Key Points
- To characterize the calcium-dependent gating kinetics and underlying kinetic states of single large-conductance calcium-activated potassium channels in rat skeletal muscle.
- Employed the patch-clamp technique on excised membrane patches from cultured rat skeletal muscle in the absence of sodium and magnesium ions at +30 mV across varied intracellular calcium concentrations.
- Analyzed approximately 200,000 open and shut intervals from five single-channel patches alongside multichannel patches, focusing on normal-mode activity (96% of intervals).
- Derived dwell-time distributions using maximum likelihood techniques with corrections for missed events to model kinetic schemes.
- Increasing intracellular calcium elevated channel open probability, exhibiting a Hill coefficient of 3.7 ± 0.8 (range 3.0-5.0) and K0.5 of 14 ± 7 µM at pH 7.0 (n = 6), and a Hill coefficient of 3.0 ± 0.5 (range 2.2-3.7) with K0.5 of 9 ± 6 µM at pH 7.2 (n = 7).
- Up to fourfold differences in K0.5 were detected among individual channels measured under identical experimental conditions.
- High Hill coefficients and exponential components from dwell-time distributions indicated a minimum of five shut states requiring four or more bound calcium ions for activation, alongside three to four distinct open states.
Structured PICO
PPopulationCultured rat skeletal muscle (large-conductance Ca(2+)-activated K+ channels)
IInterventionVarying intracellular calcium concentrations ([Ca2+]i)
OOutcomeOpen probability (Popen) and dwell-time distributions of open and shut intervalssurrogate
The study demonstrates that large-conductance Ca(2+)-activated K+ channels in rat skeletal muscle likely bind four or more Ca2+ ions to become fully activated, requiring a minimum of five shut states.