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Modal gating of human CaV2.1 (P/Q-type) calcium channels: II. the b mode and reversible uncoupling of inactivation.

Fellin T, Luvisetto S, Spagnolo M, Pietrobon D - J. Gen. Physiol. (2004)

Bottom Line: Physiol. 124:445-461).In fact, a CaV2.1 channel in the b gating mode does not inactivate during long pulses at high positive voltages, where the same channel in both fast-nb and slow-nb gating modes inactivates relatively rapidly.Moreover, a CaV2.1 channel in the b gating mode shows a larger availability to open than in the nb gating modes.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Biomedical Sciences, University of Padova, Viale G. Colombo, 3 35121 Padova, Italy.

ABSTRACT
The single channel gating properties of human CaV2.1 (P/Q-type) calcium channels were investigated with cell-attached patch-clamp recordings on HEK293 cells stably expressing these calcium channels. Human CaV2.1 channels showed a complex modal gating, which is described in this and the preceding paper (Luvisetto, S., T. Fellin, M. Spagnolo, B. Hivert, P.F. Brust, M.M. Harpold, K.A. Stauderman, M.E. Williams, and D. Pietrobon. 2004. J. Gen. Physiol. 124:445-461). Here, we report the characterization of the so-called b gating mode. A CaV2.1 channel in the b gating mode shows a bell-shaped voltage dependence of the open probability, and a characteristic low open probability at high positive voltages, that decreases with increasing voltage, as a consequence of both shorter mean open time and longer mean closed time. Reversible transitions of single human CaV2.1 channels between the b gating mode and the mode of gating in which the channel shows the usual voltage dependence of the open probability (nb gating mode) were much more frequent (time scale of seconds) than those between the slow and fast gating modes (time scale of minutes; Luvisetto et al., 2004), and occurred independently of whether the channel was in the fast or slow mode. We show that the b gating mode produces reversible uncoupling of inactivation in human CaV2.1 channels. In fact, a CaV2.1 channel in the b gating mode does not inactivate during long pulses at high positive voltages, where the same channel in both fast-nb and slow-nb gating modes inactivates relatively rapidly. Moreover, a CaV2.1 channel in the b gating mode shows a larger availability to open than in the nb gating modes. Regulation of the complex modal gating of human CaV2.1 channels could be a potent and versatile mechanism for the modulation of synaptic strength and plasticity as well as of neuronal excitability and other postsynaptic Ca2+-dependent processes.

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Reversible switching of a single human CaV2.1 channel to a noninactivating mode of gating with low open probability at +40 and +50 mV: the b mode. Cell-attached single channel patch-clamp recordings, with 90 mM Ba2+ as charge carrier, from HEK293 cells stably coexpressing human α1A-2, β1b, and α2bδ-1 subunits. Consecutive traces at +40 mV (left) and +50 mV (right) from a patch containing a single CaV2.1 channel are shown. Labeling of traces: b indicates channel activity with low open probability (b gating mode); nb indicates channel activity with the high open probability expected during periods of activity at high voltages (nb gating mode); N indicates lack of activity (). Depolarizations were 720 ms long and were delivered every 4 s from holding potentials of −80 mV. Records were sampled and filtered at 5 and 1 kHz, respectively.
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fig1: Reversible switching of a single human CaV2.1 channel to a noninactivating mode of gating with low open probability at +40 and +50 mV: the b mode. Cell-attached single channel patch-clamp recordings, with 90 mM Ba2+ as charge carrier, from HEK293 cells stably coexpressing human α1A-2, β1b, and α2bδ-1 subunits. Consecutive traces at +40 mV (left) and +50 mV (right) from a patch containing a single CaV2.1 channel are shown. Labeling of traces: b indicates channel activity with low open probability (b gating mode); nb indicates channel activity with the high open probability expected during periods of activity at high voltages (nb gating mode); N indicates lack of activity (). Depolarizations were 720 ms long and were delivered every 4 s from holding potentials of −80 mV. Records were sampled and filtered at 5 and 1 kHz, respectively.

Mentions: Single channel recordings at +40 and +50 mV (with 90 mM Ba2+ as charge carrier) on HEK293 cells, stably coexpressing human CaV2.1α1 (α1A-2), β1b, and α2bδ-1 subunits, revealed that a CaV2.1 channel in either the fast or slow gating mode (Luvisetto et al., 2004) can reversibly and frequently (in the time scale of seconds) switch to a noninactivating gating mode with low open probability, po, that we have called b mode. Fig. 1 shows consecutive traces at +40 and +50 mV from a patch containing a single CaV2.1 channel that alternates between an inactivating mode of gating with the high open probability expected during periods of activity at these high voltages (traces labeled nb) and a noninactivating gating mode with lower po, the b mode (traces labeled b). The sixth trace at +50 mV shows a transition to the b mode during the depolarization. Here, and in the following, po refers to the open probability during periods of channel activity; it is calculated excluding the last shut time due to channel entering into an inactivated state, and therefore reflects the equilibrium between short-lived open and closed states.


Modal gating of human CaV2.1 (P/Q-type) calcium channels: II. the b mode and reversible uncoupling of inactivation.

