Biophysical properties of Naᵥ 1.8/Naᵥ 1.2 chimeras and inhibition by µO-conotoxin MrVIB

Knapp, O., Nevin, S. T., Yasuda, T., Lawrence, N., Lewis, R. J. and Adams, D. J. (2012) Biophysical properties of Naᵥ 1.8/Naᵥ 1.2 chimeras and inhibition by µO-conotoxin MrVIB. British Journal of Pharmacology, 166 7: 2148-2160.


Author Knapp, O.
Nevin, S. T.
Yasuda, T.
Lawrence, N.
Lewis, R. J.
Adams, D. J.
Title Biophysical properties of Naᵥ 1.8/Naᵥ 1.2 chimeras and inhibition by µO-conotoxin MrVIB
Journal name British Journal of Pharmacology   Check publisher's open access policy
ISSN 1476-5381
0007-1188
Publication date 2012-07-09
Sub-type Article (original research)
DOI 10.1111/j.1476-5381.2012.01955.x
Volume 166
Issue 7
Start page 2148
End page 2160
Total pages 13
Place of publication Chichester, West Sussex, United Kingdom
Publisher John Wiley & Sons
Collection year 2013
Language eng
Formatted abstract Background and purpose: Voltage-gated sodium channels are expressed primarily in excitable cells and play a pivotal role in the initiation and propagation of action potentials. Nine subtypes of the pore-forming α-subunit have been identified, each with a distinct tissue distribution, biophysical properties and sensitivity to tetrodotoxin (TTX). Nav1.8, a TTX-resistant (TTX-R) subtype, is selectively expressed in sensory neurons and plays a pathophysiological role in neuropathic pain. In comparison with TTX-sensitive (TTX-S) Navα-subtypes in neurons, Nav1.8 is most strongly inhibited by the µO-conotoxin MrVIB from Conus marmoreus. To determine which domain confers Nav1.8 α-subunit its biophysical properties and MrVIB binding, we constructed various chimeric channels incorporating sequence from Nav1.8 and the TTX-S Nav1.2 using a domain exchange strategy.

Experimental approach: Wild-type and chimeric Nav channels were expressed in Xenopus oocytes, and depolarization-activated Na+ currents were recorded using the two-electrode voltage clamp technique.

Key results: MrVIB (1 µM) reduced Nav1.2 current amplitude to 69 ± 12%, whereas Nav1.8 current was reduced to 31 ± 3%, confirming that MrVIB has a binding preference for Nav1.8. A similar reduction in Na+ current amplitude was observed when MrVIB was applied to chimeras containing the region extending from S6 segment of domain I through the S5-S6 linker of domain II of Nav1.8. In contrast, MrVIB had only a small effect on Na+ current for chimeras containing the corresponding region of Nav1.2.

Conclusions and implications: Taken together, these results suggest that domain II of Nav1.8 is an important determinant of MrVIB affinity, highlighting a region of the α-subunit that may allow further nociceptor-specific ligand targeting.
Keyword Electrophysiology
Heterologous expression
Xenopus oocytes
Chimera
Tetrodotoxin
Voltage-gated sodium channels
Q-Index Code C1
Q-Index Status Confirmed Code
Institutional Status UQ
Additional Notes Article first published online: 9 July 2012.

 
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Created: Tue, 10 Jul 2012, 13:55:17 EST by Debra McMurtrie on behalf of Queensland Brain Institute