Abstract
Supplemental Digital Content is available in the text. Background - Dysregulation of voltage-gated cardiac Na + channels (Na V 1.5) by inherited mutations, disease-linked remodeling, and drugs causes arrhythmias. The molecular mechanisms whereby the Na V 1.5 voltage-sensing domains (VSDs) are perturbed to pathologically or therapeutically modulate Na + current (I Na) have not been specified. Our aim was to correlate I Na kinetics with conformational changes within the 4 (DI-DIV) VSDs to define molecular mechanisms of Na V 1.5 modulation. Method and Results - Four Na V 1.5 constructs were created to track the voltage-dependent kinetics of conformational changes within each VSD, using voltage-clamp fluorometry. Each VSD displayed unique kinetics, consistent with distinct roles in determining I Na. In particular, DIII-VSD deactivation kinetics were modulated by depolarizing pulses with durations in the intermediate time domain that modulates late I Na. We then used the DII-VSD construct to probe the molecular pathology of 2 Brugada syndrome mutations (A735V and G752R). A735V shifted DII-VSD voltage dependence to depolarized potentials, whereas G752R significantly slowed DII-VSD kinetics. Both mutations slowed I Na activation, although DII-VSD activation occurred at higher potentials (A735V) or at later times (G752R) than ionic current activation, indicating that the DII-VSD allosterically regulates the rate of I Na activation and myocyte excitability. Conclusions - Our results reveal novel mechanisms whereby the Na V 1.5 VSDs regulate channel activation and inactivation. The ability to distinguish distinct molecular mechanisms of proximal Brugada syndrome mutations demonstrates the potential of these methods to reveal how inherited mutations, post-translational modifications, and antiarrhythmic drugs alter Na V 1.5 at the molecular level.
Original language | English |
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Pages (from-to) | 1228-1239 |
Number of pages | 12 |
Journal | Circulation: Arrhythmia and Electrophysiology |
Volume | 8 |
Issue number | 5 |
DOIs | |
State | Published - Oct 1 2015 |
Keywords
- Brugada syndrome
- fluorometry
- ion channels
- mutation
- sodium channels