Na⁺ dependence of mouse Ae4 and human AE4 exchangers expressed in CHO-K1 cells. (A) CHO-K1 cells transfected with a plasmid containing the mouse Ae4 cDNA were loaded with BCECF-AM. A rapid increase in pH_i was observed upon external Cl⁻ reduction in the presence of external Na⁺ (black circles; n = 11; changing solution A to B [Table 1]). In contrast, the rise in pH_i was prevented when Na⁺ (blue circles; n = 6; changing solution C to D [Table 1])- or HCO₃⁻ (green circles; n = 8; changing solution E to F [Table 1])-free external solutions were used. (B) Similar experiments as shown in A using the same external solutions performed on CHO-K1 cells expressing mouse Ae4 but using the Cl⁻ indicator SPQ. A rapid loss of intracellular [Cl⁻] was observed upon external [Cl⁻] reduction in the presence of external Na⁺ (black circles; n = 10), but there was essentially no change in [Cl⁻]_i in HCO₃⁻ (green circles; n = 7)- or Na⁺ (blue circles; n = 6)-free external solutions. (C) There was no change in pH_i in CHO-K1 cells expressing mouse Ae4 (black circles; n = 8) and human AE4 (green circles; n = 5) in response to a decrease in the external [Cl⁻] in a NMDG-containing solution (changing solution C to D [Table 1]), but there was a dramatic increase in the pH_i when switched to a Na⁺-containing external solution. These manipulations resulted in little change in pH_i on nontransfected cells (blue circles; n = 9; changing solution D to B [Table 1]). (A–C) Results are presented as the mean ± SEM. (D) Summary of the experiments as shown in C. Data are expressed as the mean ± SEM of at least five cells per experiment from at the least three different electroporations. For nontransfected CHO-K1 cells, the blue bar is significantly less, “a” compared with “b” (P < 0.05, one-way ANOVA followed by Bonferroni’s post hoc test).