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Europace Advance Access originally published online on June 19, 2007
Europace 2007 9(8):571-577; doi:10.1093/europace/eum121
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© The European Society of Cardiology 2007. All rights reserved. For Permissions, please e-mail: journals.permissions@oxfordjournals.org

BASIC SCIENCE

Effects of heart failure on brain-type Na+ channels in rabbit ventricular myocytes

Arie O. Verkerk1, Antoni C.G. van Ginneken1, Toon A.B. van Veen2 and Hanno L. Tan1,3,*

1 Heart Failure Research Center, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands; 2 Department of Medical Physiology, University Medical Center Utrecht, Utrecht, The Netherlands; 3 Department of Cardiology, Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands

Manuscript submitted 11 January 2007. Accepted after revision 16 May 2007.

* Corresponding author. Tel: +31 20 5663265; fax: +31 20 6975458. E-mail address: h.l.tan{at}amc.uva.nl


   Abstract
Top
 Abstract
Introduction
 Methods
 Results
 Discussion
 Implications of our study
 References
 
Aims Brain-type {alpha}-subunit isoforms of the Na+ channel are present in various cardiac tissue types and may control pacemaker activity and excitation–contraction coupling. Heart failure (HF) alters pacemaker activity and excitation–contraction coupling. Here, we studied whether HF alters brain-type Na+ channel properties.

Methods and results HF was induced in rabbits by volume/pressure overload. Na+ currents of ventricular myocytes were recorded in the cell-attached mode of the patch-clamp technique using macropatches. Macropatch recordings were conducted from the middle portions of myocytes or from intercalated disc regions between cell pairs. Both areas exhibited a fast activating and inactivating current, 8.5 times larger in intercalated disc regions. Tetrodotoxin (TTX) (50 nM) did not block currents in the intercalated disc regions, but did block in the middle portions, indicating that the latter currents were TTX-sensitive brain-type Na+ currents. Macropatch recordings from these regions were used to study the effects of HF on brain-type Na+ current. Neither current density nor gating properties (activation, inactivation, recovery from inactivation, slow inactivation) differed between CTR and HF.

Conclusion The density and gating properties of brain-type Na+ current are not altered in our HF model. In the volume/pressure-overload rabbit model of HF, the role of brain-type Na+ current in HF-induced changes in excitation–contraction coupling is limited.

Key Words: Heart failure, Sodium current, Excitation–contraction coupling, Pacemaker activity


   Introduction
Top
 Abstract
 Introduction
Methods
 Results
 Discussion
 Implications of our study
 References
 
Voltage-gated Na+ (NaV) channels play a critical role in the rising phase of the action potential and are, thus, important for impulse generation and conduction in most excitable cells. They are composed of one large pore-forming {alpha}-subunit and one or more smaller ancillary ß-subunits.1Go–3Go Several {alpha}-subunit Na+ channel isoforms have been identified with distinct patterns of development and localization in the nervous system, skeletal muscle, and cardiac muscle, and different physiological and pharmacological properties.4Go Isoforms preferentially expressed in the central nervous system, i.e. NaV1.1, NaV1.2, NaV1.3, NaV1.6 (collectively called brain-type in this study), are inhibited by nanomolar concentrations of tetrodotoxin (TTX), as is the isoform present in adult skeletal muscle (NaV1.4). In contrast, the primary cardiac isoform, NaV1.5, requires micromolar concentrations of TTX for inhibition because of the presence of a cysteine instead of an aromatic residue in the pore region of domain I.5Go

