Click here to close
Hello! We notice that you are using Internet Explorer, which is not supported by Xenbase and may cause the site to display incorrectly.
We suggest using a current version of Chrome,
FireFox, or Safari.
PLoS One
2014 Jan 03;91:e85188. doi: 10.1371/journal.pone.0085188.
Show Gene links
Show Anatomy links
Functional expression of Rat Nav1.6 voltage-gated sodium channels in HEK293 cells: modulation by the auxiliary β1 subunit.
He B
,
Soderlund DM
.
Abstract
The Nav1.6 voltage-gated sodium channel α subunit isoform is abundantly expressed in the adult rat brain. To assess the functional modulation of Nav1.6 channels by the auxiliary β1 subunit we expressed the rat Nav1.6 sodium channel α subunit by stable transformation in HEK293 cells either alone or in combination with the rat β1 subunit and assessed the properties of the reconstituted channels by recording sodium currents using the whole-cell patch clamp technique. Coexpression with the β1 subunit accelerated the inactivation of sodium currents and shifted the voltage dependence of channel activation and steady-state fast inactivation by approximately 5-7 mV in the direction of depolarization. By contrast the β1 subunit had no effect on the stability of sodium currents following repeated depolarizations at high frequencies. Our results define modulatory effects of the β1 subunit on the properties of rat Nav1.6-mediated sodium currents reconstituted in HEK293 cells that differ from effects measured previously in the Xenopus oocyte expression system. We also identify differences in the kinetic and gating properties of the rat Nav1.6 channel expressed in the absence of the β1 subunit compared to the properties of the orthologous mouse and human channels expressed in this system.
Figure 2. Voltage-dependent activation of Nav1.6 and Nav1.6β1 sodium channels expressed in HEK293 cells.(A) Representative current traces recorded from a HEK-Nav1.6 cell using the indicated pulse protocol (left) and the plot of peak sodium current in these traces as a function of test potential (right). (B) Representative current traces recorded from a HEK-Nav1.6β1 cell using the pulse protocol shown in Panel A and the plot of peak sodium current in these traces as a function of test potential. (C) Conductance – voltage plots for the activation of Nav1.6 and Nav1.6β1 channels. Peak sodium currents such as those in Panels A and B were transformed to conductances (G) using the equation G = I/(Vt–Vrev), where I is the peak current, Vrev is the reversal potential, and Vt is the voltage of the test potential; conductances were then normalized to the maximum conductance (Gmax) for that cell. Values are means of 64 (Nav1.6) or 65 (Nav1.6β1) separate experiments with different cells; bars show SE values larger than the data point symbols. Curves were fitted to the mean values using the Boltzmann equation.
Figure 3. Voltage-dependent steady-state fast inactivation of Nav1.6 and Nav1.6β1 sodium channels expressed in HEK293 cells.Amplitudes of peak transient currents obtained using the indicated pulse protocol are plotted as a function of prepulse potential. Values are means of 63 (Nav1.6) or 66 (Nav1.6β1) separate experiments with different cells; bars show SE values larger than the data point symbols. Curves were fitted to the mean values using the Boltzmann equation.
Figure 4. Effect of repeated depolarization on the stability of sodium currents recorded from HEK293 cells expressing Nav1.6 and Nav1.6β1 sodium channels.Sodium currents were recorded during a 40-ms step depolarization from −120 mV to −15 mV following 0–100 conditioning prepulses (5-ms pulses from −120 mV to 10 mV at 20 Hz). Currents for each cell were normalized to the amplitude of the peak current obtained prior to repeated depolarization. Values are means of the indicated number of separate experiments with different cells; bars show SE values larger than the data point symbols.
Figure 1. Sodium currents recorded from HEK-Nav1.6 and HEK-Nav1.6β1 cells.(A) Representative sodium current traces recorded from HEK-Nav1.6 and HEK-Nav1.6β1 cells following 40-ms depolarizations from −120 mV to −15 mV. (B) Sodium currents recorded from a HEK-Nav1.6β1 cell before and after exposure to 0.5 µM TTX. Dashed lines indicate zero current.
Auld,
A rat brain Na+ channel alpha subunit with novel gating properties.
1990, Pubmed,
Xenbase
Auld,
A rat brain Na+ channel alpha subunit with novel gating properties.
1990,
Pubmed
,
Xenbase
Bezanilla,
Inactivation of the sodium channel. I. Sodium current experiments.
1978,
Pubmed
Brackenbury,
Functional reciprocity between Na+ channel Nav1.6 and beta1 subunits in the coordinated regulation of excitability and neurite outgrowth.
