Search tips
Search criteria 


Logo of jcinvestThe Journal of Clinical InvestigationCurrent IssueArchiveSubscriptionAbout the Journal
J Clin Invest. 2008 June 2; 118(6): 2260–2268.
Published online 2008 May 8. doi:  10.1172/JCI33891
PMCID: PMC2373423

Sodium channel β1 subunit mutations associated with Brugada syndrome and cardiac conduction disease in humans


Brugada syndrome is a genetic disease associated with sudden cardiac death that is characterized by ventricular fibrillation and right precordial ST segment elevation on ECG. Loss-of-function mutations in SCN5A, which encodes the predominant cardiac sodium channel α subunit NaV1.5, can cause Brugada syndrome and cardiac conduction disease. However, SCN5A mutations are not detected in the majority of patients with these syndromes, suggesting that other genes can cause or modify presentation of these disorders. Here, we investigated SCN1B, which encodes the function-modifying sodium channel β1 subunit, in 282 probands with Brugada syndrome and in 44 patients with conduction disease, none of whom had SCN5A mutations. We identified 3 mutations segregating with arrhythmia in 3 kindreds. Two of these mutations were located in a newly described alternately processed transcript, β1B. Both the canonical and alternately processed transcripts were expressed in the human heart and were expressed to a greater degree in Purkinje fibers than in heart muscle, consistent with the clinical presentation of conduction disease. Sodium current was lower when NaV1.5 was coexpressed with mutant β1 or β1B subunits than when it was coexpressed with WT subunits. These findings implicate SCN1B as a disease gene for human arrhythmia susceptibility.


Voltage-gated sodium channels are critical for the generation and propagation of the cardiac action potential, and mutations in SCN5A, the gene encoding the major pore-forming sodium channel α subunit in the heart (NaV1.5), cause multiple cardiac arrhythmia syndromes (14). Mutations producing enhanced inward current during the course of the action potential plateau, often as a consequence of destabilized fast inactivation of the channel, cause long QT syndrome type 3 (LQT3; OMIM 603830) (1). On the other hand, a reduction in sodium current leads to cardiac conduction disease, which may be progressive (OMIM 113900) (2, 3), and Brugada syndrome (OMIM 601144), characterized by ST segment elevation in the right precordial leads (V1 to V3) of the 12-lead ECG and episodes of ventricular fibrillation (4). Multiple mechanisms have been described that reduce sodium current in these syndromes, including altered gating of the channel or reduced cell-surface expression (5). In addition, mutations in SCN5A may manifest with an overlap of these different phenotypes (610). However, mutations in SCN5A are found in fewer than 30% of patients with Brugada syndrome, indicating involvement of other genes (11). A mutation in the glycerol-3-phosphate dehydrogenase 1–like gene (GPD1L) has recently been reported in a large kindred with Brugada syndrome (12); however, GPD1L mutations are rare in Brugada syndrome (13). Antzelevitch et al. have recently reported mutations in the gene encoding the L-type calcium channel (CACNA1C) or its β2b subunit (CACNB2b) in Brugada syndrome patients with unusually short QT intervals (14), but the frequency of these defects as a cause for more-typical Brugada syndrome is unknown. SCN5A mutations are also not identified in the majority of patients with cardiac conduction disease (15).

Sodium channels are multisubunit protein complexes composed not only of pore-forming α subunits but also of multiple other protein partners including auxiliary function-modifying β subunits (16, 17). In humans, 4 sodium channel β subunits (β1 to β4, encoded by SCN1B to SCN4B) have been identified, and they share a common predicted protein topology: a large extracellular N-terminal domain (including an immunoglobulin-like domain), a single transmembrane segment, and an intracellular C-terminal domain (16). Functions attributed to β subunits include an increase in sodium channel expression at the cell surface, modulation of channel gating and voltage dependence, and a role in cell adhesion and recruitment of cytosolic proteins such as ankyrin-G (16).

The β1 transcript arises from splicing of exons 1–5 of the SCN1B gene (Figure (Figure1,1, A and B). More recently, a second transcript has been described, arising from splicing of exons 1–3 with retention of a segment of intron 3 (termed exon 3A), leading to an alternate 3ι sequence (Figure (Figure1,1, A and B) (18, 19). This latter transcript encodes the β1B subunit, which, in spite of the different 3ι sequence, has a predicted protein topology similar to that of β1 (Figure (Figure1C)1C) (19). The β1B subunit has been shown to increase a neuronal sodium current (NaV1.2) (19), but its effects on NaV1.5 current have not yet been investigated, although β1 and β1B are both expressed in heart (19, 20).

Figure 1
Structure of β1 and β1B subunits.

Since loss-of-function NaV1.5 mutations cause conduction disease and Brugada syndrome, one could envision that mutations in sodium channel β subunits could also underlie these disorders by decreasing sodium current. Therefore, we tested the hypothesis that mutations in SCN1B coding sequences, for either β1 or β1B, underlie cases of conduction disease and Brugada syndrome. We identified 3 mutations segregating with arrhythmia in 3 kindreds, and 2 of the mutations were located in the newly described β1B transcript. Both β1 and β1B transcripts were expressed in the human heart and were abundant in Purkinje fibers that play a critical role in electric pulse conduction in heart. Electrophysiologic study of heterologously expressed sodium channels revealed loss of sodium current with mutant subunits.


Mutation analysis and clinical data.

We screened 282 probands with Brugada syndrome and 44 with conduction disease for mutations in exons 1–5 of SCN1B encoding the β1 subunit and in exon 3A retained in the β1B transcript (Figure (Figure1,1, A and B). SCN5A coding region mutations had been previously excluded in all 326 subjects. Three variants were identified in probands and family members (Figure (Figure2A).2A). These variants were absent in 1,404 population controls (see Methods).

Figure 2
SCN1B mutations found in patients with Brugada syndrome and conduction disease.

A missense mutation, c.259G→C (Figure (Figure2B)2B) in exon 3, resulting in p.Glu87Gln within the extracellular immunoglobulin loop of the protein (Figure (Figure1C)1C) was identified in a Turkish kindred affected by conduction disease (family 1; Figure Figure2A).2A). Alignment of the β1 subunit amino acid sequence from multiple species demonstrated that Glu87 is highly conserved, supporting the importance of glutamate at this position (Figure (Figure2C).2C). The proband was a 50-year-old white Turkish female (II-1) who presented with palpitations and dizziness. Physical examination and echocardiography were normal, and her ECG showed complete left bundle branch block. A clinical electrophysiological study revealed a prolonged His-ventricle interval of 80 ms and inducible atrioventricular nodal reentrant tachycardia; complete atrioventricular block occurred following atrial programmed stimulation and during induced tachycardia. A dual-chamber pacemaker was implanted with resolution of symptoms. The same mutation was found in her brother (II-3), who had bifascicular block (right bundle branch block and left anterior hemiblock), and her mother (I-2), who had a normal ECG. There was no family history of syncope, sudden cardiac death, or epilepsy.

