Quantitative prediction of the arrhythmogenic effects of de novo hERG mutations in computational models of human ventricular tissues

Mutations to hERG which result in changes to the rapid delayed rectifier current I _Kr can cause long and short QT syndromes and are associated with an increased risk of cardiac arrhythmias. Experimental recordings of I _Kr reveal the effects of mutations at the channel level, but how these changes translate to the cell and tissue levels remains unclear. We used computational models of human ventricular myocytes and tissues to predict and quantify the effects that de novo hERG mutations would have on cell and tissue electrophysiology. Mutations that decreased I _Kr maximum conductance resulted in an increased cell and tissue action potential duration (APD) and a long QT interval on the electrocardiogram (ECG), whereas those that caused a positive shift in the inactivation curve resulted in a decreased APD and a short QT. Tissue vulnerability to re-entrant arrhythmias was correlated with transmural dispersion of repolarisation, and any change to this vulnerability could be inferred from the ECG QT interval or T wave peak-to-end time. Faster I _Kr activation kinetics caused cell APD alternans to appear over a wider range of pacing rates and with a larger magnitude, and spatial heterogeneity in these cellular alternans resulted in discordant alternans at the tissue level. Thus, from channel kinetic data, we can predict the tissue-level electrophysiological effects of any hERG mutations and identify how the mutation would manifest clinically, as either a long or short QT syndrome with or without an increased risk of alternans and re-entrant arrhythmias.     

Reduced levels of Cacna1c attenuate mesolimbic dopamine system function

Genetic variation in CACNA1C, which codes for the L‐type calcium channel (LTCC) Ca_v1.2, is associated with clinical diagnoses of bipolar disorder, depression and schizophrenia. Dysregulation of the mesolimbic‐dopamine (ML‐DA) system is linked to these syndromes and LTCCs are required for normal DAergic neurotransmission between the ventral tegmental area (VTA) and nucleus accumbens (NAc). It is unclear, however, how variations in CACNA1C genotype, and potential subsequent changes in expression levels in these regions, modify risk. Using constitutive and conditional knockout mice, and treatment with the LTCC antagonist nimodipine, we examined the role of Cacna1c in DA‐mediated behaviors elicited by psychomotor stimulants. Using fast‐scan cyclic voltammetry, DA release and reuptake in the NAc were measured. We find that subsecond DA release in Cacna1c haploinsufficient mice lacks normal sensitivity to inhibition of the DA transporter (DAT). Constitutive haploinsufficiency of Cacna1c led to attenuation of hyperlocomotion following acute administration of stimulants specific to DAT, and locomotor sensitization of these mice to the DAT antagonist GBR12909 did not reach the same level as wild‐type mice. The maintenance of sensitization to GBR12909 was attenuated by administration of nimodipine. Sensitization to GBR12909 was attenuated in mice with reduced Cacna1c selectively in the VTA but not in the NAc. Our findings show that Cacna1c is crucial for normal behavioral responses to DA stimulants and that its activity in the VTA is required for behavioral sensitization. Cacna1c likely exerts these effects through modifications to presynaptic ML‐DA system function.     

Calcium channels and channelopathies of the central nervous system

Several inherited human neurological disorders can be caused by mutations in genes encoding Ca²⁺ channel subunits. This review deals with known human and mouse calcium channelopathies of the central nervous system (CNS). The human diseases comprise: 1) a recessive retinal disorder, X-linked congenital stationary night blindness, associated with mutations in the CACNA1F gene, encoding α₁1.4 subunits of L-type channels; and 2) a group of rare allelic autosomal dominant human neurological disorders including familial hemiplegic migraine, episodic ataxia type 2, and spinocerebellar ataxia type 6, all associated with mutations in the CACNA1A gene, encoding α₁2.1 subunits of P/Q-type calcium channels. Mutations at the mouse orthologue of the CACNA1A gene cause a group of recessive neurological disorders, including the tottering, leaner, and rocker phenotypes with ataxia and absence epilepsy, and the rolling Nagoya phenotype with ataxia without seizures. Two other spontaneous mouse mutants with ataxia and absence epilepsy, lethargic and stargazer, have mutations in genes encoding a calcium channel auxiliary β subunit and a putative calcium channel auxiliary γ subunit. For each channelopathy, the review describes disease phenotype, channel genotype, and known functional consequences of the pathological mutations; in some cases, it also describes working hypothesis and/or speculations addressing the challenging question of how the alterations in channel function lead to selective cellular dysfunction and disease.     