Fellin T, Luvisetto S, Spagnolo M, Pietrobon D - J. Gen. Physiol. (2004)

Reversible switching of a single human CaV2.1 channel to a noninactivating mode of gating with low open probability at +40 and +50 mV: the b mode. Cell-attached single channel patch-clamp recordings, with 90 mM Ba2+ as charge carrier, from HEK293 cells stably coexpressing human α1A-2, β1b, and α2bδ-1 subunits. Consecutive traces at +40 mV (left) and +50 mV (right) from a patch containing a single CaV2.1 channel are shown. Labeling of traces: b indicates channel activity with low open probability (b gating mode); nb indicates channel activity with the high open probability expected during periods of activity at high voltages (nb gating mode); N indicates lack of activity (). Depolarizations were 720 ms long and were delivered every 4 s from holding potentials of −80 mV. Records were sampled and filtered at 5 and 1 kHz, respectively.
© Copyright Policy
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC2234006&req=5

fig1: Reversible switching of a single human CaV2.1 channel to a noninactivating mode of gating with low open probability at +40 and +50 mV: the b mode. Cell-attached single channel patch-clamp recordings, with 90 mM Ba2+ as charge carrier, from HEK293 cells stably coexpressing human α1A-2, β1b, and α2bδ-1 subunits. Consecutive traces at +40 mV (left) and +50 mV (right) from a patch containing a single CaV2.1 channel are shown. Labeling of traces: b indicates channel activity with low open probability (b gating mode); nb indicates channel activity with the high open probability expected during periods of activity at high voltages (nb gating mode); N indicates lack of activity (). Depolarizations were 720 ms long and were delivered every 4 s from holding potentials of −80 mV. Records were sampled and filtered at 5 and 1 kHz, respectively.
Mentions: Single channel recordings at +40 and +50 mV (with 90 mM Ba2+ as charge carrier) on HEK293 cells, stably coexpressing human CaV2.1α1 (α1A-2), β1b, and α2bδ-1 subunits, revealed that a CaV2.1 channel in either the fast or slow gating mode (Luvisetto et al., 2004) can reversibly and frequently (in the time scale of seconds) switch to a noninactivating gating mode with low open probability, po, that we have called b mode. Fig. 1 shows consecutive traces at +40 and +50 mV from a patch containing a single CaV2.1 channel that alternates between an inactivating mode of gating with the high open probability expected during periods of activity at these high voltages (traces labeled nb) and a noninactivating gating mode with lower po, the b mode (traces labeled b). The sixth trace at +50 mV shows a transition to the b mode during the depolarization. Here, and in the following, po refers to the open probability during periods of channel activity; it is calculated excluding the last shut time due to channel entering into an inactivated state, and therefore reflects the equilibrium between short-lived open and closed states.

Bottom Line: Physiol. 124:445-461).In fact, a CaV2.1 channel in the b gating mode does not inactivate during long pulses at high positive voltages, where the same channel in both fast-nb and slow-nb gating modes inactivates relatively rapidly.Moreover, a CaV2.1 channel in the b gating mode shows a larger availability to open than in the nb gating modes.

View Article: PubMed Central - PubMed

Affiliation: Dept. of Biomedical Sciences, University of Padova, Viale G. Colombo, 3 35121 Padova, Italy.

ABSTRACT
The single channel gating properties of human CaV2.1 (P/Q-type) calcium channels were investigated with cell-attached patch-clamp recordings on HEK293 cells stably expressing these calcium channels. Human CaV2.1 channels showed a complex modal gating, which is described in this and the preceding paper (Luvisetto, S., T. Fellin, M. Spagnolo, B. Hivert, P.F. Brust, M.M. Harpold, K.A. Stauderman, M.E. Williams, and D. Pietrobon. 2004. J. Gen. Physiol. 124:445-461). Here, we report the characterization of the so-called b gating mode. A CaV2.1 channel in the b gating mode shows a bell-shaped voltage dependence of the open probability, and a characteristic low open probability at high positive voltages, that decreases with increasing voltage, as a consequence of both shorter mean open time and longer mean closed time. Reversible transitions of single human CaV2.1 channels between the b gating mode and the mode of gating in which the channel shows the usual voltage dependence of the open probability (nb gating mode) were much more frequent (time scale of seconds) than those between the slow and fast gating modes (time scale of minutes; Luvisetto et al., 2004), and occurred independently of whether the channel was in the fast or slow mode. We show that the b gating mode produces reversible uncoupling of inactivation in human CaV2.1 channels. In fact, a CaV2.1 channel in the b gating mode does not inactivate during long pulses at high positive voltages, where the same channel in both fast-nb and slow-nb gating modes inactivates relatively rapidly. Moreover, a CaV2.1 channel in the b gating mode shows a larger availability to open than in the nb gating modes. Regulation of the complex modal gating of human CaV2.1 channels could be a potent and versatile mechanism for the modulation of synaptic strength and plasticity as well as of neuronal excitability and other postsynaptic Ca2+-dependent processes.

Show MeSH
Related in: MedlinePlus