A nonsense mutation, c.536G→A in exon 3A (Figure (Figure1B1B and Figure Figure2D),2D), was identified in a French kindred affected with Brugada syndrome and conduction disease (family 2; Figure Figure2A).2A). This mutation results in p.Trp179X and is predicted to generate a prematurely truncated protein lacking the membrane-spanning segment and intracellular portion of the protein (Figure (Figure1C).1C). The proband was a 53-year-old white male (II-4) who presented with chest pain. Physical examination, echocardiography, and coronary angiography were normal. His ECG showed ST segment elevation typical of Brugada syndrome and conduction abnormalities (prolonged PR interval of 220 ms and left anterior hemiblock; Figure Figure2E)2E) (21). Ventricular fibrillation was induced by programmed electrical stimulation in basal state (in the absence of drugs). The same mutation was detected in his brother (II-1), nephew (III-1), and sister (II-2). The brother had no palpitations or history of syncope. His baseline ECG showed left anterior hemiblock and minor ST segment elevation suggestive of Brugada syndrome at baseline (type II saddleback abnormality; ref. 21); with flecainide challenge, the ST segment elevation was further exaggerated but did not meet criteria for a diagnostic (type I) pattern. The nephew had right bundle branch block and type II Brugada syndrome ECG after flecainide challenge, and the sister had a normal ECG and a negative flecainide test. There was no family history of tachyarrhythmias, syncope, sudden cardiac death, or epilepsy.

A different nonsense mutation, c.537G→A in exon 3A (Figure (Figure2D),2D), resulting in p.Trp179X, affecting the same codon as in family 2, was identified in a Dutch kindred (family 3; Figure Figure2A).2A). The proband was a 17-year-old white female (II-1). Physical examination and echocardiography were normal, and a flecainide test for Brugada syndrome was negative. Her ECG showed right bundle branch block and prolonged PR interval of 196 ms (normal upper limit in teenagers, 180 ms) (22). The same mutation was found in her father (I-1), with normal ECG and negative flecainide test. The family history was negative for syncope, sudden cardiac death, or epilepsy.

β1 and β1B transcript expression.

To confirm and extend previous reports that β1B is expressed in brain, heart, skeletal muscle, and other organs (19), we used quantitative real-time PCR in nondiseased human heart. Both β1 and β1B transcripts were detected in right and left ventricles and in Purkinje fibers (Figure (Figure3).3). The β1 transcript level was higher in Purkinje fibers (which make up the conduction system in the ventricle) than left- (2.4-fold; P < 0.05) and right-ventricular (1.6-fold; P = NS) free walls. β1B transcript levels showed an even greater difference: Purkinje fibers versus left- (4.8-fold; P < 0.001) and right-ventricular (3.7-fold; P < 0.001) free walls. Levels of both transcripts were also slightly (but not statistically significantly) higher in right- versus left-ventricular free wall (1.5-fold and 1.3-fold for β1 and β1B transcripts, respectively).

Figure 3
Expression profile of β1 and β1B transcripts in nondiseased human ventricular tissue as determined by quantitative real-time PCR.

Cellular electrophysiology.

The effects of mutant and WT β1 and β1B variants on NaV1.5 sodium current were assessed using the whole-cell patch-clamp technique in transfected CHO cells. As described in Methods, bicistronic expression vectors encoding a reporter (GFP or DsRed) with or without β subunits were cotransfected with expression vector encoding NaV1.5. Currents were compared in cells transfected with SCN5A alone or SCN5A plus WT, mutant, or both β subunits.

Figure Figure4A4A shows representative current traces in cells expressing NaV1.5 alone and NaV1.5 plus WT or mutant β1B (p.Trp179X β1B) or their combination; current densities at –30 mV are summarized in Figure Figure4B.4B. Coexpression of NaV1.5 with WT β1B significantly increased sodium current density over NaV1.5 alone, by 69%, while currents recorded with p.Trp179X β1B coexpression were no different from NaV1.5 alone. Similarly, while coexpression of WT subunit with NaV1.5 shifted the voltage dependence of both activation and inactivation to more negative potentials compared with those with NaV1.5 alone, no such shift was observed with the mutant (Figure (Figure4C4C and Table Table1).1). This result indicates that while WT β1B modulates NaV1.5 gating (in a fashion similar to WT β1; see below), the mutant exerts no such effect. Coexpression of WT or mutant β1B with NaV1.5 did not alter recovery from inactivation (Figure (Figure4D4D and Table Table1). 1).

Figure 4
Electrophysiological characteristics of the p. Trp179X β1B mutant.
Table 1
Biophysical parameters of WT and mutant β1B

To examine whether expression of the mutant influences the effect of WT β1B on NaV1.5 current (e.g., to produce a dominant negative action), cells were transfected with NaV1.5 and varying amounts of WT and p.Trp179X β1B. Figure Figure4B4B shows that the sodium current increase over NaV1.5 alone recorded with transfection of 1 μg of both β1B subunit constructs was identical to the increase with that of 1 μg of WT β1B. In addition, the increase in sodium current recorded with transfection of 0.5 μg of both β1B subunit constructs was 51% of that with 1 μg of β1B alone. These data indicate that p.Trp179X β1B does not exert a dominant negative effect on WT β1B function and further support the finding that the mutant, unlike WT, does not affect sodium channel function.

Figure Figure5A5A shows representative current traces of NaV1.5 and NaV1.5 coexpressed with WT and/or mutant β1 (p.Glu87Gln β1); current densities are summarized in Figure Figure5B.5B. Coexpression of NaV1.5 with WT β1 significantly increased sodium current density at –30 mV, by 76%, while coexpression with mutant β1 (p.Glu87Gln β1) did not increase the sodium current. The increase in sodium current recorded with coexpression of NaV1.5 and 1 μg of both WT and p.Glu87Gln β1 (+20%) was markedly smaller than the increase with coexpression of NaV1.5 with 1 μg WT β1 alone (+76%), indicating that this mutant exerts a dominant negative effect on WT β1 function. Figure Figure5C5C shows that WT β1 produced negative shifts in the voltage dependence of NaV1.5 activation and inactivation similar to those observed with WT β1B. p.Glu87Gln β1 shifted the voltage dependence of inactivation to negative potentials (similar to WT β1) but did not alter the voltage dependence of activation (Table (Table2).2). Coexpression of WT or mutant β1 with NaV1.5 did not alter recovery from inactivation (Figure (Figure5D5D and Table Table2). 2).

