Voltage-dependent inhibition of Cx50 hemichannels by extracellular BQ⁺ suggests a binding site in the pore. Recordings of single-hemichannel currents in excised patches in control (left) and in 500 µM BQ⁺ (right) at membrane potentials of +20, −30, −50, −70, and −90 mV are shown. BQ⁺ was applied to the extracellular side of the hemichannel. Dashed lines in the current traces indicate the closed state. In the absence of BQ⁺, Cx50 hemichannels predominantly reside in the open state at voltages between +20 and −70 mV. With further hyperpolarization, Cx50 hemichannels tend to gate to long-lived subconductance states (see trace at −90 mV). The application of 500 µM BQ⁺ to the extracellular side caused strong inhibition at large hyperpolarizing voltages, but little inhibition at smaller hyperpolarizing voltages (compare P_o in BQ⁺ at −30 and −90 mV). An increase in inhibition with hyperpolarization, which tends to drive BQ⁺ into the hemichannel, is suggestive of a binding site within the aqueous pore.