Steady-state voltage-dependent gating and conduction kinetics of single K+ channels in the membrane of cytoplasmic drops ofChara australis

Summary Cytoplasmic drops, covered by a membrane derived from the tonoplast, were obtained from the internodal cells ofChara australis. Patch-clamp measurements were made on this membrane using the droplet-attached configuration with the membrane patch voltage clamped at values from –250 to 50 mV. Single-channel records, filtered at 5 kHz, were analyzed to elucidate the kinetics of the ion gating reaction of the K⁺-selective channel. The current-voltage characteristics for single channels exhibit saturation and are shown to be consistent with Läuger's theory of diffusion-limited ion flow through pores (P. Läuger,Biochim. Biophys. Acta455:493–509, 1976). The time-averaged behavior of the K⁺ conductance has a maximum at –100 to –150 mV which is produced by the combination of two distinct mechanisms: (1) The channel spending more time in long-lived closed states at positive voltages and (2) a large decrease in the mean open lifetime at more negative voltages. The channel activity shows bursting behavior with opening and closing rates that are voltage-dependent. The mean open time is the kinetic parameter most sensitive to membrane potential, showing a maximum between –100 to –150 mV. The distribution of open times is dominated by one exponential component (time constant 0.3 to 10 msec). In some cases an additional rapidly decaying exponential component was detectable (time constant=0.1 msec). The closed distributions contained were observed to obtain up to four exponential components with time constants over the range 0.1 to 200 msec. However, the voltage dependence of the closed-time distributions suggests an eight-state model for this channel.