A CACNA1C Variant Associated with Reduced Voltage-Dependent Inactivation,Increased CaV1.2 Channel Window Current,and Arrhythmogenesis

Mutations in CACNA1C that increase current through the Ca_V1.2 L-type Ca²⁺ channel underlie rare forms of long QT syndrome (LQTS), and Timothy syndrome (TS). We identified a variant in CACNA1C in a male child of Filipino descent with arrhythmias and extracardiac features by candidate gene sequencing and performed functional expression studies to electrophysiologically characterize the effects of the variant on Ca_V1.2 channels. As a baby, the subject developed seizures and displayed developmental delays at 30 months of age. At age 5 years, he displayed a QTc of 520 ms and experienced recurrent VT. Physical exam at 17 years of age was notable for microcephaly, short stature, lower extremity weakness and atrophy with hyperreflexia, spastic diplegia, multiple dental caries and episodes of rhabdomyolysis. Candidate gene sequencing identified a G>C transversion at position 5731 of CACNA1C (rs374528680) predicting a glycine>arginine substitution at residue 1911 (p.G1911R) of Ca_V1.2. The allele frequency of this variant is 0.01 in Malays, but absent in 984 Caucasian alleles and in the 1000 genomes project. In electrophysiological analyses, the variant decreased voltage-dependent inactivation, thus causing a gain of function of Ca_V1.2. We also observed a negative shift of V_1/2 of activation and positive shift of V_1/2 of channel inactivation, resulting in an increase of the window current. Together, these suggest a gain-of-function effect on Ca_V1.2 and suggest increased susceptibility for arrhythmias in certain clinical settings. The p.G1911R variant was also identified in a case of sudden unexplained infant death (SUID), for which an increasing number of clinical observations have demonstrated can be associated with arrhythmogenic mutations in cardiac ion channels. In summary, the combined effects of the CACNA1C variant to diminish voltage-dependent inactivation of Ca_V1.2 and increase window current expand our appreciation of mechanisms by which a gain of function of Ca_V1.2 can contribute to QT prolongation.     

Mouse models of SCN5A-related cardiac arrhythmias

Both gain- and loss-of-function mutations in the SCN5A gene, which encodes the α-subunit of the cardiac voltage-gated Na⁺ channel Na_v1.5, are well established to underlie hereditary arrhythmic syndromes (cardiac channelopathies) such as the type 3 long QT syndrome, cardiac conduction diseases, Brugada syndrome, sick sinus syndrome, atrial standstill and numerous overlap syndromes. Although patch-clamp studies in heterologous expression systems have provided important information to understand the genotype–phenotype relationships of these diseases, they could not clarify how mutations can be responsible for such a large spectrum of diseases, the late age of onset or the progressiveness of some of them, and for the overlapping syndromes. Genetically modified mice rapidly appeared as promising tools for understanding the pathophysiological sequence of cardiac SCN5A-related channelopathies and several mouse models have been established. Here, we review the results obtained on these models that, for most of them, convincingly recapitulate the clinical phenotypes of the patients but that also have their own limitations. Mouse models turn out to be powerful tools to elucidate the pathophysiological mechanisms of SCN5A-related diseases and offer the opportunity to investigate the cellular consequences of SCN5A mutations such as the remodelling of other gene expression that might participate in the overall phenotype and explain some of the differences among patients. Finally, they also constitute useful tools for future studies addressing as yet unanswered questions, such as the role of genetic and environmental modifiers on cardiac conduction and repolarisation.     