Figure 5
Electrophysiological characteristics of the p.Glu87Gln mutant.
Table 2
Biophysical parameters of WT and mutant β1

Since Glu87 is located in a region of the protein common to both β1 and β1B, we also studied the effects of p.Glu87Gln β1B on NaV1.5 current properties (Supplemental Figure 1 and Supplemental Table 1; supplemental material available online with this article; doi: 10.1172/JCI33891DS1). While WT β1B increased NaV1.5 current by 69% (Figure (Figure4),4), p.Glu87Gln β1B did not increase the sodium current compared with NaV1.5 alone. Similarly, WT β1B produced a negative shift in voltage dependence of both activation and inactivation (Table (Table1),1), while p.Glu87Gln β1B shifted only the voltage dependence of inactivation compared with NaV1.5 alone. As with the other β subunit constructs studied, there was no change in recovery from inactivation. Thus, the effects of p.Glu87Gln were comparable in the β1 and the β1B backbones.

Since Glu87 is located in a region of the protein common to both β1 and β1B, we also studied the effects of p.Glu87Gln β1B on NaV1.5 current properties (Supplemental Figure 1 and Supplemental Table 1). While WT β1B increased NaV1.5 current by 69% (Figure (Figure4),4), p.Glu87Gln β1B did not increase the sodium current compared with NaV1.5 alone. Similarly, WT β1B produced a negative shift in voltage dependence of both activation and inactivation (Table (Table1),1), while p.Glu87Gln β1B shifted only the voltage dependence of inactivation compared with NaV1.5 alone. As with the other β subunit constructs studied, there was no change in recovery from inactivation. Thus, the effects of p.Glu87Gln were comparable in the β1 and the β1B backbones.


In this study, we provide what we believe to be the first report of mutations in SCN1B sequences encoding the β1 and β1B transcript variants in patients with conduction disease and/or Brugada syndrome. Further, we provide new data indicating that β1 and β1B transcripts in the heart vary by region; greater expression in Purkinje fibers is consistent with the conduction system phenotype we describe in mutation carrier patients. Finally, we demonstrate that the β1 and β1B variants modulate function of the major cardiac sodium channel α subunit NaV1.5 and that the identified SCN1B mutations blunt or inhibit this effect.

The 3 mutations were identified in 3 probands with conduction disease and/or Brugada syndrome as well as in other family members with or without these arrhythmia phenotypes. Formal linkage analysis was not possible because the families are too small and penetrance is incomplete. Thus, evidence in support of disease causality of these mutations (beyond their identification in subjects with clinical phenotypes) includes the findings that both β1 and β1B transcripts are expressed in heart and that the mutant subunits (p.Glu87Gln β1, p.Glu87Gln β1B, and p.Trp179X β1B) did not increase NaV1.5 currents in heterologous expression experiments, while WT β1 and β1B did. Incomplete penetrance, a well-recognized feature of the monogenic arrhythmia syndromes (12, 23), was observed. For SCN5A mutations linked to Brugada syndrome, penetrance as low as 12.5% has been described (24). A role for sex, age, and genetic modifiers (e.g., common polymorphisms) is suspected (5, 25, 26), but the mechanisms for this common clinical finding remain poorly understood.

Two types of mutations were identified. The c.536G→A and c.537G→A mutations in exon 3A both result in a stop codon at position 179, predicted to generate a β1B protein lacking the transmembrane and cytoplasmic domains and thus unable to integrate into the sarcolemma and to associate with NaV1.5. Thus, the a priori assumption is that a mutation such as this will cause disease by simple haploinsufficiency. The electrophysiologic data support this idea, since coexpression of p.Trp179X β1B failed to increase NaV1.5 current and did not modulate the effect of the WT β1B protein. Furthermore, the voltage dependencies of activation and inactivation of NaV1.5 coexpressed with p.Trp179X β1B were the same as those for NaV1.5 alone, in contrast to the shifts observed with WT β1B. While Scn1b-knockout mice display clear ECG changes (27), studies with young (17- to 18-day-old) heterozygotes identified no difference from WT. Since age-related changes in conduction are a recognized feature of cardiac conduction disease and conduction delay is one of the proposed mechanisms of Brugada syndrome (2, 28), aging may be important for the β subunit–mediated phenotype.

On the other hand, the c.259G→C mutation leads to an amino acid substitution (p.Glu87Gln) within the extracellular domain of the protein. The electrophysiological data demonstrate that the mutant subunit did modulate NaV1.5 gating (shift in the voltage dependence of inactivation, in either the β1 or β1B background), supporting the idea that it associates with NaV1.5 at the cell surface. In addition, in contrast to the p.Trp179X β1B, p.Glu87Gln did exert a dominant negative effect on the WT subunit. Thus, the 3 mutations lead to a decrease in NaV1.5 current through somewhat different mechanisms. This reduction of current is consistent with the conduction disease and Brugada syndrome phenotypes of the patients.

Normal impulse propagation in the atria, ventricles, and Purkinje network is critically dependent on normal sodium channel function. Dysfunction of the sodium channel leads to conduction delay, and loss-of-function mutations in SCN5A have been described in isolated conduction disease unassociated with structural heart disease (2, 3). Thus, our finding of SCN1B mutations associated with reduced sodium current in patients with conduction disease is consistent with previous studies of the mechanism of this disorder. The preferential expression of the β1 and β1B transcripts in human Purkinje fibers further supports the prominent conduction delay seen as part of the clinical phenotypes.

Loss-of-function mutations in SCN5A were the first reported cause of the Brugada syndrome (4). These mutations reduce sodium current by reducing NaV1.5 cell surface expression and/or altering gating (4, 5, 29). A common view is that in epicardial cells, this reduction in sodium current produces marked action potential shortening, attributed to an “unopposed” early transient outward potassium current. By contrast, reduction of sodium current in endocardial cells is thought to produce only modest action potential shortening. The resultant increased heterogeneity of repolarization predisposes to rapid reentry, resulting in ventricular fibrillation (4, 30). A common feature in Brugada syndrome — consistent with reduced sodium current — is slowed conduction (28, 31). Indeed, an alternate proposed mechanism suggests that the characteristic right-precordial ST-segment elevation on the ECG and initiation of arrhythmias is attributable primarily to right-ventricular outflow tract conduction delay (28). The trend to higher expression levels of β1B in right ventricle may thus contribute to the Brugada syndrome phenotype.

