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Allosteric voltage gating of potassium channels II. Mslo channel gating charge movement in the absence of Ca(2+).

Horrigan FT, Aldrich RW - J. Gen. Physiol. (1999)

Bottom Line: These results can be understood in terms of the allosteric voltage-gating scheme developed in the preceding paper (Horrigan, F.T., J.Physiol. 114:277-304).The model contains five open (O) and five closed (C) states arranged in parallel, and the kinetic and steady-state properties of mSlo gating currents exhibit multiple components associated with C-C, O-O, and C-O transitions.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California 94305, USA.

ABSTRACT
Large-conductance Ca(2+)-activated K(+) channels can be activated by membrane voltage in the absence of Ca(2+) binding, indicating that these channels contain an intrinsic voltage sensor. The properties of this voltage sensor and its relationship to channel activation were examined by studying gating charge movement from mSlo Ca(2+)-activated K(+) channels in the virtual absence of Ca(2+) (<1 nM). Charge movement was measured in response to voltage steps or sinusoidal voltage commands. The charge-voltage relationship (Q-V) is shallower and shifted to more negative voltages than the voltage-dependent open probability (G-V). Both ON and OFF gating currents evoked by brief (0.5-ms) voltage pulses appear to decay rapidly (tau(ON) = 60 microseconds at +200 mV, tau(OFF) = 16 microseconds at -80 mV). However, Q(OFF) increases slowly with pulse duration, indicating that a large fraction of ON charge develops with a time course comparable to that of I(K) activation. The slow onset of this gating charge prevents its detection as a component of I(gON), although it represents approximately 40% of the total charge moved at +140 mV. The decay of I(gOFF) is slowed after depolarizations that open mSlo channels. Yet, the majority of open channel charge relaxation is too rapid to be limited by channel closing. These results can be understood in terms of the allosteric voltage-gating scheme developed in the preceding paper (Horrigan, F.T., J. Cui, and R.W. Aldrich. 1999. J. Gen. Physiol. 114:277-304). The model contains five open (O) and five closed (C) states arranged in parallel, and the kinetic and steady-state properties of mSlo gating currents exhibit multiple components associated with C-C, O-O, and C-O transitions.

