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Biophysical and Pharmacological Characterization of Nav1.9 Voltage Dependent Sodium Channels Stably Expressed in HEK-293 Cells.
- Source :
-
PloS one [PLoS One] 2016 Aug 24; Vol. 11 (8), pp. e0161450. Date of Electronic Publication: 2016 Aug 24 (Print Publication: 2016). - Publication Year :
- 2016
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Abstract
- The voltage dependent sodium channel Nav1.9, is expressed preferentially in peripheral sensory neurons and has been linked to human genetic pain disorders, which makes it target of interest for the development of new pain therapeutics. However, characterization of Nav1.9 pharmacology has been limited due in part to the historical difficulty of functionally expressing recombinant channels. Here we report the successful generation and characterization of human, mouse and rat Nav1.9 stably expressed in human HEK-293 cells. These cells exhibit slowly activating and inactivating inward sodium channel currents that have characteristics of native Nav1.9. Optimal functional expression was achieved by coexpression of Nav1.9 with β1/β2 subunits. While recombinantly expressed Nav1.9 was found to be sensitive to sodium channel inhibitors TC-N 1752 and tetracaine, potency was up to 100-fold less than reported for other Nav channel subtypes despite evidence to support an interaction with the canonical local anesthetic (LA) binding region on Domain 4 S6. Nav1.9 Domain 2 S6 pore domain contains a unique lysine residue (K799) which is predicted to be spatially near the local anesthetic interaction site. Mutation of this residue to the consensus asparagine (K799N) resulted in an increase in potency for tetracaine, but a decrease for TC-N 1752, suggesting that this residue can influence interaction of inhibitors with the Nav1.9 pore. In summary, we have shown that stable functional expression of Nav1.9 in the widely used HEK-293 cells is possible, which opens up opportunities to better understand channel properties and may potentially aid identification of novel Nav1.9 based pharmacotherapies.<br />Competing Interests: The authors have declared that no competing interests exist.
- Subjects :
- Amino Acid Sequence
Anesthetics, Local chemistry
Anesthetics, Local pharmacology
Animals
Binding Sites
HEK293 Cells
Humans
Inhibitory Concentration 50
Ion Channel Gating drug effects
Lysine chemistry
Lysine metabolism
Membrane Potentials drug effects
Mice
Models, Molecular
Molecular Conformation
NAV1.9 Voltage-Gated Sodium Channel chemistry
Protein Binding
Rats
Sodium Channel Agonists chemistry
Sodium Channel Agonists pharmacology
Sodium Channel Blockers chemistry
Sodium Channel Blockers pharmacology
Gene Expression
NAV1.9 Voltage-Gated Sodium Channel genetics
NAV1.9 Voltage-Gated Sodium Channel metabolism
Subjects
Details
- Language :
- English
- ISSN :
- 1932-6203
- Volume :
- 11
- Issue :
- 8
- Database :
- MEDLINE
- Journal :
- PloS one
- Publication Type :
- Academic Journal
- Accession number :
- 27556810
- Full Text :
- https://doi.org/10.1371/journal.pone.0161450