Abstract
ABSTRACTSCN5A is expressed in cardiomyocytes and gastrointestinal (GI) smooth muscle cells (SMCs) as the voltage-gated mechanosensitive sodium channel NaV1.5. The influx of Na+ through NaV1.5 produces a fast depolarization in membrane potential, indispensable for electrical excitability in cardiomyocytes and important for electrical slow waves in GI smooth muscle. As such, abnormal NaV1.5 voltage gating or mechanosensitivity may result in channelopathies. SCN5A mutation G615E – found separately in cases of acquired long-QT syndrome, sudden cardiac death, and irritable bowel syndrome – has a relatively minor effect on NaV1.5 voltage gating. The aim of this study was to test whether G615E impacts mechanosensitivity. Mechanosensitivity of wild-type (WT) or G615E-NaV1.5 in HEK-293 cells was examined by shear stress on voltage- or current-clamped whole cells or pressure on macroscopic patches. Unlike WT, voltage-clamped G615E-NaV1.5 showed a loss in shear- and pressure-sensitivity of peak current yet a normal leftward shift in the voltage-dependence of activation. In current-clamp, shear stress led to a significant increase in firing spike frequency with a decrease in firing threshold for WT but not G615E-NaV1.5. Our results show that the G615E mutation leads to functionally abnormal NaV1.5 channels, which cause disruptions in mechanosensitivity and mechano-electrical feedback and suggest a potential contribution to smooth muscle pathophysiology.
Highlights
The voltage-gated mechanosensitive Na+ channel NaV 1.5 is expressed by SCN5A in human cardiac myocytes and gastrointestinal (GI) smooth muscle cells (SMCs) [1,2]
In the absence of shear stress, we found that voltagedependent peak conductance of G615E NaV1.5 was not different than WT NaV1.5 (GMAX 2.41 ± 0.41 nS, WT vs. 2.24 ± 0.46 nS, G615E; n = 12, P > 0.05) (Figure 1(a,c,g))
As previously described [6,16], whole-cell NaV1.5 conductance increased in response to shear stress (GMAX of WT NaV1.5: 2.41 ± 0.41 nS control to 2.91 ± 0.49 nS shear, 20.7 ± 2.0% increase, n = 12, *P < 0.001 to control) (Figure 1(a-b, g-h))
Summary
The voltage-gated mechanosensitive Na+ channel NaV 1.5 is expressed by SCN5A in human cardiac myocytes and gastrointestinal (GI) smooth muscle cells (SMCs) [1,2]. SCN5A mutations are well established to cause cardiac conduction disorders, called channelopathies. NaV1.5 channels are gated by voltage, but they are mechanosensitive [11,12], and mechanosensitivity of these channels is relevant because they are expressed in heart and gut, which are mechanically active organs. In cardiomyocytes (7), GI SMCs [14,15], and heterologous expression systems [11,12,16], mechanical stimuli alter NaV1.5 function by increasing peak Na+ current (IPEAK), hyperpolarizing the voltage dependence of activation (V1/2A) and availability (inactivation, V1/2I), and accelerating channel kinetics. The mechanism of NaV1.5 mechanosensitivity remains unclear, which limits our ability to understand the contributions of NaV1.5 mutations to pathophysiology
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