Fever-triggered ventricular arrhythmias in Brugada syndrome and type 2 long-QT syndrome

The risk for lethal ventricular arrhythmias is increased in individuals who carry mutations in genes that encode cardiac ion channels. Loss-of-function mutations in SCN5A, the gene encoding the cardiac sodium channel, are linked to Brugada syndrome (BrS). Arrhythmias in BrS are often preceded by coved-type ST-segment elevation in the right-precordial leads V1 and V2. Loss-of-function mutations in KCNH2, the gene encoding the cardiac ion channel that is responsible for the rapidly activating delayed rectifying potassium current, are linked to long-QT syndrome type 2 (LQT-2). LQT-2 is characterised by delayed cardiac repolarisation and rate-corrected QT interval (QTc) prolongation. Here, we report that the risk for ventricular arrhythmias in BrS and LQT-2 is further increased during fever. Moreover, we demonstrate that fever may aggravate coved-type ST-segment elevation in BrS, and cause QTc lengthening in LQT-2. Finally, we describe molecular mechanisms that may underlie the proarrhythmic effects of fever in BrS and LQT-2. (Neth Heart J 2010;18:165-9.)     

Dynamics of early afterdepolarization-mediated triggered activity in cardiac monolayers

Early afterdepolarizations (EADs) are voltage oscillations that occur during the repolarizing phase of the cardiac action potential and cause cardiac arrhythmias in a variety of clinical settings. EADs occur in the setting of reduced repolarization reserve and increased inward-over-outward currents, which intuitively explains the repolarization delay but does not mechanistically explain the time-dependent voltage oscillations that are characteristic of EADs. In a recent theoretical study, we identified a dual Hopf-homoclinic bifurcation as a dynamical mechanism that causes voltage oscillations during EADs, depending on the amplitude and kinetics of the L-type Ca(2+) channel (LTCC) current relative to the repolarizing K(+) currents. Here we demonstrate this mechanism experimentally. We show that cardiac monolayers exposed to the LTCC agonists BayK8644 and isoproterenol produce EAD bursts that are suppressed by the LTCC blocker nitrendipine but not by the Na(+) current blocker tetrodoxin, depletion of intracellular Ca(2+) stores with thapsigargin and caffeine, or buffering of intracellular Ca(2+) with BAPTA-AM. These EAD bursts exhibited a key dynamical signature of the dual Hopf-homoclinic bifurcation mechanism, namely, a gradual slowing in the frequency of oscillations before burst termination. A detailed cardiac action potential model reproduced the experimental observations, and identified intracellular Na(+) accumulation as the likely mechanism for terminating EAD bursts. Our findings in cardiac monolayers provide direct support for the Hopf-homoclinic bifurcation mechanism of EAD-mediated triggered activity, and raise the possibility that this mechanism may also contribute to EAD formation in clinical settings such as long QT syndromes, heart failure, and increased sympathetic output.     

Genetic determinants of QT interval variation and sudden cardiac death

Electrocardiographic QT interval prolongation or shortening is a risk factor for sudden cardiac death. The study of Mendelian syndromes in families with extreme long and short QT interval duration and ventricular arrhythmias has led to the identification of genes encoding ion channel proteins important in myocardial repolarization. Rare mutations in such ion channel genes do not individually contribute substantially to the population burden of ventricular arrhythmias and sudden cardiac death. Only now are studies systematically testing the relationship between common variants in these genes--or elsewhere in the genome--and QT interval variation and sudden cardiac death. Identification of genetic variation underlying myocardial repolarization could have important implications for the prevention of both sporadic and drug-induced arrhythmias.     

