Cloning, distribution and functional analysis of the type III sodium channel from human brain

Chen, Yu Hua; Dale, Timothy; Romanos, Michael A.; Whitaker, William R. J.; Xie, Xinmin; Clare, Jeffrey J.

doi:10.1046/j.1460-9568.2000.01336.x

Cited by 36 publications

(48 citation statements)

References 36 publications

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“…SCN3A is highly expressed during early development in humans and rodents, with expression declining into adulthood (Black et al, 1994; Chen et al, 2000; Felts et al, 1997; Whitaker et al, 2001). However, the relative reduction in adult Scn3a levels appears to be greater in the mouse.…”

Section: Discussionmentioning

confidence: 99%

SCN3A deficiency associated with increased seizure susceptibility

Lamar

Vanoye

Calhoun

et al. 2017

Neurobiology of Disease

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Mutations in voltage-gated sodium channels expressed highly in the brain (SCN1A, SCN2A, SCN3A, and SCN8A) are responsible for an increasing number of epilepsy syndromes. In particular, mutations in the SCN3A gene, encoding the pore-forming Nav1.3 α subunit, have been identified in patients with focal epilepsy. Biophysical characterization of epilepsy-associated SCN3A variants suggests that both gain- and loss-of-function SCN3A mutations may lead to increased seizure susceptibility. In this report, we identified a novel SCN3A variant (L247P) by whole exome sequencing of a child with focal epilepsy, developmental delay, and autonomic nervous system dysfunction. Voltage clamp analysis showed no detectable sodium current in a heterologous expression system expressing the SCN3A-L247P variant. Furthermore, cell surface biotinylation demonstrated a reduction in the amount of SCN3A-L247P at the cell surface, suggesting the SCN3A-L247P variant is a trafficking-deficient mutant. To further explore the possible clinical consequences of reduced SCN3A activity, we investigated the effect of a hypomorphic Scn3a allele (Scn3aHyp) on seizure susceptibility and behavior using a gene trap mouse line. Heterozygous Scn3a mutant mice (Scn3a+/Hyp) did not exhibit spontaneous seizures nor were they susceptible to hyperthermia-induced seizures. However, they displayed increased susceptibility to electroconvulsive (6 Hz) and chemiconvulsive (flurothyl and kainic acid) induced seizures. Scn3a+/Hyp mice also exhibited deficits in locomotor activity and motor learning. Taken together, these results provide evidence that loss-of-function of SCN3A caused by reduced protein expression or deficient trafficking to the plasma membrane may contribute to increased seizure susceptibility.

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Section: Discussionmentioning

confidence: 99%

SCN3A deficiency associated with increased seizure susceptibility

Lamar

Vanoye

Calhoun

et al. 2017

Neurobiology of Disease

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“…Na v 1.1 and Na v 1.2 have been proposed to conduct a persistent Na current and are modulated by G-proteins [26]. Na v 1.3 has also been shown to produce a persistent component [14]. The cloning and functional expression of Na v 1.6 raise the prospect that this channel may also carry the delayed sodium current.…”

Section: Discussionmentioning

confidence: 99%

Biophysical characterisation of the persistent sodium current of the Nav1.6 neuronal sodium channel: a single-channel analysis

Chatelier

Zhao

Bois

et al. 2010

Pflugers Arch - Eur J Physiol

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Na(v)1.6 is the major voltage-gated sodium channel at nodes of Ranvier. This channel has been shown to produce a robust persistent inward current in whole-cell experiments. Na(v)1.6 plays an important role in axonal conduction and may significantly contribute to the pathophysiology of the injured nervous system through this persistent current. However, the underlying molecular mechanisms and regulation of the persistent current are not well understood. Using the whole-cell configuration of the patch-clamp technique, we investigated the Na(v)1.6 transient and persistent currents in HEK-293. Previous studies have shown that the persistent current depended on the content of the patch electrode. Therefore, we characterised the single-channel properties of the persistent current with an intact intracellular medium using the cell-attached configuration of the patch-clamp technique. In HEK-293 cells, the Na(v)1.6 persistent current recorded in the whole-cell configuration was 3-5% of the peak transient current. In single-channel recording, the ratio between peak and persistent open probability confirmed the magnitude of the persistent current observed in the whole-cell configuration. The cell-attached configuration revealed that the molecular mechanism of the whole-cell persistent current is a consequence of single Na(v)1.6 channels reopening.

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“…Cloning and construction of a stable cell line Regions of the human brain types I, IIA and III Na + channel α subunit cDNAs were amplified by reverse transcriptase polymerase chain reaction (RT-PCR) using consensus primers (see [6]) based on the corresponding rat sequences. A mixed probe consisting of these PCR products was then used for hybridization screening of a cDNA library derived from human cerebellum (a gift from Professor I Charles, UCL, London).…”

Section: Methodsmentioning

confidence: 99%

Electrophysiological and pharmacological properties of the human brain type IIA Na + channel expressed in a stable mammalian cell line

Xie¹,

Dale²,

John³

et al. 2001

Pfl�gers Archiv European Journal of Physiology

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The human brain voltage-gated Na+ channel type IIA alpha subunit was cloned and stably expressed in Chinese hamster ovary cells and its biophysical and pharmacological properties were studied using whole-cell voltage-clamp. Fast, transient inward currents of up to -8,000 pA were elicited by membrane depolarization of the recombinant cells. Channels activated at -50 mV and reached maximal activation at -10 mV to 0 mV. The reversal potential was 62 +/- 2 mV which is close to the Na+ equilibrium potential. The half-maximal activation and inactivation voltages were -24 +/- 2 mV and -63 +/- 1 mV, respectively. Currents were reversibly blocked by tetrodotoxin with a half-maximal inhibition of 13 nM. The effects of four commonly used anti-convulsant drugs were examined for the first time on the cloned human type IIA channel. Lamotrigine and phenytoin produced concentration- and voltage-dependent inhibition of the type IIA currents, whereas, sodium valproate and gabapentin (up to 1 mM) had no effect. These results indicate that recombinant human type IIA Na+ channels conduct tetrodotoxin-sensitive Na+ currents with similar properties to those observed in recombinant rat brain type IIA and native rat brain Na+ channels. This stable cell line should provide a useful tool for more detailed characterization of therapeutic modulators of human Na+ channels.

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Cloning, distribution and functional analysis of the type III sodium channel from human brain

Cited by 36 publications

References 36 publications

SCN3A deficiency associated with increased seizure susceptibility

SCN3A deficiency associated with increased seizure susceptibility

Biophysical characterisation of the persistent sodium current of the Nav1.6 neuronal sodium channel: a single-channel analysis

Electrophysiological and pharmacological properties of the human brain type IIA Na + channel expressed in a stable mammalian cell line

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