2010,
Pubmed
Burbidge,
Molecular cloning, distribution and functional analysis of the NA(V)1.6. Voltage-gated sodium channel from human brain.
2002,
Pubmed
Burgess,
Mutation of a new sodium channel gene, Scn8a, in the mouse mutant 'motor endplate disease'.
1995,
Pubmed
Caldwell,
Sodium channel Na(v)1.6 is localized at nodes of ranvier, dendrites, and synapses.
2000,
Pubmed
Catterall,
From ionic currents to molecular mechanisms: the structure and function of voltage-gated sodium channels.
2000,
Pubmed
Chen,
Functional properties and differential neuromodulation of Na(v)1.6 channels.
2008,
Pubmed
Cummins,
Nav1.3 sodium channels: rapid repriming and slow closed-state inactivation display quantitative differences after expression in a mammalian cell line and in spinal sensory neurons.
2001,
Pubmed
,
Xenbase
Dietrich,
Functional analysis of a voltage-gated sodium channel and its splice variant from rat dorsal root ganglia.
1998,
Pubmed
,
Xenbase
Felts,
Sodium channel alpha-subunit mRNAs I, II, III, NaG, Na6 and hNE (PN1): different expression patterns in developing rat nervous system.
1997,
Pubmed
Ferrera,
Beta1-subunit modulates the Nav1.4 sodium channel by changing the surface charge.
2006,
Pubmed
Goldin,
Resurgence of sodium channel research.
2001,
Pubmed
Graham,
Characteristics of a human cell line transformed by DNA from human adenovirus type 5.
1977,
Pubmed
He,
Differential state-dependent modification of rat Na(v)1.6 sodium channels expressed in human embryonic kidney (HEK293) cells by the pyrethroid insecticides tefluthrin and deltamethrin.
2011,
Pubmed
,
Xenbase
He,
Human embryonic kidney (HEK293) cells express endogenous voltage-gated sodium currents and Na v 1.7 sodium channels.
2010,
Pubmed
Hu,
Distinct contributions of Na(v)1.6 and Na(v)1.2 in action potential initiation and backpropagation.
2009,
Pubmed
Khaliq,
The contribution of resurgent sodium current to high-frequency firing in Purkinje neurons: an experimental and modeling study.
2003,
Pubmed
Laezza,
FGF14 N-terminal splice variants differentially modulate Nav1.2 and Nav1.6-encoded sodium channels.
2009,
Pubmed
Moran,
Sodium channel heterologous expression in mammalian cells and the role of the endogenous beta1-subunits.
2002,
Pubmed
,
Xenbase
Moran,
Endogenous expression of the beta1A sodium channel subunit in HEK-293 cells.
2000,
Pubmed
,
Xenbase
Patino,
Electrophysiology and beyond: multiple roles of Na+ channel β subunits in development and disease.
2010,
Pubmed
Qu,
Differential modulation of sodium channel gating and persistent sodium currents by the beta1, beta2, and beta3 subunits.
2002,
Pubmed
Raman,
Inactivation and recovery of sodium currents in cerebellar Purkinje neurons: evidence for two mechanisms.
2001,
Pubmed
Schaller,
Expression and distribution of voltage-gated sodium channels in the cerebellum.
2003,
Pubmed
Shah,
Developmental expression of the novel voltage-gated sodium channel auxiliary subunit beta3, in rat CNS.
2001,
Pubmed
,
Xenbase
Shaw,
Preferential transformation of human neuronal cells by human adenoviruses and the origin of HEK 293 cells.
2002,
Pubmed
Tan,
Independent and joint modulation of rat Nav1.6 voltage-gated sodium channels by coexpression with the auxiliary β1 and β2 subunits.
2011,
Pubmed
,
Xenbase
Tan,
Coexpression with Auxiliary β Subunits Modulates the Action of Tefluthrin on Rat Na(v)1.6 and Na(v)1.3 Sodium Channels.
2012,
Pubmed
,
Xenbase
Thomas,
HEK293 cell line: a vehicle for the expression of recombinant proteins.
2005,
Pubmed
Welch,
Traditional AMPA receptor antagonists partially block Na v1.6-mediated persistent current.
2008,
Pubmed
Whitaker,
Comparative distribution of voltage-gated sodium channel proteins in human brain.
2001,
Pubmed
Whitaker,
Distribution of voltage-gated sodium channel alpha-subunit and beta-subunit mRNAs in human hippocampal formation, cortex, and cerebellum.
2000,
Pubmed
Yu,
Overview of the voltage-gated sodium channel family.
2003,
Pubmed
Zhao,
Regulation of Nav1.6 and Nav1.8 peripheral nerve Na+ channels by auxiliary β-subunits.
2011,
Pubmed