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Changes in OFF kinetics with channel activation. (A) A family of IgOFF evoked at 2100 mV after pulses to +140 mV of 0.06–20 ms duration (from Fig. 6 A). Current amplitude is maximal after a 0.5-ms pulse, but IgOFF decays more slowly as pulse duration increases. The baseline for each record is set to the mean current during an interval 4–5 ms after the pulse. (B) The decay of OFF currents are fit by double-exponential functions with τF = 15.5 μs and τM = 59 μs. (C) QOFF obtained by integrating IgOFF from A achieves a steady state within 300 μs after a 0.06-ms pulse (arrow) but relaxes more slowly after longer pulses. (D) The kinetics of QOFF relaxation after a brief (0.06 ms) or prolonged (10–20 ms) pulses are compared by plotting QOFF–Qss on a semilog scale. Qss is the steady-state value of QOFF measured 3 ms after the pulse. The 0.06-ms trace is fit by a single-exponential function (τF = 15.5 μs). The 10–20-ms trace, representing an average of 10-, 15-, and 20-ms records, is fit by a triple exponential (solid line, τF = 15.5 μs, τM = 59 μs, τS = 448 μs) where the individual components are indicated by dashed lines. (E) A family of QOFF–Qss for the data in C. Traces are fit with triple exponential functions with the time constants determined from D. (F) QOFF component amplitudes from these fits are plotted versus pulse duration. The relaxation of all three components is fit by exponential functions (solid lines) with a time constant of 4.22 ms. Error bars represent the component amplitudes obtained when τM is changed by ±10% (with τF and τS held constant). The fast component of ON charge (Qfast) is indicated by an arrow. (G) Fast and Medium IgOFF component amplitudes determined from B are plotted versus pulse duration. Solid lines represent exponential fits with a time constant of 4.22 ms. (H) The allosteric model predicts three components of QOFF relaxation corresponding to the indicated transitions in the gating scheme.
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Figure 7: Changes in OFF kinetics with channel activation. (A) A family of IgOFF evoked at 2100 mV after pulses to +140 mV of 0.06–20 ms duration (from Fig. 6 A). Current amplitude is maximal after a 0.5-ms pulse, but IgOFF decays more slowly as pulse duration increases. The baseline for each record is set to the mean current during an interval 4–5 ms after the pulse. (B) The decay of OFF currents are fit by double-exponential functions with τF = 15.5 μs and τM = 59 μs. (C) QOFF obtained by integrating IgOFF from A achieves a steady state within 300 μs after a 0.06-ms pulse (arrow) but relaxes more slowly after longer pulses. (D) The kinetics of QOFF relaxation after a brief (0.06 ms) or prolonged (10–20 ms) pulses are compared by plotting QOFF–Qss on a semilog scale. Qss is the steady-state value of QOFF measured 3 ms after the pulse. The 0.06-ms trace is fit by a single-exponential function (τF = 15.5 μs). The 10–20-ms trace, representing an average of 10-, 15-, and 20-ms records, is fit by a triple exponential (solid line, τF = 15.5 μs, τM = 59 μs, τS = 448 μs) where the individual components are indicated by dashed lines. (E) A family of QOFF–Qss for the data in C. Traces are fit with triple exponential functions with the time constants determined from D. (F) QOFF component amplitudes from these fits are plotted versus pulse duration. The relaxation of all three components is fit by exponential functions (solid lines) with a time constant of 4.22 ms. Error bars represent the component amplitudes obtained when τM is changed by ±10% (with τF and τS held constant). The fast component of ON charge (Qfast) is indicated by an arrow. (G) Fast and Medium IgOFF component amplitudes determined from B are plotted versus pulse duration. Solid lines represent exponential fits with a time constant of 4.22 ms. (H) The allosteric model predicts three components of QOFF relaxation corresponding to the indicated transitions in the gating scheme.

Mentions: The large slow component of Qp(t) observed at V ≥ +140 mV in Fig. 6 B indicates that QOFF increases with pulse duration. In contrast, the peak amplitude of IgOFF remains roughly constant or decreases with pulse duration at the same voltages (Fig. 6 A). That IgOFF can decrease or remain constant while its integral (QOFF) increases implies that the kinetics of OFF current change with pulse duration. This change is obvious in Fig. 7 A, which compares OFF currents evoked at −100 mV after pulses to +140 mV of different duration (0.06–20 ms). Two components of IgOFF are evident from these records. After brief pulses (0.06 or 0.11 ms), OFF current decays with a rapid exponential time course, but an additional slower component appears as pulse duration is increased. The decay of IgOFF at all pulse durations can be well fit by double-exponential functions with time constants of 15.5 and 59 μs (Fig. 7 B). Both components decay within 300 μs and therefore appear to be fast relative to the time course of channel closing. Potassium tail currents decay with a time constant of 172 ± 15 μs at −80 mV (Horrigan et al. 1999) and therefore require approximately 5τ(IK) = 900 μs to decay completely. However, a slower component of OFF charge movement can be detected by plotting the integral of IgOFF (QOFF(t); Fig. 7 C). QOFF(t) measured after a brief (0.06 ms) voltage pulse achieves a steady state within 300 μs (Fig. 7 C, arrow), consistent with the rapid decay of IgOFF. In contrast, QOFF(t) measured after a 20-ms pulse requires >1 ms to reach a steady state, indicating a slow component of charge relaxation. This component of QOFF is not evident in the corresponding IgOFF trace because it is slow and represents <20% of the total OFF charge.


Allosteric voltage gating of potassium channels II. Mslo channel gating charge movement in the absence of Ca(2+).