The TTX-resistant NaV1.5 is most prominent in various cardiac preparations, but recent studies show that the TTX-sensitive brain-type isoforms are also present in sinoatrial node (SAN),6Go–8Go Purkinje cells,9Go and ventricular myocytes.9Go–14Go The contribution of TTX-sensitive brain-type isoforms to total Na+ current in myocytes varies between cardiac tissues. In primary (SAN) and secondary (Purkinje) pacemaker cells it is approximately 45 and 22%, respectively,8Go,9Go whereas it varies between 10 and 20% in ventricular myocytes of various species.9Go,11Go,13Go,14Go In ventricular myocytes, NaV1.5 and the brain-type Na+ channels may have distinct subcellular localizations and functions. Immunocytochemistry studies show that NaV1.1, NaV1.3, and NaV1.6 are present uniformly across the sarcolemma in the transverse (t-) tubular system, whereas NaV1.5 channels are clustered primarily at unique sites within the intercalated discs.11Go,12Go,15Go Nav1.5 channels are involved in initiation and propagation of the cardiac action potential from cell to cell. A functional role of brain-type Na+ channels in cardiac tissues is thought to involve excitation–contraction coupling by coordination and synchronization of the action potential from the sarcolemma into the interior via the t-tubules.11Go This is believed to underlie the synchronous Ca2+ release from the sarcoplasmic reticulum, which results in ventricular contractions (for review, see Brette and Orchard14). The role of brain-type Na+ channels in excitation–contraction coupling, however, is debated.16Go Block of TTX-sensitive brain-type Na+ channels by 100 nM TTX increased the cycle length in intact heart, intact SAN preparations and isolated SAN cells of mice by 15–60%,7Go,8Go suggesting that brain-type Na+ channels play a substantial role in SAN pacemaker activity.

Both excitation–contraction coupling17Go and pacemaker activity18Go are affected in failing hearts, although there are marked differences in reported changes between various HF models and/or species (see Discussion). We hypothesized that changes in brain-type Na+ channel properties may partially underlie these changes. Accordingly, we studied the biophysical properties of brain-type Na+ channels in isolated control (CTR) and heart failure (HF) cells using a well-characterized rabbit model of pressure and volume overload-induced HF. In this study, we focused on ventricular myocytes and did not study SAN properties, whereas we recognized that ventricular myocytes and SAN may be subject to different HF-related remodelling responses.


   Methods
Top
 Abstract
 Introduction
 Methods
Results
 Discussion
 Implications of our study
 References
 
Cell preparation
Cardiac failure was induced in New Zealand-White male rabbits (n = 6) by combined volume and pressure overload as described previously.19Go Hearts were excised 3–4 months after creating both aortic regurgitation by catheter-induced damage to the aortic valve, and abdominal aortic stenosis by ligation of the abdominal aorta. At the time of sacrifice, a HF index based on relative heart weight, relative lung weight, left ventricular end-diastolic pressure, third heart sound, and ascites were calculated.19Go We performed experiments if at least three of the five parameters were abnormal,18Go indicating severe HF. A previous study from our laboratory indicated that differences between non-operated and sham-operated rabbits are negligible.19Go Therefore, non-operated age-matched healthy animals (n = 10) served as CTR. All management and experimental procedures conducted during this study were performed in accordance with the institutional guidelines and complied with the Declaration of Helsinki.

Cells were isolated by enzymatic dissociation from the midmyocardium of the left ventricular free wall as described previously.20Go Small aliquots of cell suspension were put in a recording chamber on the stage of an inverted microscope. Cells were allowed to adhere for 5 min after which superfusion with Tyrode's solution was started. Tyrode's solution (36±0.2°C) contained (in mmol/L): NaCl 140, KCl 5.4, CaCl2 1.8, MgCl2 1.0, glucose 5.5, HEPES 5.0, pH 7.4 (NaOH). Single specimens and cell-pairs of rod-shaped myocytes that had smooth surfaces and clear cross-striations were selected for the measurements.

Electrophysiology
Na+ currents were recorded in the macropatch cell-attached mode of the patch-clamp technique,21Go using an Axopatch 200B amplifier (Axon Instruments). Both superfusion and pipette solutions were Tyrode's solution. Pipettes were pulled from borosilicate glass capillaries using a vertical microelectrode puller and were heat-polished. Open tip resistances of pipettes were 1.2–1.6 M{Omega}. Currents were filtered at 5 kHz and digitized at 10 kHz.

