Fig. 3. Apical Na⁺/H⁺ exchange rates in NHE3-expressing Caco-2BBe cells are sensitive to tenapanor. (A) TEER curve of hNHE3vsv expressing Caco-2BBe cells after 5 acid suicide selection cycles on transwell membranes from day 0 to day 14 after seeding. (B-D) The NHE function was estimated by first using NH₄Cl prepulse followed by Na⁺ free buffer C to cause intracellular acidification, and then Na⁺ was added selectively to the apical side during the recovery phase to stimulate apical NHE activity. Inhibitors were added 5 minutes before the addition of Na⁺. (B) pH_i curve of total apical NHE activity of C2NHE3vsv AS5 cells. (C) pH_i curve of apical NHE activity in the presence of 60 μM HOE642 in the apical solution. (D) pH_i curve in the presence of 60 μM HOE642 and 1 μM tenapanor in the apical solution. (E) Comparisons of apical NHE activities of C2NHE3vsv cells under different inhibitory conditions. (n=3, mean ± SEM, one-way ANOVA with Tukey's multiple comparison tests, **p<0.001).