Cloning, sequencing and identification of single nucleotide polymorphisms of partial sequence on the porcine CACNA1S gene

CACNA1S gene encodes the α1 subunit of the calcium channel. The mutation of CACNA1S gene can cause hypokalemic periodic paralysis (HypoKPP) and maliglant hyperthermia synarome (MHS) in hu-man beings. Current research on CACNA1S was mainly in human being and model animal, but rarely in livestock and poultry. In this study, Yorkshire pigs (23), Pietrain pigs (30), Jinhua pigs (115) and the second generation (126) of crossbred of Jinhua and Pietrain were used. Primers were designed ac-cording to the sequence of human CACNA1S gene and PCR was carried out using pig genome DNA. PCR products were sequenced and compared with that of human, and then single nucleotide poly-morphisms (SNPs) were investigated by PCR-SSCP, while PCR-RFLP tests were performed to validate the mutations. Results indicated: (1) the 5211 bp DNA fragments of porcine CACNA1S gene were ac-quired (GenBank accession number: DQ767693 ) and the identity of the exon region was 82.6% be-tween human and pig; (2) fifty-seven mutations were found within the cloned sequences, among which 24 were in exon region; (3) the results of PCR-RFLP were in accordance with that of PCR-SSCP. Ac-cording to the EST of porcine CACNA1S gene published in GenBank (Bx914582, Bx666997), 8 of the 11 SNPs identified in the present study were consistent with the base difference between two EST frag-ments.     

CACNA1H missense mutations associated with amyotrophic lateral sclerosis alter Cav3.2 T-type calcium channel activity and reticular thalamic neuron firing

Amyotrophic lateral sclerosis (ALS) is a progressive neurodegenerative disease that affects nerve cells in the brain and the spinal cord. In a recent study by Steinberg and colleagues, 2 recessive missense mutations were identified in the Ca_v3.2 T-type calcium channel gene (CACNA1H), in a family with an affected proband (early onset, long duration ALS) and 2 unaffected parents. We have introduced and functionally characterized these mutations using transiently expressed human Ca_v3.2 channels in tsA-201 cells. Both of these mutations produced mild but significant changes on T-type channel activity that are consistent with a loss of channel function. Computer modeling in thalamic reticular neurons suggested that these mutations result in decreased neuronal excitability of thalamic structures. Taken together, these findings implicate CACNA1H as a susceptibility gene in amyotrophic lateral sclerosis.     

hERG1 channel activators: A new anti-arrhythmic principle

The cardiac action potential is the result of an orchestrated function of a number of different ion channels. Action potential repolarisation in humans relies on three potassium current components named I_Kr, I_Ks and I_K1 with party overlapping functions. The ion channel α-subunits conducting these currents are hERG1 (Kv11.1), KCNQ1 (Kv7.1) and Kir2.1. Loss-of-function in any of these currents can result in long QT syndrome. Long QT is a pro-arrhythmic disease with increased risk of developing lethal ventricular arrhythmias such as Torsade de Pointes and ventricular fibrillation. In addition to congenital long QT, acquired long QT can also constitute a safety risk. Especially unintended inhibition of the hERG1 channel constitutes a major concern in the development of new drugs. Based on this knowledge is has been speculated whether activation of the hERG1 channel could be anti-arrhythmic and thereby constitute a new principle in treatment of cardiac arrhythmogenic disorders. The first hERG1 channel agonist was reported in 2005 and a limited number of such compounds are now available. In the present text we review results obtained by hERG1 channel activation in a number of cardiac relevant settings from in vitro to in vivo. It is demonstrated how the principle of hERG1 channel activation under certain circumstances can constitute a new anti-arrhythmogenic principle. Finally, important conceptual differences between the short QT syndrome and the hERG1 channel activation, are evaluated.     