The Na+ currents were characterized by determination of the voltage dependence of activation, voltage-dependent inactivation, recovery from inactivation, and slow inactivation using custom voltage-clamp protocols modified from those published previously.22Go,23Go Similar to the study of Murray et al.,21Go the current amplitudes were averaged from multiple pulses; activation and inactivation protocols were repeated two times, recovery and slow inactivation protocols were repeated three times. Each cell was used to measure one parameter only (activation, inactivation, recovery from inactivation, or slow inactivation), within 4 min after the Giga seal was achieved. Details of each pulse protocol are given schematically in the figures and explained in the ‘Results’ section. All pipette command potentials in this study were relative to the resting membrane potential of the myocytes, which we determined previously24Go in a series of whole-cell patch-clamp experiments using Tyrode's solution as indicated above and pipette solution containing (in mmol/L): K-glucose 125, KCl 20, NaCl 5, HEPES 10; pH 7.2 (KOH). The resting membrane potentials of the myocytes averaged – 81.5 ± 0.9 and – 81.7 ± 0.6 mV in CTR (n = 37) and failing (n = 32) myocytes, respectively. To facilitate the protocols used in this study, the pipette command potentials were corrected by – 82 mV. INa was defined as the difference between peak current and the current at the end of the depolarizing voltage step.

Statistics
Data are expressed as mean ± SEM. Group comparisons were made using the unpaired Student's t-test. Statistical significance was defined as P < 0.05.


   Results
Top
 Abstract
 Introduction
 Methods
 Results
Discussion
 Implications of our study
 References
 
Presence and density of CTR and HF brain-type Na+ channels
The localization of brain-type Na+ channels is clearly distinct from the cardiac NaV1.5 channels. NaV1.5 channel are concentrated primarily at intercalated discs, whereas brain-type Na+ channel subunits are localized uniformly across the sarcolemma.11Go–12Go,15Go We conducted cell-attached recordings, using an electrode with a relatively large opening to measure from a macropatch on the surface membrane. We hypothesized that membrane patches from the middle portion of the myocytes contain brain-type Na+ channels, while the intercalated discs at the margins of the myocytes predominantly contain NaV1.5 channels. To test this hypothesis, electrode tips were placed on the middle region of single myocytes (Figure 1A, right panel) or in intact intercalated disc regions between cell pairs (Figure 1A, left panel). These experiments were performed in CTR myocytes. The success rate in finding cell pairs with clear cell boundaries by positioning the electrode tips and obtaining Giga seal recordings from intercalated disc regions was low. Nevertheless, the macropatch recordings of both regions clearly demonstrated the presence of a fast activating and inactivating current (Figure 1B). The ‘quasi-macroscopic’ currents in the intercalated disc region, however, were 8.5 times larger than those obtained from the middle regions of the myocytes (Figure 1B, inset). ‘Quasi-macroscopic’ currents from the middle region of the myocytes were not observed when 50 nM TTX was added to the pipette solution (Figure 1B, inset, and below). In contrast, large ‘quasi-macroscopic’ currents still could be recorded from the intercalated discs in the presence of 50 nM TTX (Figure 1B, inset). When 30 µM TTX was added to the pipette solution, however, ‘quasi-macroscopic’ currents were not observed from the intercalated discs (n = 5; data not shown). These findings confirm that the observed current in membrane areas from the middle region of the myocytes was TTX-sensitive brain-type Na+ current, whereas intercalated disc Na+ current are carried by clusters of TTX-insensitive NaV1.5 channels.


Figure 1
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  		Figure 1 Functional localization of NaV1.5 and brain-type Na+ channels in control rabbit ventricular myocytes. (A) Positioning of the patch-clamp microelectrode in the intercalated disc region of a cell pair (left panel) and middle portion of a single myocyte (right panel). The patch-clamp microelectrode comes from the right. (B) Typical examples of Na+ currents recorded in a macropatch of the intercalated disc region (left panel) and middle region of the myocyte (right panel). Currents were generated from a holding potential of – 140 mV by applying 50 ms voltage steps to – 40 mV. (Inset) Average Na+ current amplitudes in the absence and presence of 50 nM tetrodotoxin (TTX). Numbers indicate myocytes/heart.