This idea is further supported by functional studies in a single large kindred in which a GPD1L mutation was linked to Brugada syndrome: coexpression of mutant GPD1L with NaV1.5 was reported to decrease sodium current, consistent with the observation that loss-of-function mutations in SCN5A cause Brugada syndrome (12). In principle, reduction in L-type calcium current might also produce differential effects in epicardial and endocardial sites and thus cause Brugada syndrome; rare kindreds with this mechanism have now been described (14).

Conduction disease was observed in families 1 and 3, while in family 2, mutation carriers presented either solely with conduction disease or conduction disease in combination with ECGs typical of Brugada syndrome. This phenomenon of overlapping clinical phenotypes is common in individuals with SCN5A mutations leading to loss of sodium channel function (6, 7), and conversely, in vitro electrophysiologic analysis of SCN5A mutations linked to Brugada syndrome or isolated conduction disease consistently reveals loss of NaV1.5 channel function (2, 4). Indeed, a single mutation segregating in a given family can lead to conduction disease in some family members and Brugada syndrome in others (6, 7). What determines the ultimate phenotype — Brugada syndrome versus isolated conduction disease — is unknown. Sex, age, and genetic modifiers (e.g., common polymorphisms) have been proposed as modulators of the clinical phenotypes (5, 25, 26).

The reported effects of β1 on NaV1.5 channels are controversial (32). Some groups have reported that β1 increases NaV1.5 currents with or without affecting voltage dependence or channel kinetics, while others have reported no effect of β1 on NaV1.5 current (20, 3337). The β1B variant has to date only been studied in coexpression studies with the neuronal sodium channel NaV1.2 (encoded by SCN2A), where it was shown to increase sodium current and cause a small negative shift in voltage dependence of activation (19). In our experiments, WT β1 and β1B had similar effects on NaV1.5 current: both increased sodium currents and led to hyperpolarizing (negative) shifts in voltage dependence of activation and inactivation.

Not only were the effects of the WT β subunits on NaV1.5 current similar, but the effects of the p.Glu87Gln mutation in the β1 background (p.Glu87Gln β1) were also similar to those in the β1B background (p.Glu87Gln β1B). Although the β1 and β1B variants share the same topology (an N-terminal extracellular immunoglobulin domain, a transmembrane domain, and a C-terminal cytoplasmic domain), their sequence identity is limited to the extracellular immunoglobulin domain; the C-terminal half of β1B, residues 150–268, has only approximately 17% amino acid sequence identity with β1 (19). Taken together, the data suggest that the molecular determinants of β1 and β1B modulation of NaV1.5 cell-surface expression and gating likely reside in the extracellular immunoglobulin domain. This is in line with previous studies of skeletal muscle (NaV1.4 encoded by SCN4A) and neuronal (NaV1.2) sodium channel α subunits that have shown that deletion of the intracellular domain of the β1 subunit has no effect on its modulation of α subunit function, whereas deletions within the extracellular domain block modulation (3840). Alternatively, specific residues may not be as important as preservation of overall structural motifs, as suggested by the data of Zimmer and Benndorf, who reported that the β1 subunit modulates NaV1.5 via the membrane anchor plus additional intracellular or extracellular regions (41).

In addition to modulation of sodium channel α subunit expression and function, other roles have been suggested for β subunits: these include acting as adhesion molecules or as participants in signal transduction (16, 32). The different transmembrane and C-terminal domains of β1 and β1B might therefore lead to participation in different signaling pathways. For instance, phosphorylation of the tyrosine at position 181 of the β1 C terminus regulates its interaction with ankyrin-G (42), which is thought to be critical for ankyrin-G localization within cardiomyocytes (intercalated discs versus T tubules). β1B lacks this tyrosine in its C-terminal domain, so a role for β1B as a modulator of this function seems less likely.

Another mechanism regulating function of β subunits is the potential for cleavage by β-site amyloid precursor protein–cleaving enzyme (BACE1) and γ-secretase, resulting in the release of the N- and C-terminal fragments (43). The processed C-terminal fragment of β2 and β4 has been reported to be associated with cell adhesion, migration, and morphogenesis in neuronal cells as well as regulation of the expression level of the neuronal sodium channel NaV1.1 (4446). Thus, p.Trp179X β1B may result in absence of functions depending on the generation of a β subunit C-terminal fragment by BACE1. However, a role for BACE1 cleavage has not been studied in either human β1 subunit or cardiomyocytes, and the cleavage site located at the common juxtamembrane domain in β1 and β1B is not conserved between human and mice (19, 43).

Mutations in SCN1B have been previously reported in generalized epilepsy with febrile seizures plus (47), and β1-null mice exhibit a severe seizure disorder and die at approximately 3 weeks of age (48). In addition, these mice exhibit bradycardia and prolonged rate-corrected QT intervals (27). These changes suggest that β1 plays an important role in the murine heart, although it is possible that the changes are a consequence of the severe overall developmental phenotype in this model (48). To our knowledge, defects in cardiac function have not been investigated in SCN1B mutation carriers presenting with epilepsy (32, 49). Conversely, we have observed no neurological phenotype in our patients. Four SCN1B mutations have been linked to epilepsy to date (47, 49, 50), all of which localize to the extracellular immunoglobulin-like fold of the protein, as does the p.Glu87Gln mutation reported here. One additional possible link between the cardiac and neurological phenotypes associated with β1 mutations is the syndrome of sudden unexpected death in epilepsy (SUDEP) (51), where a role for cardiac bradyarrhythmias has been proposed (52).

To date, SCN5A mutations are the most common cause identified in cases of Brugada syndrome, and SCN5A is the only identified causative gene in conduction disease (2, 11). However, SCN5A mutations are not identified in the majority of patients, and it has been reported that the frequency of mutations in other implicated genes (CACNA1C, CACNB2b, GPD1L) is also low in Brugada syndrome (1214). In this study, SCN1B mutations were identified in less than 1% of probands with Brugada syndrome and less than 5% of probands with conduction disease and thus account for a small subset of these inherited arrhythmic syndromes.

