Time-irreversible Subconductance Gating Associated with Ba(2+) Block of Large Conductance Ca(2+)-activated K(+) Channels

Ba(2+) block of large conductance Ca(2+)-activated K(+) channels was studied in patches of membrane excised from cultures of rat skeletal muscle using the patch clamp technique. Under conditions in which a blocking Ba(2+) ion would dissociate to the external solution (150 mM N-methyl-d-glucamine(+) (o), 500 mM K(+) (i), 10 μM Ba(2+) (i), +30 mV, and 100 μM Ca(2+) (i) to fully activate the channel), Ba(2+) blocks with a mean duration of ∼2 s occurred, on average, once every ∼100 ms of channel open time. Of these Ba(2+) blocks, 78% terminated with a single step in the current to the fully open level and 22% terminated with a transition to a subconductance level at ∼0.26 of the fully open level (preopening) before stepping to the fully open level. Only one apparent preclosing was observed in ∼10,000 Ba(2+) blocks. Thus, the preopenings represent Ba(2+)-induced time-irreversible subconductance gating. The fraction of Ba(2+) blocks terminating with a preopening and the duration of preopenings (exponentially distributed, mean = 0.75 ms) appeared independent of changes in [Ba(2+)](i) or membrane potential. The fractional conductance of the preopenings increased from 0.24 at +10 mV to 0.39 at +90 mV. In contrast, the average subconductance level during normal gating in the absence of Ba(2+) was independent of membrane potential, suggesting different mechanisms for preopenings and normal subconductance levels. Preopenings were also observed with 10 mM Ba(2+) (o) and no added Ba(2+) (i). Adding K(+), Rb(+), or Na(+) to the external solution decreased the fraction of Ba(2+) blocks with preopenings, with K(+) and Rb(+) being more effective than Na(+). These results are consistent with models in which the blocking Ba(2+) ion either induces a preopening gate, and then dissociates to the external solution, or moves to a site located on the external side of the Ba(2+) blocking site and acts directly as the preopening gate.