 
To study the effects of HF on the biophysical properties of brain-type Na+ channels, we next conducted cell-attached recordings from the middle regions of the myocytes. A clear fast activating and inactivating current was observed in both CTR and HF myocytes (Figure 2A and C). Baba et al.25Go found that cell surface localization of NaV1.5 channels may differ between myocytes of normal hearts and those isolated from infarct regions. In both CTR and HF myocytes, ‘quasi-macroscopic’ currents were absent in the presence of 50 nM TTX (Figure 2B and C), indicating that the observed currents were the TTX-sensitive brain-type Na+ current. The amplitudes of brain-type Na+ currents in CTR and HF were similar. The average current–voltage relationships were not significantly different between CTR and HF brain-type Na+ currents (Figure 2C).


Figure 2
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  		Figure 2 Presence and densities of CTR and HF brain-type Na+ channels. (A, B) Examples of brain-type Na+ current traces recorded in a control (CTR) and heart failure (HF) cell in the absence (A) and presence of 50 nM TTX (B). Currents were generated from a holding potential of – 140 mV by applying 50 ms voltage steps to – 40 mV. (C) Current–voltage relationship for CTR and HF brain-type Na+ channels. Numbers indicate myocytes/heart.

 
(In)activation gating properties of CTR and HF brain-type Na+ channels
To determine the activation characteristics of CTR and HF quasi-macroscopic brain-type Na+ channels, current–voltage curves, as shown in Figure 2C, were corrected for changes in driving force, normalized to maximum peak current, and fitted with a Boltzmann distribution curve to determine V1/2 [membrane potential for half-maximal (in)activation] and the slope factor k. The resulting curves for voltage-dependence of activation of CTR and HF brain-type Na+ channels are shown in Figure 3A. Activation curves of CTR and HF brain-type Na+ channels matched closely. Accordingly, V1/2 (membrane potential for half-maximal activation) and slope factor k values for CTR and HF brain-type Na+ channel activation were not significantly different. Activation V1/2 averaged – 50.6 ± 2.7 and – 48.7 ± 1.9 mV, and slope factors 5.8 ± 0.4 and 5.9 ± 0.5 mV (CTR and HF, respectively).


Figure 3
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  		Figure 3 Activation and inactivation properties of CTR and HF brain-type Na+ channels. (A) Voltage-dependence of activation for CTR and HF brain-type Na+ channels. Currents were generated from a holding potential of – 140 mV by applying 50 ms voltage steps from – 140 to + 50 mV in 5 mV increments and a cycle length of 5 s (see protocol inset). The resulting current amplitude at each voltage was divided by the maximum peak Na+ current and plotted versus the corresponding voltage to obtain the voltage-dependent activation curve. Solid lines represent the Boltzmann fits of the average data; numbers indicate myocytes/heart. (B) Voltage-dependence of inactivation of CTR and HF brain-type Na+ channels. Voltage-dependence of inactivation was obtained by measuring the peak Na+ current during a 50 ms test step to – 40 mV, which followed a 500 ms prepulse to membrane potentials between – 160 and + 40 mV from a holding potential of – 140 mV (see protocol inset). Test pulse current amplitude was divided by its maximum current and plotted versus the voltage of the prepulse to construct the voltage-dependent inactivation curve (cycle lengths: 5 s). Solid lines represent the Boltzmann fits of the average data; numbers indicate myocytes/heart.

 
Voltage-dependent inactivation relationships were obtained by measuring the peak currents during a 50 ms test step to – 40 mV, which followed a 500 ms prepulse to membrane potentials between – 160 and + 40 mV to induce inactivation. Test pulse current amplitude was normalized to its maximum current and plotted versus the prepulse voltage to assess the voltage-dependent inactivation curve. This curve was fitted with the Boltzmann distribution function {y = A/[1 + exp((V – V1/2)/k)]} to determine the V1/2 (membrane potential at which 50% of Na+ channels have entered an inactivated state) and slope factor k of voltage-dependent inactivation. The voltage-dependence of inactivation for CTR and HF brain-type Na+ channels cells are shown in Figure 3B. The inactivation properties of HF Na+ channels were indistinguishable from those of CTR (Figure 3B). V1/2 and k were – 88.1 ± 3.0 and – 7.9 ± 0.9 mV (CTR), and – 88.6 ± 1.4 and – 7.7 ± 0.9 mV (HF), respectively.

