ANO5-PLS associated ionic currents are nonselective. (a) Representative whole-cell I–V curves recorded from ANO5-expressing HEK293 cells with 200 µM free Ca²⁺ in the pipette with external solutions containing 150 mM (black) or 15 mM (red) CsCl. Currents were generated by ramps from −100 to +100 mV. (b) Cation permeability of ANO5 and ANO1 currents relative to Cl⁻ (P_x/P_Cl) were calculated by changes in reversal potential as shown in panel a. Numbers in or above column indicate the number of cells assayed. Differences between ANO1 and ANO5 were evaluated for each condition using a paired Student's t test (**, P = 0.0012; ****, P = 0.0001). Differences in cation permeability for ANO5 were compared using a one-way ANOVA (P = 0.2693). (c) Ionic current amplitudes (black) and PLS (red) of HEK293 cells expressing ANO1, ANO5, and ANO1 harboring the ANO5 SCRD (ANO151) and ANO5 harboring the ANO1 SCRD ANO515. Differences in cells scrambling versus not scrambling were compared using a Fisher exact test (**, P = 0.0021; ****, P < 0.0001; ns, P = 0.5658). Numbers below indicate the number of cells assayed. Error bars indicate SEM.