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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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Voltage dependence and kinetics of fast charge movement. (A1) The normalized Qfast–V relationships for many experiments are fit with Boltzmann functions (z = 0.59 e, dashed lines). The solid line is a Boltzmann function indicating the mean half-activation voltage (〈Vh〉 = 155 mV, z = 0.59 e). (A2) The data from A1 (open symbols) are aligned by shifting them along the voltage axis by ΔVh = (〈Vh〉 − Vh). The mean Qfast–V (filled circles, mean ± SEM) is superimposed on the data together with two Boltzmann fits (Vh = 155 mV; solid line: z = 0.59e, dashed line: z = 0.55 e) and was determined by averaging the shifted data in 15-mV bins. (B1) Time constants of fast Ig relaxation (τgFast) were determined from exponential fits to ON and OFF currents for the experiments in A and are plotted on a log scale versus voltage. (C1) Three τgFast–V relationships from B1 that cover a large voltage range are compared. (B2 and C2) Data from B1 and C1 were shifted along the voltage axis by ΔVh (determined from A) and then normalized to the mean τgFast measured from +100 to +180 mV (59 μs). The solid line in B2 indicates the best fit of a two-state model of voltage-sensor activation where the relationship between the forward (α) and backward (β) rates are constrained such that J = α/β = 1 at +155 mV (zα = +0.30 e, zβ = −0.21 e, α(0) = 1,310 s−1, β(0) = 30,160 s−1). Dashed lines in A2, B2, and C2 represent the parameters ultimately used in the allosteric model to describe closed-state charge movement (zα = +0.33 e, zβ = −0.22 e, α(0) = 1,100 s−1, β(0) = 32,120 s−1).
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Figure 4: Voltage dependence and kinetics of fast charge movement. (A1) The normalized Qfast–V relationships for many experiments are fit with Boltzmann functions (z = 0.59 e, dashed lines). The solid line is a Boltzmann function indicating the mean half-activation voltage (〈Vh〉 = 155 mV, z = 0.59 e). (A2) The data from A1 (open symbols) are aligned by shifting them along the voltage axis by ΔVh = (〈Vh〉 − Vh). The mean Qfast–V (filled circles, mean ± SEM) is superimposed on the data together with two Boltzmann fits (Vh = 155 mV; solid line: z = 0.59e, dashed line: z = 0.55 e) and was determined by averaging the shifted data in 15-mV bins. (B1) Time constants of fast Ig relaxation (τgFast) were determined from exponential fits to ON and OFF currents for the experiments in A and are plotted on a log scale versus voltage. (C1) Three τgFast–V relationships from B1 that cover a large voltage range are compared. (B2 and C2) Data from B1 and C1 were shifted along the voltage axis by ΔVh (determined from A) and then normalized to the mean τgFast measured from +100 to +180 mV (59 μs). The solid line in B2 indicates the best fit of a two-state model of voltage-sensor activation where the relationship between the forward (α) and backward (β) rates are constrained such that J = α/β = 1 at +155 mV (zα = +0.30 e, zβ = −0.21 e, α(0) = 1,310 s−1, β(0) = 30,160 s−1). Dashed lines in A2, B2, and C2 represent the parameters ultimately used in the allosteric model to describe closed-state charge movement (zα = +0.33 e, zβ = −0.22 e, α(0) = 1,100 s−1, β(0) = 32,120 s−1).

Mentions: Fig. 4 A1 plots the normalized Qfast–V relationships for many experiments. The data were initially fit with Boltzmann functions where all parameters were allowed to vary, yielding a mean equivalent charge <z> = 0.59 ± 0.03 e (mean ± SEM, n = 10). The Q–Vs were then refit with z = <z> and normalized as shown in Fig. 4 A1. Although the individual plots are reasonably fit using identical values of z, they are scattered in their position along the voltage axis, similar to the mSlo GK–V relationships (Horrigan et al. 1999). To compare the shapes of the Q–Vs, the individual records were aligned as shown in Fig. 4 A2 (open symbols) by shifting them along the voltage axis by ΔV = 〈Vh〉 −Vh where Vh is the half-activation voltage of an individual Q–V and 〈Vh〉 is the mean (155 ± 6.5 mV, n = 10) determined from Fig. 4 A1. These voltage-shifted plots were then used to determine the average Q–V (Fig. 4 A2, filled symbols). A Boltzmann function with z = 0.59 e and Vh = 155 is superimposed on the data (solid line).