Recovery from inactivation properties of CTR and HF brain-type Na+ channels
The recovery time course is shown in Figure 4. Recovery from inactivation was measured using a two pulse protocol, where a conditioning pulse (P1) to – 40 mV inactivated Na+ channels, followed by a test pulse (P2) to – 40 mV after a variable recovery interval ranging between 1 and 1000 ms at a recovery potential of – 140 mV. The peak current in response to P2 was normalized to the peak current at P1, and plotted versus the recovery time interval. The resulting curve was fitted with a double-exponential function to obtain the time constants of the fast and the slow components of recovery from inactivation: y = [Af x exp(–t/{tau}f)] + [As x exp(– t/{tau}s)], where t is the recovery time interval, {tau}f and {tau}s the time constants of fast and slow components, and Af and As the fractions of the fast and slow component. Time constants of recovery from inactivation were not significantly different between CTR and HF brain-type Na+ channels (Figure 4). Time constants of fast and slow components of recovery ({tau}f and {tau}s, respectively) were 10.0 ± 2.5 and 1090 ± 194 ms in CTR and 10.5 ± 2.5 and 978 ± 297 ms in HF.


Figure 4
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  		Figure 4 Recovery from inactivation of CTR and HF brain-type Na+ channels. Recovery from inactivation was measured using a two-pulse protocol with a conditioning pulse (P1) to – 40 mV, followed by a 20 ms long test pulse (P2) after a variable recovery interval at a hyperpolarizing voltage of – 140 mV ranging between 1 and 1000 ms (see protocol insets). The fraction of channels that had recovered following the intervals, was calculated by dividing the peak Na+ currents evoked by P2 by the peak current measured during P1, and plotted versus the recovery interval (cycle lengths: 5 s). Solid lines are fits using double-exponential functions (for details, see Results section) of the average data; numbers indicate myocytes/heart.

 
Slow inactivation properties of CTR and HF brain-type Na+ channels
The slow inactivation course is shown in Fig. 5. To assess slow inactivation, we used a two-pulse protocol with an initial conditioning prepulse to – 40 mV (P1) of variable duration ranging between 1 and 1000 ms, followed by a step to – 140 mV of 30 ms to allow the channels to recover from fast inactivation, and a final test pulse to – 40 mV (P2) to assess the fraction of channels available for activation. This fraction was obtained by dividing the current amplitude during P2 by the current amplitude during P1, plotted versus the duration of P1, and designated Peak INa (P2/P1). Consequently, the fraction that entered slow inactivation equals 1 – Peak INa (P2/P1). HF brain-type Na+ channels did not exhibit significantly more slow inactivation when compared with CTR brain-type Na+ channels [Figure 5; 20.6 ± 3.6% (CTR) vs. 22.6 ± 3.1% (HF) at P2 of 1000 ms].


Figure 5
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  		Figure 5 Slow inactivation of CTR and HF brain-type Na+ channels. A two-pulse protocol was used with an initial conditioning prepulse to – 40 mV (P1) of variable duration ranging between 1 and 1000 ms, followed by a 30 ms step to – 140 mV to allow the channels to recover from fast inactivation but not slow inactivation, and a final test pulse to – 40 mV (P2) to assess the fraction of channels that had escaped slow inactivation and were available for activation (see protocol inset). This fraction was obtained by dividing the current during P2 by the current during P1, and plotted versus the duration of P1 (cycle lengths: 5 s). Solid lines represent fits using double-exponential functions, y = [Af x exp(–t/{tau}f)]+[As x exp(–t/{tau}s)], of the average data; numbers indicate myocytes/heart.