L-type channel inactivation balances the increased peak calcium current due to absence of Rad in cardiomyocytes

The L-type Ca²⁺ channel (LTCC) provides trigger calcium to initiate cardiac contraction in a graded fashion that is regulated by L-type calcium current (I_Ca,L) amplitude and kinetics. Inactivation of LTCC is controlled to fine-tune calcium flux and is governed by voltage-dependent inactivation (VDI) and calcium-dependent inactivation (CDI). Rad is a monomeric G protein that regulates I_Ca,L and has recently been shown to be critical to β-adrenergic receptor (β-AR) modulation of I_Ca,L. Our previous work showed that cardiomyocyte-specific Rad knockout (cRadKO) resulted in elevated systolic function, underpinned by an increase in peak I_Ca,L, but without pathological remodeling. Here, we sought to test whether Rad-depleted LTCC contributes to the fight-or-flight response independently of β-AR function, resulting in I_Ca,L kinetic modifications to homeostatically balance cardiomyocyte function. We recorded whole-cell I_Ca,L from ventricular cardiomyocytes from inducible cRadKO and control (CTRL) mice. The kinetics of I_Ca,L stimulated with isoproterenol in CTRL cardiomyocytes were indistinguishable from those of unstimulated cRadKO cardiomyocytes. CDI and VDI are both enhanced in cRadKO cardiomyocytes without differences in action potential duration or QT interval. To confirm that Rad loss modulates LTCC independently of β-AR stimulation, we crossed a β₁,β₂-AR double-knockout mouse with cRadKO, resulting in a Rad-inducible triple-knockout mouse. Deletion of Rad in cardiomyocytes that do not express β₁,β₂-AR still yielded modulated I_Ca,L and elevated basal heart function. Thus, in the absence of Rad, increased Ca²⁺ influx is homeostatically balanced by accelerated CDI and VDI. Our results indicate that the absence of Rad can modulate the LTCC without contribution of β₁,β₂-AR signaling and that Rad deletion supersedes β-AR signaling to the LTCC to enhance in vivo heart function.     

Rare Mutations of CACNB2 Found in Autism Spectrum Disease-Affected Families Alter Calcium Channel Function

Autism Spectrum Disorders (ASD) are complex neurodevelopmental diseases clinically defined by dysfunction of social interaction. Dysregulation of cellular calcium homeostasis might be involved in ASD pathogenesis, and genes coding for the L-type calcium channel subunits Ca_V1.2 (CACNA1C) and Ca_Vβ2 (CACNB2) were recently identified as risk loci for psychiatric diseases. Here, we present three rare missense mutations of CACNB2 (G167S, S197F, and F240L) found in ASD-affected families, two of them described here for the first time (G167S and F240L). All these mutations affect highly conserved regions while being absent in a sample of ethnically matched controls. We suggest the mutations to be of physiological relevance since they modulate whole-cell Ba²⁺ currents through calcium channels when expressed in a recombinant system (HEK-293 cells). Two mutations displayed significantly decelerated time-dependent inactivation as well as increased sensitivity of voltage-dependent inactivation. In contrast, the third mutation (F240L) showed significantly accelerated time-dependent inactivation. By altering the kinetic parameters, the mutations are reminiscent of the CACNA1C mutation causing Timothy Syndrome, a Mendelian disease presenting with ASD. In conclusion, the results of our first-time biophysical characterization of these three rare CACNB2 missense mutations identified in ASD patients support the hypothesis that calcium channel dysfunction may contribute to autism.     