A conventional heterologous mammalian expression system was used for functional assessment of the mutations. The environment in this approach is different from that in native cardiomyocytes, and other proteins known to associate with the sodium channel complex, such as other β subunits, are generally absent. Despite these limitations, the in vitro characteristics of the mutations were concordant with the phenotype observed in the patients, which, in combination with the genetic data presented, supports the hypothesis of a causal relationship between the mutations and disease.

In summary, we have for the first time to our knowledge identified SCN1B mutations in families with conduction disease and Brugada syndrome. We have shown expression of the β1 subunit transcript and the alternate β1B subunit transcript variant in human heart and demonstrated reduced NaV1.5 sodium current as a result of loss or altered β subunit modulation of Nav1.5 current. These findings implicate SCN1B as a disease gene for human arrhythmia susceptibility.


Study populations.

The study populations consisted of: (a) unrelated Brugada syndrome probands identified and characterized at the Academic Medical Center, Amsterdam (n = 38), l’institut du thorax, Nantes, (n = 216), and the Niigata University Graduate School of Medical and Dental Sciences (n = 28); and (b) patients with cardiac conduction disease were identified and characterized at the Academic Medical Center (n = 2), l’institut du thorax (n = 39), and Ege University School of Medicine (n = 3). The study was performed according to a protocol approved by the Medical Ethical Committee, Academic Medical Center, Amsterdam; Comité de Protection des Personnes, Nantes; Ege University Research Ethics Committee; and Medical Research Ethics Committee, Niigata University Graduate School of Medical and Dental Sciences. Informed consent was obtained from all patients. Coding region and splice site mutations in SCN5A had been previously excluded in all probands by single-strand confirmation polymorphism analysis, denaturing HPLC sequencing, or direct sequencing using primers in flanking intronic sequences.

Mutation analysis.

Probands with Brugada syndrome and cardiac conduction disease were screened for mutations in regions of the SCN1B gene encoding β1 and β1B, except for Japanese probands, who were screened only in the regions of SCN1B gene encoding β1B. Screening for mutations was performed by PCR amplification of coding regions and flanking intronic sequences, followed by direct sequencing of amplicons on an ABI PRISM 3730 DNA Sequence Detection System (Applied Biosystems). Primer sequences are listed in Supplemental Table 2.

Control populations.

We screened randomly selected and unrelated white Dutch individuals (n = 176); white individuals (n = 702) selected from the KORA S4 survey, which included population-based southern German individuals (n = 4,261) surveyed between 1999 and 2001 (53); unrelated white Turkish individuals (n = 150); and 4 different ethnic groups (white, African American, Hispanic, Asian; n = 94 for each group) from the Coriell Cell Repositories. The Coriell samples were resequenced as described above by the J. Craig Venter Institute through the NHLBI DNA Resequencing and Genotyping Program. The other control samples were genotyped at the identified mutation sites.

Subunit mRNA abundances in human cardiac tissue.

Real-time RT-PCR was used to quantify subunit abundances. Assays were conducted in nondiseased human hearts obtained from the University of Szeged, Szeged, Hungary, that were technically unusable for transplantation based on logistic considerations (54). Before cardiac explantation, organ donor patients did not receive medication except dobutamine, furosemide, and plasma expanders. The investigations conformed to the principles outlined in the Helsinki Declaration of the World Medical Association. All experimental protocols were approved by the Ethical Review Board of the Medical Center of the University of Szeged (no. 51-57/1997 OEJ). The left ventricles from 6 donors and the right ventricles from 6 donors were dissected and stored in cardioplegic solution at 4°C for approximately 4–8 hours before being frozen in liquid nitrogen. Purkinje fiber mRNA was extracted from false tendons dissected from the ventricles of 6 donors. Further information on the donors is presented in Supplemental Table 3.

Total RNA from each cardiac sample was isolated and DNase treated with the RNeasy Fibrous Tissue Mini Kit (QIAGEN) following the manufacturer’s instructions. The quality of total RNA was assessed with PAGE (2100 Bioanalyzer; Agilent Technologies). Absence of genomic DNA contamination was verified by PCR. First-strand cDNA was synthesized from 2 μg total RNA with High-Capacity cDNA Archive Kit (Applied Biosystems). Real-time PCR was performed on a TaqMan system with predesigned 6-carboxyfluorescein–labeled (FAM-labeled) fluorogenic TaqMan probe and primers for β1, custom-designed TaqMan probe and primers for β1B (located in the retained segment of intron 3), and 1× TaqMan Universal PCR Master Mix (Applied Biosystems). PCR efficiency of the β1 and β1B fluorescent probes was estimated at approximately 98%. After 2 minutes at 50°C and 10 minutes at 95°C, 40 cycles of amplification were performed with the ABI PRISM 7900HT Sequence Detection System (Applied Biosystems). Data were collected with instrument spectral compensation by Applied Biosystems SDS 2.1 software and analyzed with the Ct relative quantification method (55). Fluorescence signals were normalized to the housekeeping gene hypoxanthine phosphoribosyl transferase 1 (HPRT1). For each sample, β1 and β1B transcripts were quantified in duplicate. The values were averaged and then used for the 2–ΔCt × 100 calculation, where 2–ΔCt corresponds to expression relative to HPRT. Primer and probe sequences are listed in Supplemental Table 4.

Generation of expression vectors.

Full-length human β1 cDNA (GenBank accession number NM_001037) subcloned into a bicistronic vector also carrying the cDNA for enhanced eGFP (pEGFP-IRES; BD Biosciences — Clontech) was supplied by Alfred George Jr. (Vanderbilt University, Nashville, Tennessee, USA). Full-length human β1B cDNA (GenBank accession number NM_199037) was cloned from human ventricular mRNA, supplied by Katherine Murray (Vanderbilt University). The β1B cDNA was subcloned into a pEGFP-IRES vector (BD Biosciences — Clontech). Mutant constructs were prepared using the QuikChange II XL Site-Directed Mutagenesis Kit (Stratagene) according to the manufacturer’s instructions. The inserts were subsequently sequenced to ensure that there was no other mutation besides the intended one.

Transient transfection in CHO cells.