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)

Voltage dependence and kinetics of fast charge movement. (A1) The normalized Qfast–V relationships for many experiments are fit with Boltzmann functions (z = 0.59 e, dashed lines). The solid line is a Boltzmann function indicating the mean half-activation voltage (〈Vh〉 = 155 mV, z = 0.59 e). (A2) The data from A1 (open symbols) are aligned by shifting them along the voltage axis by ΔVh = (〈Vh〉 − Vh). The mean Qfast–V (filled circles, mean ± SEM) is superimposed on the data together with two Boltzmann fits (Vh = 155 mV; solid line: z = 0.59e, dashed line: z = 0.55 e) and was determined by averaging the shifted data in 15-mV bins. (B1) Time constants of fast Ig relaxation (τgFast) were determined from exponential fits to ON and OFF currents for the experiments in A and are plotted on a log scale versus voltage. (C1) Three τgFast–V relationships from B1 that cover a large voltage range are compared. (B2 and C2) Data from B1 and C1 were shifted along the voltage axis by ΔVh (determined from A) and then normalized to the mean τgFast measured from +100 to +180 mV (59 μs). The solid line in B2 indicates the best fit of a two-state model of voltage-sensor activation where the relationship between the forward (α) and backward (β) rates are constrained such that J = α/β = 1 at +155 mV (zα = +0.30 e, zβ = −0.21 e, α(0) = 1,310 s−1, β(0) = 30,160 s−1). Dashed lines in A2, B2, and C2 represent the parameters ultimately used in the allosteric model to describe closed-state charge movement (zα = +0.33 e, zβ = −0.22 e, α(0) = 1,100 s−1, β(0) = 32,120 s−1).
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Figure 4: Voltage dependence and kinetics of fast charge movement. (A1) The normalized Qfast–V relationships for many experiments are fit with Boltzmann functions (z = 0.59 e, dashed lines). The solid line is a Boltzmann function indicating the mean half-activation voltage (〈Vh〉 = 155 mV, z = 0.59 e). (A2) The data from A1 (open symbols) are aligned by shifting them along the voltage axis by ΔVh = (〈Vh〉 − Vh). The mean Qfast–V (filled circles, mean ± SEM) is superimposed on the data together with two Boltzmann fits (Vh = 155 mV; solid line: z = 0.59e, dashed line: z = 0.55 e) and was determined by averaging the shifted data in 15-mV bins. (B1) Time constants of fast Ig relaxation (τgFast) were determined from exponential fits to ON and OFF currents for the experiments in A and are plotted on a log scale versus voltage. (C1) Three τgFast–V relationships from B1 that cover a large voltage range are compared. (B2 and C2) Data from B1 and C1 were shifted along the voltage axis by ΔVh (determined from A) and then normalized to the mean τgFast measured from +100 to +180 mV (59 μs). The solid line in B2 indicates the best fit of a two-state model of voltage-sensor activation where the relationship between the forward (α) and backward (β) rates are constrained such that J = α/β = 1 at +155 mV (zα = +0.30 e, zβ = −0.21 e, α(0) = 1,310 s−1, β(0) = 30,160 s−1). Dashed lines in A2, B2, and C2 represent the parameters ultimately used in the allosteric model to describe closed-state charge movement (zα = +0.33 e, zβ = −0.22 e, α(0) = 1,100 s−1, β(0) = 32,120 s−1).
Mentions: Fig. 4 A1 plots the normalized Qfast–V relationships for many experiments. The data were initially fit with Boltzmann functions where all parameters were allowed to vary, yielding a mean equivalent charge <z> = 0.59 ± 0.03 e (mean ± SEM, n = 10). The Q–Vs were then refit with z = <z> and normalized as shown in Fig. 4 A1. Although the individual plots are reasonably fit using identical values of z, they are scattered in their position along the voltage axis, similar to the mSlo GK–V relationships (Horrigan et al. 1999). To compare the shapes of the Q–Vs, the individual records were aligned as shown in Fig. 4 A2 (open symbols) by shifting them along the voltage axis by ΔV = 〈Vh〉 −Vh where Vh is the half-activation voltage of an individual Q–V and 〈Vh〉 is the mean (155 ± 6.5 mV, n = 10) determined from Fig. 4 A1. These voltage-shifted plots were then used to determine the average Q–V (Fig. 4 A2, filled symbols). A Boltzmann function with z = 0.59 e and Vh = 155 is superimposed on the data (solid line).

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