Common Genetic Variation Near the Phospholamban Gene Is Associated with Cardiac Repolarisation: Meta-Analysis of Three Genome-Wide Association Studies

To identify loci affecting the electrocardiographic QT interval, a measure of cardiac repolarisation associated with risk of ventricular arrhythmias and sudden cardiac death, we conducted a meta-analysis of three genome-wide association studies (GWAS) including 3,558 subjects from the TwinsUK and BRIGHT cohorts in the UK and the DCCT/EDIC cohort from North America. Five loci were significantly associated with QT interval at P<1×10⁻⁶. To validate these findings we performed an in silico comparison with data from two QT consortia: QTSCD (n = 15,842) and QTGEN (n = 13,685). Analysis confirmed the association between common variants near NOS1AP (P = 1.4×10⁻⁸³) and the phospholamban (PLN) gene (P = 1.9×10⁻²⁹). The most associated SNP near NOS1AP (rs12143842) explains 0.82% variance; the SNP near PLN (rs11153730) explains 0.74% variance of QT interval duration. We found no evidence for interaction between these two SNPs (P = 0.99). PLN is a key regulator of cardiac diastolic function and is involved in regulating intracellular calcium cycling, it has only recently been identified as a susceptibility locus for QT interval. These data offer further mechanistic insights into genetic influence on the QT interval which may predispose to life threatening arrhythmias and sudden cardiac death.     

Novel SNPs of the bovine <Emphasis Type="Italic">CACNA2D1</Emphasis> gene and their association with carcass and meat quality traits

Calcium channel, voltage-dependent, alpha-2/delta subunit 1 (CACNA2D1) gene encodes a member of the alpha-2/delta subunit family, proteins are accessory molecules associated with voltage-gated calcium channels, and increase the density at the plasma membrane of calcium channels activated by high voltage. The main objective of the present study was to identify polymorphisms of CACNA2D1 gene, and to analyze associations between these polymorphisms and carcass and meat quality traits in cattle. In this study, through PCR-SSCP and DNA sequencing methods, two new allelic variant corresponding to the C → G and G → T mutations at positions 526740 and 537917 in the exon25 and exon35 of bovine CACNA2D1 gene, respectively, could be detected. SNP C526740G is a nonsynonymous mutation, resulting in a Cysteine (Cys) to Tryptophan (Trp) amino acid replacement and SNP G537917T resulting in an Aspartic (Asp) to Tyrosine (Tyr) amino acid replacement. The gene-specific SNP markers association analysis was investigated. The C526740G was significantly associated with Meat color (MC) (P = 0.0297) and Backfat thickness (BF) (P < 0.001). The G537917A indicated significant association with Dressing percentage (DP) (P = 0.0485). No significant association, however, was detected between any of the marker genotype and other traits measured in this study. Results from this study initially suggested that CACNA2D1 gene is one of the potential candidate genes influencing carcass and meat quality traits and gene-specific SNPs may be a useful marker for MAS programs in cattle breeding.     

Congenital Short QT Syndrome

The Short QT Syndrome is a recently described new genetic disorder, characterized by abnormally short QT interval, paroxysmal atrial fibrillation and life threatening ventricular arrhythmias. This autosomal dominant syndrome can afflict infants, children, or young adults; often a remarkable family background of cardiac sudden death is elucidated. At electrophysiological study, short atrial and ventricular refractory periods are found, with atrial fibrillation and polymorphic ventricular tachycardia easily induced by programmed electrical stimulation. Gain of function mutations in three genes encoding K⁺ channels have been identified, explaining the abbreviated repolarization seen in this condition: KCNH2 for I_kr (SQT1), KCNQ1 for I_ks (SQT2) and KCNJ2 for I_k1 (SQT3). The currently suggested therapeutic strategy is an ICD implantation, although many concerns exist for asymptomatic patients, especially in pediatric age. Pharmacological treatment is still under evaluation; quinidine has shown to prolong QT and reduce the inducibility of ventricular arrhythmias, but awaits additional confirmatory clinical data.     