Horrigan FT, Aldrich RW - J. Gen. Physiol. (1999)

Changes in OFF kinetics with channel activation. (A) A family of IgOFF evoked at 2100 mV after pulses to +140 mV of 0.06–20 ms duration (from Fig. 6 A). Current amplitude is maximal after a 0.5-ms pulse, but IgOFF decays more slowly as pulse duration increases. The baseline for each record is set to the mean current during an interval 4–5 ms after the pulse. (B) The decay of OFF currents are fit by double-exponential functions with τF = 15.5 μs and τM = 59 μs. (C) QOFF obtained by integrating IgOFF from A achieves a steady state within 300 μs after a 0.06-ms pulse (arrow) but relaxes more slowly after longer pulses. (D) The kinetics of QOFF relaxation after a brief (0.06 ms) or prolonged (10–20 ms) pulses are compared by plotting QOFF–Qss on a semilog scale. Qss is the steady-state value of QOFF measured 3 ms after the pulse. The 0.06-ms trace is fit by a single-exponential function (τF = 15.5 μs). The 10–20-ms trace, representing an average of 10-, 15-, and 20-ms records, is fit by a triple exponential (solid line, τF = 15.5 μs, τM = 59 μs, τS = 448 μs) where the individual components are indicated by dashed lines. (E) A family of QOFF–Qss for the data in C. Traces are fit with triple exponential functions with the time constants determined from D. (F) QOFF component amplitudes from these fits are plotted versus pulse duration. The relaxation of all three components is fit by exponential functions (solid lines) with a time constant of 4.22 ms. Error bars represent the component amplitudes obtained when τM is changed by ±10% (with τF and τS held constant). The fast component of ON charge (Qfast) is indicated by an arrow. (G) Fast and Medium IgOFF component amplitudes determined from B are plotted versus pulse duration. Solid lines represent exponential fits with a time constant of 4.22 ms. (H) The allosteric model predicts three components of QOFF relaxation corresponding to the indicated transitions in the gating scheme.
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Related In: Results  -  Collection

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Figure 7: Changes in OFF kinetics with channel activation. (A) A family of IgOFF evoked at 2100 mV after pulses to +140 mV of 0.06–20 ms duration (from Fig. 6 A). Current amplitude is maximal after a 0.5-ms pulse, but IgOFF decays more slowly as pulse duration increases. The baseline for each record is set to the mean current during an interval 4–5 ms after the pulse. (B) The decay of OFF currents are fit by double-exponential functions with τF = 15.5 μs and τM = 59 μs. (C) QOFF obtained by integrating IgOFF from A achieves a steady state within 300 μs after a 0.06-ms pulse (arrow) but relaxes more slowly after longer pulses. (D) The kinetics of QOFF relaxation after a brief (0.06 ms) or prolonged (10–20 ms) pulses are compared by plotting QOFF–Qss on a semilog scale. Qss is the steady-state value of QOFF measured 3 ms after the pulse. The 0.06-ms trace is fit by a single-exponential function (τF = 15.5 μs). The 10–20-ms trace, representing an average of 10-, 15-, and 20-ms records, is fit by a triple exponential (solid line, τF = 15.5 μs, τM = 59 μs, τS = 448 μs) where the individual components are indicated by dashed lines. (E) A family of QOFF–Qss for the data in C. Traces are fit with triple exponential functions with the time constants determined from D. (F) QOFF component amplitudes from these fits are plotted versus pulse duration. The relaxation of all three components is fit by exponential functions (solid lines) with a time constant of 4.22 ms. Error bars represent the component amplitudes obtained when τM is changed by ±10% (with τF and τS held constant). The fast component of ON charge (Qfast) is indicated by an arrow. (G) Fast and Medium IgOFF component amplitudes determined from B are plotted versus pulse duration. Solid lines represent exponential fits with a time constant of 4.22 ms. (H) The allosteric model predicts three components of QOFF relaxation corresponding to the indicated transitions in the gating scheme.
Mentions: The large slow component of Qp(t) observed at V ≥ +140 mV in Fig. 6 B indicates that QOFF increases with pulse duration. In contrast, the peak amplitude of IgOFF remains roughly constant or decreases with pulse duration at the same voltages (Fig. 6 A). That IgOFF can decrease or remain constant while its integral (QOFF) increases implies that the kinetics of OFF current change with pulse duration. This change is obvious in Fig. 7 A, which compares OFF currents evoked at −100 mV after pulses to +140 mV of different duration (0.06–20 ms). Two components of IgOFF are evident from these records. After brief pulses (0.06 or 0.11 ms), OFF current decays with a rapid exponential time course, but an additional slower component appears as pulse duration is increased. The decay of IgOFF at all pulse durations can be well fit by double-exponential functions with time constants of 15.5 and 59 μs (Fig. 7 B). Both components decay within 300 μs and therefore appear to be fast relative to the time course of channel closing. Potassium tail currents decay with a time constant of 172 ± 15 μs at −80 mV (Horrigan et al. 1999) and therefore require approximately 5τ(IK) = 900 μs to decay completely. However, a slower component of OFF charge movement can be detected by plotting the integral of IgOFF (QOFF(t); Fig. 7 C). QOFF(t) measured after a brief (0.06 ms) voltage pulse achieves a steady state within 300 μs (Fig. 7 C, arrow), consistent with the rapid decay of IgOFF. In contrast, QOFF(t) measured after a 20-ms pulse requires >1 ms to reach a steady state, indicating a slow component of charge relaxation. This component of QOFF is not evident in the corresponding IgOFF trace because it is slow and represents <20% of the total OFF charge.

