mutants and WT showed net H+ influx and slight intracellular acidification in the meristem zone. Treatment with amiloride led to intracellular acidification and lower net H+ flux in WT plants and to a decrease in intracellular Ca2+ in WT and sos1 plants. WT plants pretreated with amiloride did not show positive net H+ flux and intracellular acidification. After NaCl application, internal pH shifted to higher values in WT and sos1 plants. However, absolute values of H+ fluxes were higher and internal pH values were lower in WT plants pretreated with amiloride compared with sos1 mutants. Therefore, the SOS1 transporter is involved in H+ influx into the meristem zone of Arabidopsis roots, or it may function as a Na+/H+ antiporter. Amiloride affects SOS1 and other Na+/H+ antiporters in plant cells because of its ability to decrease the H+ gradient across the plasma membrane.