For functional analysis, cultured CHO cells were transiently transfected with the constructs described above using FuGENE 6 (Roche Applied Science). Constructs encoding β1 or β1B subunits (1 μg, unless otherwise specified) were cotransfected with the pBK-CMV vector (1 μg; Stratagene) encoding SCN5A (GenBank accession number NM_000335), supplied by Alfred George Jr. To study dominant negative effects, mutant β1 or β1B construct (0.5 μg or 1 μg) was cotransfected with the same amount of construct for WT β1 or β1B subunit that had been subcloned into a bicistronic vector also carrying cDNA for red fluorescent protein from Discosoma version T3 (pDsRed-IRES; supplied by Alfred George Jr.) along with SCN5A (1 μg). When SCN5A was transfected without β subunits, the plasmid encoding the eGFP (pEGFP-IRES; BD Biosciences — Clontech) with no β subunit insert was cotransfected. Cells were grown for 48 hours after transfection before study.


Cells displaying green fluorescence were chosen for study; in experiments with transfection of both WT and mutant β subunits, cells displaying both green and red fluorescence were chosen. Sodium currents were measured at room temperature using the whole-cell configuration of the patch-clamp technique with an Axopatch 200B amplifier (Molecular Devices). The extracellular bath solution contained (in mmol/l): 145 NaCl, 4.0 KCl, 1.0 MgCl2, 1.8 CaCl2, 10 glucose, 10 HEPES, pH 7.4 (NaOH). Patch pipettes (~1.5 MΩ) contained (in mmol/l): 10 NaF, 110 CsF, 20 CsCl, 10 EGTA, and 10 HEPES, pH 7.4 (CsOH). Currents were filtered at 5 kHz and digitized at 50 kHz. Cell capacitance and series resistance were compensated for by at least 80%. Voltage control, data acquisition, and analysis were accomplished using pCLAMP 9.2 and Clampfit 9.2 software (Molecular Devices).

Sodium current properties were determined by voltage clamp protocols as shown in the relevant figures. Cells were held at –120 mV, and currents were elicited with 50-ms depolarizing pulses from –80 to 60 mV in 10-mV increments. Voltage dependence of inactivation was studied using 500-ms prepulses from –120 to –20 mV in 10-mV increments, followed by a test pulse to –20 mV. The rate of recovery from inactivation was examined by 50-ms conditioning pulse to –20 mV from a holding potential of –120 mV, followed by a varying recovery duration and a 10-ms test pulse to –20 mV. All currents were normalized to cell capacitance. The voltage dependence of sodium current was determined by fitting a Boltzmann function (y = [1 + exp{(VV1/2) / k}]–1), yielding the voltage required to achieve half-maximal conductance or channel availability (V1/2) and slope factor (k). The time constants of recovery from inactivation were determined using a double-exponential function (y = Af{1 – exp(–t / τf)} + As{1 – exp(–t / τs)}), where τf and τs are the time constants of fast and slow components, and Af and As are the fractions of the fast and slow components.


Electrophysiological data are expressed as mean ± SEM. Gene expression data are expressed as median ± median absolute deviation. All statistical analyses were conducted with SPSS version 12.0. To test for significant differences among groups, an unpaired 2-tailed t test or ANOVA was used. The level of statistical significance was P < 0.05.

Supplementary Material

Supplemental data:


We thank Leander Beekman, Peter van Tintelen, Arie O. Verkerk, Carol Ann Remme, Alfred George Jr., Katherine Murray, Sabina Kupershmidt, Kai Liu, Sameer Chopra, Nathalie Gaborit, Satoru Komura, Mahmut Akyol, and Moritz Sinner for their contributions to performing and/or analyzing this work and for helpful discussions. This work was supported by grants from the NIH (HL46681 and HL65962 to D.M. Roden), a Fondation Leducq Trans-Atlantic Network of Excellence grant (05 CVD 01, Preventing Sudden Death, to D. Escande, J.-J. Schott, A.M. Wilde, S. Kääb, and D.M. Roden), Netherlands Heart Foundation grant 2003T302 (to A.A.M. Wilde), the Interuniversity Cardiology Institute of The Netherlands (project 27, to A.A.M. Wilde), Agence Nationale de la Recherche grant 05-MRAR-028 (to J.-J. Schott), Agence Nationale de la Recherche grants 05-MRAR-028 and 06-MRAR-022 (to J.-J. Schott), German Federal Ministry of Education and Research (BMBF) grants 01GI0204, 01GS0499, 01GI0204, and 01GR0103 (to S. Kääb, A. Pfeufer, and H.-E. Wichmann), and a Sumitomo Life Social Foundation grant (to H. Watanabe). The KORA platform is funded by the BMBF and by the State of Bavaria. Resequencing of Coriell samples was performed by the J. Craig Venter Institute through the NHLBI Resequencing and Genotyping Program. We also thank Andras Varro (University of Szeged) for providing the human tissues. C.R. Bezzina is an Established Investigator of The Netherlands Heart Foundation (grant 2005/T024).


Conflict of interest: The authors have declared that no conflict of interest exists.

Citation for this article: J. Clin. Invest. 118:2260–2268 (2008). doi:10.1172/JCI33891

Hiroshi Watanabe, Tamara T. Koopmann, and Solena Le Scouarnec contributed equally to this work.


