Panel A shows structural views of the 8A/8C channel, with the extracellular domain, transmembrane domain, intracellular domain, and leucine rich repeat domain labeled. Panel B shows representative whole cell current traces for 8A/8C under isotonic, hypotonic, and 10 micromolar DCPIB conditions. Scale bars indicate 500 picoamperes and 250 milliseconds. Panel C shows a line graph of current density versus membrane voltage for isotonic, hypotonic, and DCPIB conditions. The x-axis represents membrane voltage in millivolts, and the y-axis represents current density in picoamperes per picofarad. Panel D shows concentration response curves for DCPIB inhibition of 8A/8C and several paired residue variants. The x-axis represents DCPIB concentration in micromolar on a logarithmic scale, and the y-axis represents percentage inhibition. Panel E shows representative whole cell current traces for the indicated paired residue variants: R103/L105R, E6A/E6A, T48D/M48D, Y124F/Y126F, L134E/L136E, and L134K/L136K. Scale bars indicate 500 picoamperes and 250 milliseconds.
Mutational analysis of DCPIB sensitivity in heteromeric 8A/8C currents. (A) Side (left) and top (right) views of 8A/8C heteromer (PDB ID: 8B41) (Rutz et al., 2023). (B) Representative whole-cell heteromeric 8A/8C current recorded from a cell dialyzed with normal pipette solution and cell swelling with hypotonic buffer (left), after bath application of 10 μM DCPIB (right). The scale is the same for both traces. (C) I–V relationships of the 8A/8C heteromer from cells in isotonic, hypotonic, and hypotonic buffer with 10 μM DCPIB (n = 5). (D) CRC data recorded at +100 mV for heteromeric 8A/8C WT and the indicated mutants (8A/8C n = 5–6, mutants n = 5 each). Data are mean ± SEM. (E) Representative whole-cell currents of the indicated mutants recorded from a cell dialyzed with normal pipette solution and cell swelling with hypotonic buffer. The scale is the same for all traces.