Effects of the Transient Outward Potassium Current on Action Potential Upstroke Velocities Tested Using the Dynamic Clamp Technique

Verkerk, Arie O.; Veerman, Christiaan C.; Zegers, Jan G.; Wilders, Ronald

doi:10.22489/cinc.2016.076-143

Cited by 2 publications

(4 citation statements)

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“…Thirdly, we assessed upstroke velocities in the HEK293-Na V 1.5 cells, transfected with either IRES-GFP or KCND3 -IRES-GFP plasmids. Noteworthy, HEK293 cells expressing Na V 1.5 display fast depolarizations upon switching from voltage clamp (VC) to current clamp (CC) due to Na + channel activation as previously shown (Berecki et al, 2010 ; Verkerk et al, 2016 ; Lieve et al, 2017 ). Thus, the alternating VC/CC technique allows for a more dynamic assessment of Na V 1.5 current as compared to measurements in VC configuration, in the setting of more physiological temperature and Na + gradients across the membrane.…”

Section: Resultsmentioning

confidence: 67%

“…Using dynamic clamp, we have recently assessed the effect of an I to -like current on the upstroke velocity in HEK293 cells transiently overexpressing Na V 1.5 channels. That study revealed only a minor influence of I to , if any, on upstroke velocity (Verkerk et al, 2016 ). Here, we confirmed these results using computer simulations based on the current densities recorded (Figure 5 ).…”

Section: Discussionmentioning

confidence: 91%

“…The characterization of various knock-out mouse models of α-subunits generating the fast component of I to ( Kcnd3, Kcnd2 ) confirmed the involvement of I to in phase-1 repolarization, but its impact on AP upstroke velocity or I Na density were not investigated (Niwa et al, 2008 ; Liu et al, 2015 ). We recently evaluated the effects of an in silico I to injection on AP upstroke and repolarization velocity using the dynamic clamp technique (Verkerk et al, 2016 ). In that study, we observed a minimal effect (≈2%) of in-silico I to injection on upstroke velocity but only when the injected current was large and rapidly activated at very negative potentials (around −50 mV).…”

Section: Introductionmentioning

confidence: 99%

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KV4.3 Expression Modulates NaV1.5 Sodium Current

et al. 2018

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In cardiomyocytes, the voltage-gated transient outward potassium current (Ito) is responsible for the phase-1 repolarization of the action potential (AP). Gain-of-function mutations in KCND3, the gene encoding the Ito carrying KV4.3 channel, have been associated with Brugada syndrome (BrS). While the role of Ito in the pro-arrhythmic mechanism of BrS has been debated, recent studies have suggested that an increased Ito may directly affect cardiac conduction. However, the effects of an increased Ito on AP upstroke velocity or sodium current at the cellular level remain unknown. We here investigated the consequences of KV4.3 overexpression on NaV1.5 current and consequent sodium channel availability. We found that overexpression of KV4.3 protein in HEK293 cells stably expressing NaV1.5 (HEK293-NaV1.5 cells) significantly reduced NaV1.5 current density without affecting its kinetic properties. In addition, KV4.3 overexpression decreased AP upstroke velocity in HEK293-NaV1.5 cells, as measured with the alternating voltage/current clamp technique. These effects of KV4.3 could not be explained by alterations in total NaV1.5 protein expression. Using computer simulations employing a multicellular in silico model, we furthermore demonstrate that the experimentally observed increase in KV4.3 current and concurrent decrease in NaV1.5 current may result in a loss of conduction, underlining the potential functional relevance of our findings. This study gives the first proof of concept that KV4.3 directly impacts on NaV1.5 current. Future studies employing appropriate disease models should explore the potential electrophysiological implications in (patho)physiological conditions, including BrS associated with KCND3 gain-of-function mutations.

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Section: Resultsmentioning

confidence: 67%

Section: Discussionmentioning

confidence: 91%

Section: Introductionmentioning

confidence: 99%

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KV4.3 Expression Modulates NaV1.5 Sodium Current

et al. 2018

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“…However, a recent study in HEK293 cells, combining in silico injection of I to using dynamic clamp with measurement of I Na -driven upstrokes using alternating voltage clamp/current clamp demonstrated that I to hardly affected upstroke velocity. 28 Furthermore V max can be influenced by sodium channel availability, which is set by both the resting ) cycling, which may modulate I Na . 29 It can be speculated that a decrease in Ca 2+ i may result in a faster V max .…”

Section: Hey2mentioning

confidence: 99%

The Brugada Syndrome Susceptibility Gene HEY2 Modulates Cardiac Transmural Ion Channel Patterning and Electrical Heterogeneity

Veerman

Podliesna

Tadros

et al. 2017

Circulation Research

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Rationale: Genome-wide association studies previously identified an association of rs9388451 at chromosome 6q22.3 (near HEY2 ) with Brugada syndrome. The causal gene and underlying mechanism remain unresolved. Objective: We used an integrative approach entailing transcriptomic studies in human hearts and electrophysiological studies in Hey2 +/− ( Hey2 heterozygous knockout) mice to dissect the underpinnings of the 6q22.31 association with Brugada syndrome. Methods and Results: We queried expression quantitative trait locus data acquired in 190 human left ventricular samples from the genotype-tissue expression consortium for cis -expression quantitative trait locus effects of rs9388451, which revealed an association between Brugada syndrome risk allele dosage and HEY2 expression (β=+0.159; P =0.0036). In the same transcriptomic data, we conducted genome-wide coexpression analysis for HEY2 , which uncovered KCNIP2 , encoding the β-subunit of the channel underlying the transient outward current ( I to ), as the transcript most robustly correlating with HEY2 expression (β=+1.47; P =2×10 −34 ). Transcript abundance of Hey2 and the I to subunits Kcnip2 and Kcnd2 , assessed by quantitative reverse transcription–polymerase chain reaction, was higher in subepicardium versus subendocardium in both left and right ventricles, with lower levels in Hey2 +/− mice compared with wild type. Surface ECG measurements showed less prominent J waves in Hey2 +/− mice compared with wild-type. In wild-type mice, patch-clamp electrophysiological studies on cardiomyocytes from right ventricle demonstrated a shorter action potential duration and a lower V max in subepicardium compared with subendocardium cardiomyocytes, which was paralleled by a higher I to and a lower sodium current ( I Na ) density in subepicardium versus subendocardium. These transmural differences were diminished in Hey2 +/− mice because of changes in subepicardial cardiomyocytes. Conclusions: This study uncovers a role of HEY2 in the normal transmural electrophysiological gradient in the ventricle and provides compelling evidence that genetic variation at 6q22.31 (rs9388451) is associated with Brugada syndrome through a HEY2 -dependent alteration of ion channel expression across the cardiac ventricular wall.

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Effects of the Transient Outward Potassium Current on Action Potential Upstroke Velocities Tested Using the Dynamic Clamp Technique

Abstract: The

Cited by 2 publications

References 6 publications

KV4.3 Expression Modulates NaV1.5 Sodium Current

KV4.3 Expression Modulates NaV1.5 Sodium Current

The Brugada Syndrome Susceptibility Gene HEY2 Modulates Cardiac Transmural Ion Channel Patterning and Electrical Heterogeneity

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