 

   Discussion
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
Implications of our study
 References
 
The Na+ channel has various isoforms. Although the cardiac isoform NaV1.5 predominates in the heart, other isoforms found predominantly in the brain have been reported to be present in the heart. We employed macropatch cell-attached patch clamp methodology to: (i) demonstrate for the first time with a functional assay that various Na channel isoforms have different locations in the sarcolemmal membrane; (ii) characterize the electrophysiological effects of HF on brain-type Na+ current in a well-established rabbit model of pressure and volume overload-induced HF. This is the first report, which uses macropatches to characterize the biophysiological properties of brain-type Na+ current in cardiac myocytes. Moreover, it is also the first report on possible functional consequences, if any, of HF on brain-type Na+ channels.

Macropatch measurements of brain-type Na+ current in cardiac myocytes
So far, electrophysiological studies into the properties of brain-type Na+ channels have been frequently conducted by the use of the whole-cell patch-clamp technique and 100–200 µM TTX with the aim of distinguishing the resulting Na+ currents from those of Nav1.5.8Go,9Go,11Go,13Go Whether such TTX concentrations only block the brain-type Na+ channels, however, is questionable, given that the EC50 of NaV1.5 channels is 2–6 µM.26Go To circumvent potential problems regarding TTX-sensitivity, we recorded quasi-macroscopic Na+ currents in the cell-attached mode using an electrode with a relatively large opening to measure from a macropatch on the surface membrane. We reasoned that, due to the differences in localization of the various Na+ channel isoforms, an isolated membrane patch from the middle portion of the cell would only contain brain-type Na+ channels. These quasi-macroscopic currents were absent in the presence of nanomolar concentrations (50 nM) of TTX (Figures 1 and 2), confirming that the observed currents were indeed brain-type Na+ currents. This finding agrees with immunolocalization findings of Maier,11Go but contrasts with findings of Cohen,15Go who reported, using immunolocalization labelling, that NaV1.5 is also present along the cell surface and at the z-lines in adult rat ventricular myocytes. According to Maier,11Go this finding by Cohen may be due to the fact that antibodies had also recognized NaV1.3.

We found that quasi-macroscopic currents from the intercalated disc region were insensitive to 50 nM of TTX, while they were abolished by 30 µM TTX. Moreover, the Na+ current amplitudes obtained in the intercalated disc region were 8.5 times larger than the middle portion of the myocytes. These findings suggest that the channels in the intercalated disc region constitute the NaV1.5 isoform. Our electrophysiological study agrees with previous studies, using immunolocalization methods, which reported that NaV1.5 is predominantly present in the intercalated disc region.11Go–12Go,15Go

The use of the macropatch technique has the advantages that the normal cytoplasmic environment is maintained, and that experiments could be performed at close-to-physiological temperatures and ion concentrations.21Go,27Go Nevertheless, while the intracellular milieu is left intact, cell-attached recording methods retain the potential problem that time-dependent changes in Na+ current (in)activation properties may occur.28Go–30Go In addition, due to the close-to-physiological temperature and ion concentrations, and the absence of specific ion channel blockers, the presence of other types of voltage-dependent ion channels in the patch recordings cannot be excluded. Moreover, Na+ channels are attached to the cytoskeletal elements under normal conditions.31Go In the cell-attached mode of the patch-clamp technique, the membrane is pulled into the opening of the pipette tip, forming a ohm-shaped deformation.32Go This may detach the channels in the membrane from these cytoskeletal elements, and culminate in altered properties. Finally, in the present study, we used an extracellular solution with 5.4 mM K+. Thus, the myocytes had a resting membrane potential close to the Nernst equation for K+ ions, i.e. – 82 mV.24Go The inward currents flowing through the macropatch may depolarize the cell, thereby confounding the findings of the voltage dependence of (in)activation.