1. Wang Q., et al. SCN5A mutations associated with an inherited cardiac arrhythmia, long QT syndrome. Cell. 1995;80:805–811. doi: 10.1016/0092-8674(95)90359-3. [PubMed] [Cross Ref]
2. Schott J.J., et al. Cardiac conduction defects associate with mutations in SCN5A. Nat. Genet. 1999;23:20–21. [PubMed]
3. Tan H.L., et al. A sodium-channel mutation causes isolated cardiac conduction disease. Nature. 2001;409:1043–1047. doi: 10.1038/35059090. [PubMed] [Cross Ref]
4. Chen Q., et al. Genetic basis and molecular mechanism for idiopathic ventricular fibrillation. Nature. 1998;392:293–296. doi: 10.1038/32675. [PubMed] [Cross Ref]
5. Tan H.L., Bezzina C.R., Smits J.P., Verkerk A.O., Wilde A.A. Genetic control of sodium channel function. Cardiovasc. Res. 2003;57:961–973. doi: 10.1016/S0008-6363(02)00714-9. [PubMed] [Cross Ref]
6. Bezzina C., et al. A single Na(+) channel mutation causing both long-QT and Brugada syndromes. Circ. Res. 1999;85:1206–1213. [PubMed]
7. Kyndt F., et al. Novel SCN5A mutation leading either to isolated cardiac conduction defect or Brugada syndrome in a large French family. Circulation. 2001;104:3081–3086. doi: 10.1161/hc5001.100834. [PubMed] [Cross Ref]
8. Grant A.O., et al. Long QT syndrome, Brugada syndrome, and conduction system disease are linked to a single sodium channel mutation. J. Clin. Invest. 2002;110:1201–1209. [PMC free article] [PubMed]
9. Valdivia C.R., et al. A novel SCN5A arrhythmia mutation, M1766L, with expression defect rescued by mexiletine. Cardiovasc. Res. 2002;55:279–289. doi: 10.1016/S0008-6363(02)00445-5. [PubMed] [Cross Ref]
10. Remme C.A., et al. Overlap syndrome of cardiac sodium channel disease in mice carrying the equivalent mutation of human SCN5A-1795insD. Circulation. 2006;114:2584–2594. doi: 10.1161/CIRCULATIONAHA.106.653949. [PubMed] [Cross Ref]
11. Shimizu W., Aiba T., Kamakura S. Mechanisms of disease: current understanding and future challenges in Brugada syndrome. Nat. Clin. Pract. Cardiovasc. Med. 2005;2:408–414. doi: 10.1038/ncpcardio0268. [PubMed] [Cross Ref]
12. London B., et al. Mutation in glycerol-3-phosphate dehydrogenase 1 like gene (GPD1-L) decreases cardiac Na+ current and causes inherited arrhythmias. Circulation. 2007;116:2260–2268. doi: 10.1161/CIRCULATIONAHA.107.703330. [PMC free article] [PubMed] [Cross Ref]
13. Koopmann T.T., et al. Exclusion of multiple candidate genes and large genomic rearrangements in SCN5A in a Dutch Brugada syndrome cohort. Heart Rhythm. 2007;4:752–755. doi: 10.1016/j.hrthm.2007.02.021. [PubMed] [Cross Ref]
14. Antzelevitch C., et al. Loss-of-function mutations in the cardiac calcium channel underlie a new clinical entity characterized by ST-Segment elevation, short QT intervals, and sudden cardiac death. Circulation. 2007;15:442–449. [PMC free article] [PubMed]
15. Moric E., et al. The implications of genetic mutations in the sodium channel gene (SCN5A). Europace. 2003;5:325–334. doi: 10.1016/S1099-5129(03)00085-0. [PubMed] [Cross Ref]
16. Isom L.L. Sodium channel beta subunits: anything but auxiliary. Neuroscientist. 2001;7:42–54. [PubMed]
17. Abriel H., Kass R.S. Regulation of the voltage-gated cardiac sodium channel Nav1.5 by interacting proteins. Trends Cardiovasc. Med. 2005;15:35–40. doi: 10.1016/j.tcm.2005.01.001. [PubMed] [Cross Ref]
18. Kazen-Gillespie K.A., et al. Cloning, localization, and functional expression of sodium channel beta1A subunits. J. Biol. Chem. 2000;275:1079–1088. doi: 10.1074/jbc.275.2.1079. [PubMed] [Cross Ref]
19. Qin N., et al. Molecular cloning and functional expression of the human sodium channel beta1B subunit, a novel splicing variant of the beta1 subunit. Eur. J. Biochem. 2003;270:4762–4770. doi: 10.1046/j.1432-1033.2003.03878.x. [PubMed] [Cross Ref]
20. Nuss H.B., et al. Functional association of the beta 1 subunit with human cardiac (hH1) and rat skeletal muscle (mu 1) sodium channel alpha subunits expressed in Xenopus oocytes. J. Gen. Physiol. 1995;106:1171–1191. doi: 10.1085/jgp.106.6.1171. [PMC free article] [PubMed] [Cross Ref]
21. Wilde A.A., et al. Proposed diagnostic criteria for the Brugada syndrome: consensus report. Circulation. 2002;106:2514–2519. doi: 10.1161/01.CIR.0000034169.45752.4A. [PubMed] [Cross Ref]
22. Dickinson D.F. The normal ECG in childhood and adolescence. Heart. 2005;91:1626–1630. doi: 10.1136/hrt.2004.057307. [PMC free article] [PubMed] [Cross Ref]
23. Priori S.G., Napolitano C., Schwartz P.J. Low penetrance in the long-QT syndrome: clinical impact. Circulation. 1999;99:529–533. [PubMed]
24. Priori S.G., et al. Clinical and genetic heterogeneity of right bundle branch block and ST-segment elevation syndrome: a prospective evaluation of 52 families. Circulation. 2000;102:2509–2515. [PubMed]
25. Viswanathan P.C., Benson D.W., Balser J.R. A common SCN5A polymorphism modulates the biophysical effects of an SCN5A mutation. J. Clin. Invest. 2003;111:341–346. [PMC free article] [PubMed]
26. Bezzina C.R., et al. Common sodium channel promoter haplotype in Asian subjects underlies variability in cardiac conduction. Circulation. 2006;113:338–344. doi: 10.1161/CIRCULATIONAHA.105.580811. [PubMed] [Cross Ref]
27. Lopez-Santiago L.F., et al. Sodium channel Scn1b null mice exhibit prolonged QT and RR intervals. J. Mol. Cell. Cardiol. 2007;43:636–647. doi: 10.1016/j.yjmcc.2007.07.062. [PMC free article] [PubMed] [Cross Ref]
28. Meregalli P.G., Wilde A.A., Tan H.L. Pathophysiological mechanisms of Brugada syndrome: depolarization disorder, repolarization disorder, or more? Cardiovasc. Res. 2005;67:367–378. doi: 10.1016/j.cardiores.2005.03.005. [PubMed] [Cross Ref]
29. Baroudi G., et al. Novel mechanism for Brugada syndrome: defective surface localization of an SCN5A mutant (R1432G). Circ. Res. 2001;88:e78–e83. doi: 10.1161/hh1201.093270. [PubMed] [Cross Ref]