Bottom Line: These results can be understood in terms of the allosteric voltage-gating scheme developed in the preceding paper (Horrigan, F.T., J.Physiol. 114:277-304).The model contains five open (O) and five closed (C) states arranged in parallel, and the kinetic and steady-state properties of mSlo gating currents exhibit multiple components associated with C-C, O-O, and C-O transitions.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Physiology, Howard Hughes Medical Institute, Stanford University School of Medicine, Stanford, California 94305, USA.

ABSTRACT
Large-conductance Ca(2+)-activated K(+) channels can be activated by membrane voltage in the absence of Ca(2+) binding, indicating that these channels contain an intrinsic voltage sensor. The properties of this voltage sensor and its relationship to channel activation were examined by studying gating charge movement from mSlo Ca(2+)-activated K(+) channels in the virtual absence of Ca(2+) (<1 nM). Charge movement was measured in response to voltage steps or sinusoidal voltage commands. The charge-voltage relationship (Q-V) is shallower and shifted to more negative voltages than the voltage-dependent open probability (G-V). Both ON and OFF gating currents evoked by brief (0.5-ms) voltage pulses appear to decay rapidly (tau(ON) = 60 microseconds at +200 mV, tau(OFF) = 16 microseconds at -80 mV). However, Q(OFF) increases slowly with pulse duration, indicating that a large fraction of ON charge develops with a time course comparable to that of I(K) activation. The slow onset of this gating charge prevents its detection as a component of I(gON), although it represents approximately 40% of the total charge moved at +140 mV. The decay of I(gOFF) is slowed after depolarizations that open mSlo channels. Yet, the majority of open channel charge relaxation is too rapid to be limited by channel closing. These results can be understood in terms of the allosteric voltage-gating scheme developed in the preceding paper (Horrigan, F.T., J. Cui, and R.W. Aldrich. 1999. J. Gen. Physiol. 114:277-304). The model contains five open (O) and five closed (C) states arranged in parallel, and the kinetic and steady-state properties of mSlo gating currents exhibit multiple components associated with C-C, O-O, and C-O transitions.

Show MeSH
Related in: MedlinePlus