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Title: Alternative splicing in sodium channels : biophysical and functional effects in NaV1.1, NaV1.2 & NaV1.7
Author: Liavas, A.
ISNI:       0000 0004 8503 1155
Awarding Body: UCL (University College London)
Current Institution: University College London (University of London)
Date of Award: 2015
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Alternative splicing in voltage-gated sodium channels can affect pathophysiological conditions, including epilepsy and pain. A conserved alternative splicing event in sodium channel genes, including SCN1A, SCN2A and SCN9A, gives rise to the neonatal (5N) and adult (5A) isoforms. Differences in the ratio of 5A/5N in Nav1.1 (encoded by SCN1A) in patients may lead to different predisposition to epilepsy or response to antiepileptic drugs (AED). Previous HEK293T whole-cell voltage-clamp recordings showed that Nav1.1-5N channels recover more quickly from fast inactivation than 5A. However it was unknown whether this effect is conserved in Nav1.2 (encoded by SCN2A) and Nav1.7 (SCN9A) channels, or what the functional consequences of this splicing event are for neurons. This project used whole-cell voltage-clamp recordings on heterologously expressed neonatal and adult channels to compare the biophysical properties of the splice isoforms for all three channel types and their modulation by AEDs. It also used current-clamp and dynamic-clamp recordings on transfected hippocampal cultured neurons to assess the effect of splicing on neuronal properties during epileptiform activity. Biophysical analysis in HEK293T cells revealed that splicing profoundly regulates fast inactivation and channel availability during fast, repetitive stimulation, with neonatal channels showing higher availability compared to adult channels and this difference was conserved among Nav1.1, Nav1.2 and Nav1.7. The change in inactivation imposed by splicing can be modeled as a modification of the stability of the inactivation statein resting channels. This change can be eradicated by administration of the AEDs phenytoin and carbamazepine. Current-clamp recordings in transfected neurons showed that the alternatively spliced variantmodifies the rising phase of action potentials for Nav1.1 and Nav1.2 at high firing frequencies, implying a consistent splice-dependent modulation of channel availability. For Nav1.1 in interneurons, this translated to higher firing frequency for the neonatal isoform, which also conferred a higher maximal firing rate during epileptiform events imposed under dynamic-clamp recordings.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available