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State-dependent inactivation of the alpha1G T-type calcium channel.

Serrano JR, Perez-Reyes E, Jones SW - J. Gen. Physiol. (1999)

Bottom Line: Recovery was similar after 60-ms steps to -20 mV or 600-ms steps to -70 mV, suggesting rapid equilibration of open- and closed-state inactivation.The results are well described by a kinetic model where inactivation is allosterically coupled to the movement of the first three voltage sensors to activate.One consequence of state-dependent inactivation is that alpha1G channels continue to inactivate after repolarization, primarily from the open state, which leads to cumulative inactivation during repetitive pulses.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106, USA.

ABSTRACT
We have examined the kinetics of whole-cell T-current in HEK 293 cells stably expressing the alpha1G channel, with symmetrical Na(+)(i) and Na(+)(o) and 2 mM Ca(2+)(o). After brief strong depolarization to activate the channels (2 ms at +60 mV; holding potential -100 mV), currents relaxed exponentially at all voltages. The time constant of the relaxation was exponentially voltage dependent from -120 to -70 mV (e-fold for 31 mV; tau = 2.5 ms at -100 mV), but tau = 12-17 ms from-40 to +60 mV. This suggests a mixture of voltage-dependent deactivation (dominating at very negative voltages) and nearly voltage-independent inactivation. Inactivation measured by test pulses following that protocol was consistent with open-state inactivation. During depolarizations lasting 100-300 ms, inactivation was strong but incomplete (approximately 98%). Inactivation was also produced by long, weak depolarizations (tau = 220 ms at -80 mV; V(1/2) = -82 mV), which could not be explained by voltage-independent inactivation exclusively from the open state. Recovery from inactivation was exponential and fast (tau = 85 ms at -100 mV), but weakly voltage dependent. Recovery was similar after 60-ms steps to -20 mV or 600-ms steps to -70 mV, suggesting rapid equilibration of open- and closed-state inactivation. There was little current at -100 mV during recovery from inactivation, consistent with

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Recovery from closed-state inactivation. In this cell, a 600-ms step to −70 mV produced a small inward current , but a subsequent 10-ms test pulse to −20 mV demonstrated 40% inactivation (compared with 8% predicted open-state inactivation). The main figure shows the time course of recovery from inactivation at −100 mV, fitted to an exponential function with . Currents were measured during the steps to −20 mV, and were normalized to the current recorded during test pulses to −20 mV (with no prepulse to −70 mV) given after the protocol. The inset shows records for recovery intervals of 25, 50, 100, 250, 500, and 1,000 ms (cell d8o28, 500 Hz Gaussian filter). In this cell, the peak PO,r and the amount of inactivation at −70 mV were both less than typically observed (compare with Fig. 6, Fig. 7, and Fig. 9).
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Figure 10: Recovery from closed-state inactivation. In this cell, a 600-ms step to −70 mV produced a small inward current , but a subsequent 10-ms test pulse to −20 mV demonstrated 40% inactivation (compared with 8% predicted open-state inactivation). The main figure shows the time course of recovery from inactivation at −100 mV, fitted to an exponential function with . Currents were measured during the steps to −20 mV, and were normalized to the current recorded during test pulses to −20 mV (with no prepulse to −70 mV) given after the protocol. The inset shows records for recovery intervals of 25, 50, 100, 250, 500, and 1,000 ms (cell d8o28, 500 Hz Gaussian filter). In this cell, the peak PO,r and the amount of inactivation at −70 mV were both less than typically observed (compare with Fig. 6, Fig. 7, and Fig. 9).

Mentions: To determine whether inactivation from closed states is a fundamentally different kinetic process from open-state inactivation, we examined recovery from inactivation after 600-ms steps to −70 mV (Fig. 10). Recovery from inactivation was similar, whether inactivation was produced primarily from open or closed states (Table ). Notably, there was little voltage dependence to recovery (varying approximately twofold from −120 to −80 mV), and recovery could be quite rapid (τ ∼ 100 ms at −120 mV). These results suggest that the inactivated states reached from open and closed states interconvert rapidly. Alternatively, it is possible that a single inactivated state is accessed from both open and closed states.


State-dependent inactivation of the alpha1G T-type calcium channel.

