Identification of a novel loss-of-function calcium channel gene mutation in short QT syndrome (SQTS6)

Short QT syndrome (SQTS) is a genetically determined ion-channel disorder, which may cause malignant tachyarrhythmias and sudden cardiac death. Thus far, mutations in five different genes encoding potassium and calcium channel subunits have been reported. We present, for the first time, a novel loss-of-function mutation coding for an L-type calcium channel subunit.

Duplication of the sodium channel gene cluster on 2q24 in children with early onset epilepsy

Sodium channel gene aberrations are associated with a wide range of seizure disorders, particularly Dravet syndrome. They usually consist of missense or truncating gene mutations or deletions. Duplications involving multiple genes encoding for different sodium channels are not widely known. This article summarizes the clinical, radiologic, and genetic features of patients with 2q24 duplication involving the sodium channel gene cluster.

A Spanish founder mutation in the chloride channel gene, CLCNKB, as a cause of atypical Bartter syndrome in adult age

BACKGROUND: Mutations in the chloride channel gene, CLCNKB, usually cause classic Bartter syndrome (cBS) or a mixed Bartter-Gitelman phenotype in the first years of life. METHODS: We report an adult woman with atypical BS caused by a homozygous missense mutation, A204T, in the CLCNKB gene, which has previously been described as the apparently unique cause of cBS in Spain. RESULTS: The evaluation of this patient revealed an overlap of phenotypic features ranging from severe biochemical and systemic disturbances typical of cBS to scarce symptoms and diagnosis in the adult age typical of Gitelman syndrome. The tubular disease caused a dramatic effect on mental, growth and puberal development leading to low IQ, final short stature and abnormal ovarian function. Furthermore, low serum PTH concentrations with concomitant nephrocalcinosis and normocalcaemia were observed. Both ovarian function and serum PTH levels were normalized after treatment with cyclooxygenase inhibitors. CONCLUSIONS: The present report confirms a weak genotype-phenotype correlation in patients with CLCNKB mutations and supports the founder effect of the A204T mutation in Spain. In our country, the genetic diagnosis of adult patients with hereditary hypokalaemic tubulopathies should include a screening of A204T mutation in the CLCNKB gene.

Marked disturbance of calcium homeostasis in mice with targeted disruption of the Trpv6 calcium channel gene

We report the phenotype of mice with targeted disruption of the Trpv6 (Trpv6 KO) epithelial calcium channel. The mice exhibit disordered Ca(2+) homeostasis, including defective intestinal Ca(2+) absorption, increased urinary Ca(2+) excretion, decreased BMD, deficient weight gain, and reduced fertility. Although our Trpv6 KO affects the closely adjacent EphB6 gene, the phenotype reported here is not related to EphB6 dysfunction. INTRODUCTIOn: The mechanisms underlying intestinal Ca(2+) absorption are crucial for overall Ca(2+) homeostasis, because diet is the only source of all new Ca(2+) in the body. Trpv6 encodes a Ca(2+)-permeable cation channel responsible for vitamin D-dependent intestinal Ca(2+) absorption. Trpv6 is expressed in the intestine and also in the skin, placenta, kidney, and exocrine organs. MATERIALS AND METHODS: To determine the in vivo function of TRPV6, we generated mice with targeted disruption of the Trpv6 (Trpv6 KO) gene. RESULTS: Trpv6 KO mice are viable but exhibit disordered Ca(2+) homeostasis, including a 60% decrease in intestinal Ca(2+) absorption, deficient weight gain, decreased BMD, and reduced fertility. When kept on a regular (1% Ca(2+)) diet, Trpv6 KO mice have deficient intestinal Ca(2+) absorption, despite elevated levels of serum PTH (3.8-fold) and 1,25-dihydroxyvitamin D (2.4-fold). They also have decreased urinary osmolality and increased Ca(2+) excretion. Their serum Ca(2+) is normal, but when challenged with a low (0.25%) Ca(2+) diet, Trpv6 KO mice fail to further increase serum PTH and vitamin D, ultimately developing hypocalcemia. Trpv6 KO mice have normal urinary deoxypyridinoline excretion, although exhibiting a 9.3% reduction in femoral mineral density at 2 months of age, which is not restored by treatment for 1 month with a high (2%) Ca(2+) "rescue" diet. In addition to their deranged Ca(2+) homeostasis, the skin of Trpv6 KO mice has fewer and thinner layers of stratum corneum, decreased total Ca(2+) content, and loss of the normal Ca(2+) gradient. Twenty percent of all Trpv6 KO animals develop alopecia and dermatitis. CONCLUSIONS: Trpv6 KO mice exhibit an array of abnormalities in multiple tissues/organs. At least some of these are caused by tissue-specific mechanisms. In addition, the kidneys and bones of Trpv6 KO mice do not respond to their elevated levels of PTH and 1,25-dihydroxyvitamin D. These data indicate that the TRPV6 channel plays an important role in Ca(2+) homeostasis and in other tissues not directly involved in this process.