Circadian rhythm in QT interval is preserved in mice deficient of potassium channel interacting protein 2

Potassium Channel Interacting Protein 2 (KChIP2) is suggested to be responsible for the circadian rhythm in repolarization duration, ventricular arrhythmias, and sudden cardiac death. We investigated the hypothesis that there is no circadian rhythm in QT interval in the absence of KChIP2. Implanted telemetric devices recorded electrocardiogram continuously for 5 days in conscious wild-type mice (WT, n = 9) and KChIP2^?/? mice (n = 9) in light:dark periods and in complete darkness. QT intervals were determined from all RR intervals and corrected for heart rate (QT₁₀₀ = QT/(RR/100)^1/2). Moreover, QT intervals were determined from complexes within the RR range of mean-RR ± 1% in the individual mouse (QT_mean-RR). We find that RR intervals are 125 ± 5 ms in WT and 123 ± 4 ms in KChIP2^?/? (p = 0.81), and QT intervals are 52 ± 1 and 52 ± 1 ms, respectively(p = 0.89). No ventricular arrhythmias or sudden cardiac deaths were observed. We find similar diurnal (light:dark) and circadian (darkness) rhythms of RR intervals in WT and KChIP2^?/? mice. Circadian rhythms in QT₁₀₀ intervals are present in both groups, but at physiological small amplitudes: 1.6 ± 0.2 and 1.0 ± 0.3 ms in WT and KChIP2^?/?, respectively (p = 0.15). A diurnal rhythm in QT₁₀₀ intervals was only found in WT mice. QT_mean-RR intervals display clear diurnal and circadian rhythms in both WT and KChIP2^?/?. The amplitude of the circadian rhythm in QT_mean-RR is 4.0 ± 0.3 and 3.1 ± 0.5 ms in WT and KChIP2^?/?, respectively (p = 0.16). In conclusion, KChIP2 expression does not appear to underlie the circadian rhythm in repolarization duration.     

Caveolae, ion channels and cardiac arrhythmias

Caveolae are specialized membrane microdomains enriched in cholesterol and sphingolipids which are present in multiple cell types including cardiomyocytes. Along with the essential scaffolding protein caveolin-3, a number of different ion channels and transporters have been localized to caveolae in cardiac myocytes including L-type Ca²⁺ channels (Ca_v1.2), Na⁺ channels (Na_v1.5), pacemaker channels (HCN4), Na⁺/Ca²⁺ exchanger (NCX1) and others. Closely associated with these channels are specific macromolecular signaling complexes that provide highly localized regulation of the channels. Mutations in the caveolin-3 gene (CAV3) have been linked with the congenital long QT syndrome (LQT9), and mutations in caveolar-localized ion channels may contribute to other inherited arrhythmias. Changes in the caveolar microdomain in acquired heart disease may also lead to dysregulation and dysfunction of ion channels, altering the risk of arrhythmias in conditions such as heart failure. This review highlights the existing evidence identifying and characterizing ion channels localized to caveolae in cardiomyocytes and their role in arrhythmogenesis.     

Characterization of CaV1.2 exon 33 heterozygous knockout mice and negative correlation between Rbfox1 and CaV1.2 exon 33 expressions in human heart failure

Recently, we reported that homozygous deletion of alternative exon 33 of Ca_V1.2 calcium channel in the mouse resulted in ventricular arrhythmias arising from increased Ca_V1.2_Δ33 I_CaL current density in the cardiomyocytes. We wondered whether heterozygous deletion of exon 33 might produce cardiac phenotype in a dose-dependent manner, and whether the expression levels of RNA splicing factors known to regulate alternative splicing of exon 33 might change in human heart failure. Unexpectedly, we found that exon 33^+/? cardiomyocytes showed similar Ca_V1.2 channel properties as wild-type cardiomyocyte, even though Ca_V1.2_Δ33 channels exhibit a gain-in-function. In human hearts, we found that the mRNA level of splicing factor Rbfox1, but not Rbfox2, was downregulated in dilated cardiomyopathy, and CACNA1C mRNA level was dramatically decreased in the both of dilated and ischemic cardiomyopathy. These data imply Rbfox1 may be involved in the development of cardiomyopathies via regulating the alternative splicing of Ca_V1.2 exon 33. (149 words)