30. Yan G.X., Antzelevitch C. Cellular basis for the Brugada syndrome and other mechanisms of arrhythmogenesis associated with ST-segment elevation. Circulation. 1999;100:1660–1666. [PubMed]
31. Antzelevitch C., et al. Brugada syndrome: report of the second consensus conference: endorsed by the Heart Rhythm Society and the European Heart Rhythm Association. Circulation. 2005;111:659–670. doi: 10.1161/01.CIR.0000152479.54298.51. [PubMed] [Cross Ref]
32. Meadows L.S., Isom L.L. Sodium channels as macromolecular complexes: implications for inherited arrhythmia syndromes. Cardiovasc. Res. 2005;67:448–458. doi: 10.1016/j.cardiores.2005.04.003. [PubMed] [Cross Ref]
33. Qu Y., et al. Modulation of cardiac Na+ channel expression in Xenopus oocytes by beta 1 subunits. J. Biol. Chem. 1995;270:25696–25701. doi: 10.1074/jbc.270.43.25696. [PubMed] [Cross Ref]
34. Ko S.H., Lenkowski P.W., Lee H.C., Mounsey J.P., Patel M.K. Modulation of Na(v)1.5 by beta1- and beta3-subunit co-expression in mammalian cells. Pflugers Arch. 2005;449:403–412. doi: 10.1007/s00424-004-1348-4. [PubMed] [Cross Ref]
35. Johnson D., Montpetit M.L., Stocker P.J., Bennett E.S. The sialic acid component of the beta1 subunit modulates voltage-gated sodium channel function. J. Biol. Chem. 2004;279:44303–44310. doi: 10.1074/jbc.M408900200. [PubMed] [Cross Ref]
36. Fahmi A.I., et al. The sodium channel beta-subunit SCN3b modulates the kinetics of SCN5a and is expressed heterogeneously in sheep heart. J. Physiol. 2001;537:693–700. doi: 10.1113/jphysiol.2001.012691. [PubMed] [Cross Ref]
37. Makita N., Bennett P.B., Jr., George A.L., Jr. Voltage-gated Na+ channel beta 1 subunit mRNA expressed in adult human skeletal muscle, heart, and brain is encoded by a single gene. J. Biol. Chem. 1994;269:7571–7578. [PubMed]
38. Chen C., Cannon S.C. Modulation of Na+ channel inactivation by the beta 1 subunit: a deletion analysis. Pflugers Arch. 1995;431:186–195. doi: 10.1007/BF00410190. [PubMed] [Cross Ref]
39. McCormick K.A., et al. Molecular determinants of Na+ channel function in the extracellular domain of the beta1 subunit. J. Biol. Chem. 1998;273:3954–3962. doi: 10.1074/jbc.273.7.3954. [PubMed] [Cross Ref]
40. McCormick K.A., Srinivasan J., White K., Scheuer T., Catterall W.A. The extracellular domain of the beta1 subunit is both necessary and sufficient for beta1-like modulation of sodium channel gating. J. Biol. Chem. 1999;274:32638–32646. doi: 10.1074/jbc.274.46.32638. [PubMed] [Cross Ref]
41. Zimmer T., Benndorf K. The human heart and rat brain IIA Na+ channels interact with different molecular regions of the beta1 subunit. J. Gen. Physiol. 2002;120:887–895. doi: 10.1085/jgp.20028703. [PMC free article] [PubMed] [Cross Ref]
42. Malhotra J.D., Thyagarajan V., Chen C., Isom L.L. Tyrosine-phosphorylated and nonphosphorylated sodium channel beta1 subunits are differentially localized in cardiac myocytes. J. Biol. Chem. 2004;279:40748–40754. doi: 10.1074/jbc.M407243200. [PubMed] [Cross Ref]
43. Wong H.K., et al. beta Subunits of voltage-gated sodium channels are novel substrates of beta-site amyloid precursor protein-cleaving enzyme (BACE1) and gamma-secretase. J. Biol. Chem. 2005;280:23009–23017. doi: 10.1074/jbc.M414648200. [PubMed] [Cross Ref]
44. Kim D.Y., Ingano L.A., Carey B.W., Pettingell W.H., Kovacs D.M. Presenilin/gamma-secretase-mediated cleavage of the voltage-gated sodium channel beta2-subunit regulates cell adhesion and migration. J. Biol. Chem. 2005;280:23251–23261. doi: 10.1074/jbc.M412938200. [PubMed] [Cross Ref]
45. Kim D.Y., et al. BACE1 regulates voltage-gated sodium channels and neuronal activity. Nat. Cell Biol. 2007;9:755–764. doi: 10.1038/ncb1602. [PMC free article] [PubMed] [Cross Ref]
46. Miyazaki H., et al. BACE1 modulates filopodia-like protrusions induced by sodium channel beta4 subunit. Biochem. Biophys. Res. Commun. 2007;361:43–48. doi: 10.1016/j.bbrc.2007.06.170. [PubMed] [Cross Ref]
47. Wallace R.H., et al. Febrile seizures and generalized epilepsy associated with a mutation in the Na+-channel beta1 subunit gene SCN1B. Nat. Genet. 1998;19:366–370. doi: 10.1038/1252. [PubMed] [Cross Ref]
48. Chen C., et al. Mice lacking sodium channel beta1 subunits display defects in neuronal excitability, sodium channel expression, and nodal architecture. J. Neurosci. 2004;24:4030–4042. doi: 10.1523/JNEUROSCI.4139-03.2004. [PubMed] [Cross Ref]
49. Scheffer I.E., et al. Temporal lobe epilepsy and GEFS+ phenotypes associated with SCN1B mutations. Brain. 2007;130:100–109. doi: 10.1093/brain/awl272. [PubMed] [Cross Ref]
50. Audenaert D., et al. A deletion in SCN1B is associated with febrile seizures and early-onset absence epilepsy. Neurology. 2003;61:854–856. [PubMed]
51. Nashef L., Hindocha N., Makoff A. Risk factors in sudden death in epilepsy (SUDEP): the quest for mechanisms. Epilepsia. 2007;48:859–871. doi: 10.1111/j.1528-1167.2007.01082.x. [PubMed] [Cross Ref]
52. Rugg-Gunn F.J., Simister R.J., Squirrell M., Holdright D.R., Duncan J.S. Cardiac arrhythmias in focal epilepsy: a prospective long-term study. Lancet. 2004;364:2212–2219. doi: 10.1016/S0140-6736(04)17594-6. [PubMed] [Cross Ref]
53. Wichmann H.E., Gieger C., Illig T. KORA-gen — resource for population genetics, controls and a broad spectrum of disease phenotypes. Gesundheitswesen. 2005;67(Suppl. 1):S26–S30. [PubMed]
54. Gaborit N., Le Bouter S., Szuts V., Varro A., Escande D., Nattel S., Demolombe S. Regional and tissue specific transcript signatures of ion channel genes in the non-diseased human heart. J. Physiol. 2007;582:675–693. doi: 10.1113/jphysiol.2006.126714. [PubMed] [Cross Ref]
55. Livak K.J., Schmittgen T.D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods. 2001;25:402–408. doi: 10.1006/meth.2001.1262. [PubMed] [Cross Ref]

Articles from The Journal of Clinical Investigation are provided here courtesy of American Society for Clinical Investigation