Patients with biallelic mutations in the chloride channel gene CLCNKB: long-term management and outcome

BACKGROUND: Little information on the management and long-term follow-up of patients with biallelic mutations in the chloride channel gene CLCNKB is available. METHODS: Long-term follow-up was evaluated from 5.0 to 24 years (median, 14 years) after diagnosis in 13 patients with homozygous (n = 10) or compound heterozygous (n = 3) mutations. RESULTS: Medical treatment at last follow-up control included supplementation with potassium in 12 patients and sodium in 2 patients and medical treatment with indomethacin in 9 patients. At the end of follow-up, body height was 2.0 standard deviation score or less in 6 patients; 2 of these patients had growth hormone deficiency. Body weight (<or=2.0 standard deviation score in 6 patients) significantly increased (P < 0.05) at the end of follow-up in comparison to values at diagnosis. Nonpostural persistent proteinuria was present in 6 patients, and 4 patients had a glomerular filtration rate less than 75 mL/min/1.73 m(2) (<1.25 mL/s). CONCLUSION: These data show that some patients with biallelic mutations in the chloride channel gene CLCNKB tend to present with pathological proteinuria and impaired kidney function after a median follow-up of 14 years, and growth retardation is common and sometimes related to growth hormone deficiency in these patients.

F90927: a new member in the class of cardioactive steroids

F90927 is a newly developed cardioactive drug with a steroid-like structure. It acts directly and agonistically on the cardiac L-type Ca2+ channel by shifting its voltage-dependent activation toward more negative potentials. This leads to an increased influx of Ca2+ and, therefore, to a stronger contraction; however, no arrhythmias occur. Calcium current stimulation can already be observed at nanomolar concentrations, but higher concentrations of F90927 elevate intracellular Ca2+ concentration, causing a reduction of the myocardial compliance and an increased diastolic blood pressure. Vessels also react to F90927 and contract in its presence. Binding of F90927 with the L-type Ca2+ channel presumably occurs in the vicinity of the transmembrane domains III and IV of the alpha1 subunit. F90927 exhibits no use dependence and interacts with Ca2+ channel inhibitors of all three known classes of channel modulators (dihydropyridines, phenylalkylamines, and benzothiazepines), suggesting that it is a member of a new class of Ca2+ channel modulators. Due to its adverse effects on blood pressure and vessel contraction, F90927 is not an ideal drug candidate. It has, however, some unique properties, which makes it a promising tool to study the function of the L-type Ca2+ channel.

Increased expression of the auxiliary beta2-subunit of ventricular L-type Ca2+ channels leads to single-channel activity characteristic of heart failure

BACKGROUND: Increased activity of single ventricular L-type Ca(2+)-channels (L-VDCC) is a hallmark in human heart failure. Recent findings suggest differential modulation by several auxiliary beta-subunits as a possible explanation. METHODS AND RESULTS: By molecular and functional analyses of human and murine ventricles, we find that enhanced L-VDCC activity is accompanied by altered expression pattern of auxiliary L-VDCC beta-subunit gene products. In HEK293-cells we show differential modulation of single L-VDCC activity by coexpression of several human cardiac beta-subunits: Unlike beta(1) or beta(3) isoforms, beta(2a) and beta(2b) induce a high-activity channel behavior typical of failing myocytes. In accordance, beta(2)-subunit mRNA and protein are up-regulated in failing human myocardium. In a model of heart failure we find that mice overexpressing the human cardiac Ca(V)1.2 also reveal increased single-channel activity and sarcolemmal beta(2) expression when entering into the maladaptive stage of heart failure. Interestingly, these animals, when still young and non-failing ("Adaptive Phase"), reveal the opposite phenotype, viz: reduced single-channel activity accompanied by lowered beta(2) expression. Additional evidence for the cause-effect relationship between beta(2)-subunit expression and single L-VDCC activity is provided by newly engineered, double-transgenic mice bearing both constitutive Ca(V)1.2 and inducible beta(2) cardiac overexpression. Here in non-failing hearts induction of beta(2)-subunit overexpression mimicked the increase of single L-VDCC activity observed in murine and human chronic heart failure. CONCLUSIONS: Our study presents evidence of the pathobiochemical relevance of beta(2)-subunits for the electrophysiological phenotype of cardiac L-VDCC and thus provides an explanation for the single L-VDCC gating observed in human and murine heart failure.

A deleterious gene-by-environment interaction imposed by calcium channel blockers in Marfan syndrome.

Calcium channel blockers (CCBs) are prescribed to patients with Marfan syndrome for prophylaxis against aortic aneurysm progression, despite limited evidence for their efficacy and safety in the disorder. Unexpectedly, Marfan mice treated with CCBs show accelerated aneurysm expansion, rupture, and premature lethality. This effect is both extracellular signal-regulated kinase (ERK1/2) dependent and angiotensin-II type 1 receptor (AT1R) dependent. We have identified protein kinase C beta (PKCβ) as a critical mediator of this pathway and demonstrate that the PKCβ inhibitor enzastaurin, and the clinically available anti-hypertensive agent hydralazine, both normalize aortic growth in Marfan mice, in association with reduced PKCβ and ERK1/2 activation. Furthermore, patients with Marfan syndrome and other forms of inherited thoracic aortic aneurysm taking CCBs display increased risk of aortic dissection and need for aortic surgery, compared to patients on other antihypertensive agents.