We cannot completely exclude the possibility that these disadvantages of the cell-attached macropatch methodology have influenced our absolute values of V1/2 of (in)activation. Yet, we reasoned that these possible confounders were only of limited significance when brain-type Na+ current properties in CTR and HF myocytes were compared. First, each cell was used to measure one biophysiological parameter only within 4 min after Giga seal formation. This procedure minimized possible time-dependent changes in Na+ current (in)activation properties in both CTR and HF. Secondly, Na+ channels in myocytes have a relatively high density when compared with other channels, which suggests that the contribution of other types of voltage-dependent ion channels in our measurements was limited. This suggestion was supported by our observation that, after TTX addition, remaining voltage-dependent ion channels were virtually absent. Thirdly, brief treatments (< 10–15 min) of cell-attached patches with cytochalasin D (Cyto-D), an agent that interferes with actin polymerization, induced short bursts of Na+ channel openings and prolonged decays of ensemble-averaged currents. Such Na+ current behaviour was absent without Cyto-D, indicating that, under normal conditions, cell-attached patches did not modify the interaction between the Na+ channel and the cytoskeletal elements much.33Go Finally, in our rabbit model of HF, background K+ conductance at resting membrane potential is not diminished.34Go Also, the current density of brain-type Na+ current did not differ between CTR and HF myocytes (Figure 2C). In both CTR and HF myocytes, we calculated that the depolarizations due to the macropatch currents did not exceed 2 mV.

Biophysical properties of brain-type Na+ channels in CTR and HF
Ionic remodelling during HF of various outward K+ currents in both animal models and in humans is well known.35Go,36Go The effects of HF on inward Na+ currents are less well-established. Peak cardiac Na+ current density was reported to be unaltered in models of hypertrophy in rats37Go and human atrium,38Go in pressure and volume overload-induced HF in rabbit,24Go and in pacing-induced HF in dogs.39Go However, another study of peak cardiac Na+ current density in a pacing-induced HF model in dogs showed a 39% decrease.40Go Peak cardiac Na+ current density was also significantly decreased by 39% in an ischaemia-induced HF model in dog,41Go by 30% in a guinea pig hypertrophy model,42Go and by 57% in human heart failure,40Go respectively. In the present study, we found no effects of HF on the density of brain-type Na+ current. This finding agrees with quantitative measures of mRNA, including RNAse protection assays and real-time quantitative PCR, in a pacing-induced HF model in dog, which showed no changes in mRNA levels of three {alpha}-subunit isoforms (NaV1.1, 1.3, 1.5).40Go

Activation properties and inactivation properties of cardiac Na+ current are not altered by HF in dog and human ventricular myocytes.40Go In the present study, we found no effects of HF on (in)activation properties of brain-type Na+ current. Also, recovery from inactivation and slow inactivation were unchanged.

Altogether, the reported effects of HF on cardiac Na+ current density are not consistent. The discrepancies between the various studies may be due to differences in the methods to induce HF, species, or tissues. It is unknown whether the absence of effects on electrophysiological properties of HF on brain-type Na+ current, as found here, is specific to our HF model, species used, or tissue studied.


   Implications of our study
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 Implications of our study
References
 
The present work demonstrates that the density and gating properties of TTX-sensitive brain-type Na+ currents are not changed during HF in ventricular myocytes. This observation is important because it excludes a role of brain-type Na+ current in HF-induced changes in excitation–contraction coupling. It must be noted, however, that this conclusion is based on studies in our HF model and that, given the reported variability in observed changes between various HF models and/or species, it is not immediately clear whether this conclusion also applies to other cell types, HF models, or humans.


   Acknowledgments
 
We thank Charly N.W. Belterman, Antonius Baartscheer, and Cees A. Schumacher for their excellent technical assistance. This study was supported by the Netherlands Organization for Scientific Research Grant 916.36.012 (T.A.B.V.), the Netherlands Heart Foundation Grant 2002B191 (H.L.T.), the Royal Netherlands Academy of Arts and Sciences KNAW (H.L.T.), and the Bekales Foundation (H.L.T.).

Conflict of interest. none declared.


   References
Top
 Abstract
 Introduction
 Methods
 Results
 Discussion
 Implications of our study
 References
 
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[7] Maier SK, Westenbroek RE, Yamanushi TT, Dobrzynski H, Boyett MR, Catterall WA, et al. An unexpected requirement for brain-type sodium channels for control of heart rate in the mouse sinoatrial node. Proc Natl Acad Sci USA (2003) 100:3507–12.[Abstract/Free Full Text]

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Effects of heart failure on brain-type Na+ channels in rabbit ventricular myocytes
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