Figure 1.

Na⁺/H⁺ transport assays were performed using the cell-permeable pH-sensitive fluorophore SNARF-4 F acetoxymethyl ester and multi-photon laser scanning microscopy. Differentiated human duodenal enteroids were first incubated in the presence of NH₄Cl buffer, then pre-pulsed with Na⁺-free buffer (tetramethylammonium chloride, TMA) to induce intracellular pH acidification, and finally transferred to Na⁺ buffer to follow the rate of alkalinization. Specific inhibition of NHE3 with S3226 (20 μmol/L) dramatically decreased the rate of Na⁺ driven alkalinization (NHE activity) in 3-D differentiated human duodenal enteroids relative to specimens treated with HOE-694 (50 μmol/L) to block activity of other NHE isoforms. Please note the similarity of Na⁺-dependent alkalinization in the presence of HOE-694 plus S3226 and S3226 alone, supporting that nearly all NHE activities are related to NHE3. Graph depicts a representative experiment for each condition; each condition was evaluated in triplicate. (A color version of this figure is available in the online journal.)