It has been shown that β auxiliary subunits increase current amplitude in voltage-dependent calcium channels. In this study, however, we found a novel inhibitory effect of β3 subunit on macroscopic Ba2+ currents through recombinant N- and R-type calcium channels expressed in Xenopus oocytes. Overexpressed β3 (12.5 ng/cell cRNA) significantly suppressed N- and R-type, but not L-type, calcium channel currents at “physiological” holding potentials (HPs) of −60 and −80 mV. At a HP of −80 mV, coinjection of various concentrations (0–12.5 ng) of the β3 with Cav2.2α1 and α2δ enhanced the maximum conductance of expressed channels at lower β3 concentrations but at higher concentrations (>2.5 ng/cell) caused a marked inhibition. The β3-induced current suppression was reversed at a HP of −120 mV, suggesting that the inhibition was voltage dependent. A high concentration of Ba2+ (40 mM) as a charge carrier also largely diminished the effect of β3 at −80 mV. Therefore, experimental conditions (HP, divalent cation concentration, and β3 subunit concentration) approaching normal physiological conditions were critical to elucidate the full extent of this novel β3 effect. Steady-state inactivation curves revealed that N-type channels exhibited “closed-state” inactivation without β3, and that β3 caused an ∼40-mV negative shift of the inactivation, producing a second component with an inactivation midpoint of approximately −85 mV. The inactivation of N-type channels in the presence of a high concentration (12.5 ng/cell) of β3 developed slowly and the time-dependent inactivation curve was best fit by the sum of two exponential functions with time constants of 14 s and 8.8 min at −80 mV. Similar “ultra-slow” inactivation was observed for N-type channels without β3. Thus, β3 can have a profound negative regulatory effect on N-type (and also R-type) calcium channels by causing a hyperpolarizing shift of the inactivation without affecting “ultra-slow” and “closed-state” inactivation properties.
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1 April 2004
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March 15 2004
Overexpressed Cavβ3 Inhibits N-type (Cav2.2) Calcium Channel Currents through a Hyperpolarizing Shift of “Ultra-slow” and “Closed-state” Inactivation
Takahiro Yasuda,
Takahiro Yasuda
1School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia
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Richard J. Lewis,
Richard J. Lewis
1School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia
2Institute for Molecular Bioscience, The University of Queensland, Queensland 4072, Australia
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David J. Adams
David J. Adams
1School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia
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Takahiro Yasuda
1School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia
Richard J. Lewis
1School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia
2Institute for Molecular Bioscience, The University of Queensland, Queensland 4072, Australia
David J. Adams
1School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia
Address correspondence to David J. Adams School of Biomedical Sciences, The University of Queensland, Queensland 4072, Australia. Fax: (07) 3365-4933; email: [email protected]
Abbreviations used in this paper: AID, α1 subunit interaction domain; HP, holding potential; HVI, high-voltage inactivation; LVI, low-voltage inactivation; VDCC, voltage-dependent calcium channel.
Received:
October 28 2003
Accepted:
February 13 2004
Online ISSN: 1540-7748
Print ISSN: 0022-1295
The Rockefeller University Press
2004
J Gen Physiol (2004) 123 (4): 401–416.
Article history
Received:
October 28 2003
Accepted:
February 13 2004
Citation
Takahiro Yasuda, Richard J. Lewis, David J. Adams; Overexpressed Cavβ3 Inhibits N-type (Cav2.2) Calcium Channel Currents through a Hyperpolarizing Shift of “Ultra-slow” and “Closed-state” Inactivation . J Gen Physiol 1 April 2004; 123 (4): 401–416. doi: https://doi.org/10.1085/jgp.200308967
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