Serrano JR, Perez-Reyes E, Jones SW - J. Gen. Physiol. (1999)

Recovery from closed-state inactivation. In this cell, a 600-ms step to −70 mV produced a small inward current , but a subsequent 10-ms test pulse to −20 mV demonstrated 40% inactivation (compared with 8% predicted open-state inactivation). The main figure shows the time course of recovery from inactivation at −100 mV, fitted to an exponential function with . Currents were measured during the steps to −20 mV, and were normalized to the current recorded during test pulses to −20 mV (with no prepulse to −70 mV) given after the protocol. The inset shows records for recovery intervals of 25, 50, 100, 250, 500, and 1,000 ms (cell d8o28, 500 Hz Gaussian filter). In this cell, the peak PO,r and the amount of inactivation at −70 mV were both less than typically observed (compare with Fig. 6, Fig. 7, and Fig. 9).
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC2230639&req=5

Figure 10: Recovery from closed-state inactivation. In this cell, a 600-ms step to −70 mV produced a small inward current , but a subsequent 10-ms test pulse to −20 mV demonstrated 40% inactivation (compared with 8% predicted open-state inactivation). The main figure shows the time course of recovery from inactivation at −100 mV, fitted to an exponential function with . Currents were measured during the steps to −20 mV, and were normalized to the current recorded during test pulses to −20 mV (with no prepulse to −70 mV) given after the protocol. The inset shows records for recovery intervals of 25, 50, 100, 250, 500, and 1,000 ms (cell d8o28, 500 Hz Gaussian filter). In this cell, the peak PO,r and the amount of inactivation at −70 mV were both less than typically observed (compare with Fig. 6, Fig. 7, and Fig. 9).
Mentions: To determine whether inactivation from closed states is a fundamentally different kinetic process from open-state inactivation, we examined recovery from inactivation after 600-ms steps to −70 mV (Fig. 10). Recovery from inactivation was similar, whether inactivation was produced primarily from open or closed states (Table ). Notably, there was little voltage dependence to recovery (varying approximately twofold from −120 to −80 mV), and recovery could be quite rapid (τ ∼ 100 ms at −120 mV). These results suggest that the inactivated states reached from open and closed states interconvert rapidly. Alternatively, it is possible that a single inactivated state is accessed from both open and closed states.

Bottom Line: Recovery was similar after 60-ms steps to -20 mV or 600-ms steps to -70 mV, suggesting rapid equilibration of open- and closed-state inactivation.The results are well described by a kinetic model where inactivation is allosterically coupled to the movement of the first three voltage sensors to activate.One consequence of state-dependent inactivation is that alpha1G channels continue to inactivate after repolarization, primarily from the open state, which leads to cumulative inactivation during repetitive pulses.

View Article: PubMed Central - PubMed

Affiliation: Department of Physiology and Biophysics, Case Western Reserve University, Cleveland, Ohio 44106, USA.

ABSTRACT
We have examined the kinetics of whole-cell T-current in HEK 293 cells stably expressing the alpha1G channel, with symmetrical Na(+)(i) and Na(+)(o) and 2 mM Ca(2+)(o). After brief strong depolarization to activate the channels (2 ms at +60 mV; holding potential -100 mV), currents relaxed exponentially at all voltages. The time constant of the relaxation was exponentially voltage dependent from -120 to -70 mV (e-fold for 31 mV; tau = 2.5 ms at -100 mV), but tau = 12-17 ms from-40 to +60 mV. This suggests a mixture of voltage-dependent deactivation (dominating at very negative voltages) and nearly voltage-independent inactivation. Inactivation measured by test pulses following that protocol was consistent with open-state inactivation. During depolarizations lasting 100-300 ms, inactivation was strong but incomplete (approximately 98%). Inactivation was also produced by long, weak depolarizations (tau = 220 ms at -80 mV; V(1/2) = -82 mV), which could not be explained by voltage-independent inactivation exclusively from the open state. Recovery from inactivation was exponential and fast (tau = 85 ms at -100 mV), but weakly voltage dependent. Recovery was similar after 60-ms steps to -20 mV or 600-ms steps to -70 mV, suggesting rapid equilibration of open- and closed-state inactivation. There was little current at -100 mV during recovery from inactivation, consistent with

Show MeSH
Related in: MedlinePlus