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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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Time course of inactivation and recovery. (A) Effect of prepulses of variable voltage and duration on the current evoked by a test pulse to −20 mV. The currents were normalized to that observed during a test pulse given alone (with no prepulse). The smooth curves are exponential functions, with time constants of 202 ms at −80 mV, and 210 ms at −70 mV. (B) The time course of recovery from inactivation, after 60-ms pulses to −20 mV. The values are the current during the test pulse to −20 mV, divided by the current at the corresponding time in the 60-ms prepulse. Time constants for recovery were 86 ms at −120 mV, 74 ms at −100 mV, and 175 ms at −80 mV. (C) The protocol for recovery from inactivation. Records (analogue filtering at 1 kHz) are shown for recovery at the holding potential of −100 mV, with recovery intervals of 8, 20, 40, 80, and 200 ms. All data in this figure are from cell a8612.
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Figure 6: Time course of inactivation and recovery. (A) Effect of prepulses of variable voltage and duration on the current evoked by a test pulse to −20 mV. The currents were normalized to that observed during a test pulse given alone (with no prepulse). The smooth curves are exponential functions, with time constants of 202 ms at −80 mV, and 210 ms at −70 mV. (B) The time course of recovery from inactivation, after 60-ms pulses to −20 mV. The values are the current during the test pulse to −20 mV, divided by the current at the corresponding time in the 60-ms prepulse. Time constants for recovery were 86 ms at −120 mV, 74 ms at −100 mV, and 175 ms at −80 mV. (C) The protocol for recovery from inactivation. Records (analogue filtering at 1 kHz) are shown for recovery at the holding potential of −100 mV, with recovery intervals of 8, 20, 40, 80, and 200 ms. All data in this figure are from cell a8612.

Mentions: Substantial inactivation was observed at voltages as negative as −80 mV (Fig. 6 A). Pulses to −120 mV had little effect, implying that there is little resting fast inactivation at our holding potential of −100 mV. At −80 mV, inactivation proceeded with , and was 70 ± 5% complete . Inactivation was nearly complete at −70 mV .


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

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

Time course of inactivation and recovery. (A) Effect of prepulses of variable voltage and duration on the current evoked by a test pulse to −20 mV. The currents were normalized to that observed during a test pulse given alone (with no prepulse). The smooth curves are exponential functions, with time constants of 202 ms at −80 mV, and 210 ms at −70 mV. (B) The time course of recovery from inactivation, after 60-ms pulses to −20 mV. The values are the current during the test pulse to −20 mV, divided by the current at the corresponding time in the 60-ms prepulse. Time constants for recovery were 86 ms at −120 mV, 74 ms at −100 mV, and 175 ms at −80 mV. (C) The protocol for recovery from inactivation. Records (analogue filtering at 1 kHz) are shown for recovery at the holding potential of −100 mV, with recovery intervals of 8, 20, 40, 80, and 200 ms. All data in this figure are from cell a8612.
© Copyright Policy
Related In: Results  -  Collection

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

Figure 6: Time course of inactivation and recovery. (A) Effect of prepulses of variable voltage and duration on the current evoked by a test pulse to −20 mV. The currents were normalized to that observed during a test pulse given alone (with no prepulse). The smooth curves are exponential functions, with time constants of 202 ms at −80 mV, and 210 ms at −70 mV. (B) The time course of recovery from inactivation, after 60-ms pulses to −20 mV. The values are the current during the test pulse to −20 mV, divided by the current at the corresponding time in the 60-ms prepulse. Time constants for recovery were 86 ms at −120 mV, 74 ms at −100 mV, and 175 ms at −80 mV. (C) The protocol for recovery from inactivation. Records (analogue filtering at 1 kHz) are shown for recovery at the holding potential of −100 mV, with recovery intervals of 8, 20, 40, 80, and 200 ms. All data in this figure are from cell a8612.
Mentions: Substantial inactivation was observed at voltages as negative as −80 mV (Fig. 6 A). Pulses to −120 mV had little effect, implying that there is little resting fast inactivation at our holding potential of −100 mV. At −80 mV, inactivation proceeded with , and was 70 ± 5% complete . Inactivation was nearly complete at −70 mV .

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