Cardiac sodium channel Na(v)1.5 and interacting proteins: Physiology and pathophysiology

The cardiac voltage-gated Na(+) channel Na(v)1.5 generates the cardiac Na(+) current (INa). Mutations in SCN5A, the gene encoding Na(v)1.5, have been linked to many cardiac phenotypes, including the congenital and acquired long QT syndrome, Brugada syndrome, conduction slowing, sick sinus syndrome, atrial fibrillation, and dilated cardiomyopathy. The mutations in SCN5A define a sub-group of Na(v)1.5/SCN5A-related phenotypes among cardiac genetic channelopathies. Several research groups have proposed that Na(v)1.5 may be part of multi-protein complexes composed of Na(v)1.5-interacting proteins which regulate channel expression and function. The genes encoding these regulatory proteins have also been found to be mutated in patients with inherited forms of cardiac arrhythmias. The proteins that associate with Na(v)1.5 may be classified as (1) anchoring/adaptor proteins, (2) enzymes interacting with and modifying the channel, and (3) proteins modulating the biophysical properties of Na(v)1.5 upon binding. The aim of this article is to review these Na(v)1.5 partner proteins and to discuss how they may regulate the channel's biology and function. These recent investigations have revealed that the expression level, cellular localization, and activity of Na(v)1.5 are finely regulated by complex molecular and cellular mechanisms that we are only beginning to understand.

Nerve excitability studies characterize Kv1.1 fast potassium channel dysfunction in patients with episodic ataxia type 1

Episodic ataxia type 1 is a neuronal channelopathy caused by mutations in the KCNA1 gene encoding the fast K(+) channel subunit K(v)1.1. Episodic ataxia type 1 presents with brief episodes of cerebellar dysfunction and persistent neuromyotonia and is associated with an increased incidence of epilepsy. In myelinated peripheral nerve, K(v)1.1 is highly expressed in the juxtaparanodal axon, where potassium channels limit the depolarizing afterpotential and the effects of depolarizing currents. Axonal excitability studies were performed on patients with genetically confirmed episodic ataxia type 1 to characterize the effects of K(v)1.1 dysfunction on motor axons in vivo. The median nerve was stimulated at the wrist and compound muscle action potentials were recorded from abductor pollicis brevis. Threshold tracking techniques were used to record strength-duration time constant, threshold electrotonus, current/threshold relationship and the recovery cycle. Recordings from 20 patients from eight kindreds with different KCNA1 point mutations were compared with those from 30 normal controls. All 20 patients had a history of episodic ataxia and 19 had neuromyotonia. All patients had similar, distinctive abnormalities: superexcitability was on average 100% higher in the patients than in controls (P < 0.00001) and, in threshold electrotonus, the increase in excitability due to a depolarizing current (20% of threshold) was 31% higher (P < 0.00001). Using these two parameters, the patients with episodic ataxia type 1 and controls could be clearly separated into two non-overlapping groups. Differences between the different KCNA1 mutations were not statistically significant. Studies of nerve excitability can identify K(v)1.1 dysfunction in patients with episodic ataxia type 1. The simple 15 min test may be useful in diagnosis, since it can differentiate patients with episodic ataxia type 1 from normal controls with high sensitivity and specificity.

Nedd4-2-dependent ubiquitylation and regulation of the cardiac potassium channel hERG1

The voltage-gated cardiac potassium channel hERG1 (human ether-à-gogo-related gene 1) plays a key role in the repolarization phase of the cardiac action potential (AP). Mutations in its gene, KCNH2, can lead to defects in the biosynthesis and maturation of the channel, resulting in congenital long QT syndrome (LQTS). To identify the molecular mechanisms regulating the density of hERG1 channels at the plasma membrane, we investigated channel ubiquitylation by ubiquitin ligase Nedd4-2, a post-translational regulatory mechanism previously linked to other ion channels. We found that whole-cell hERG1 currents recorded in HEK293 cells were decreased upon neural precursor cell expressed developmentally down-regulated 4-2 (Nedd4-2) co-expression. The amount of hERG1 channels in total HEK293 lysates and at the cell surface, as assessed by Western blot and biotinylation assays, respectively, were concomitantly decreased. Nedd4-2 and hERG1 interact via a PY motif located in the C-terminus of hERG1. Finally, we determined that Nedd4-2 mediates ubiquitylation of hERG1 and that deletion of this motif affects Nedd4-2-dependent regulation. These results suggest that ubiquitylation of the hERG1 protein by Nedd4-2, and its subsequent down-regulation, could represent an important mechanism for modulation of the duration of the human cardiac action potential.

Cardiac Sodium Channel Nav1.5 Mechanosensitivity is Inhibited by Ranolazine

The cardiac action potential (AP) is initiated by the depolarizing inward sodium current (I(Na)). The pore-forming subunit of the cardiac sodium channel, Na(v)1.5, is the main ion channel that conducts I(Na) in cardiac cells. Despite the large number of studies investigating Na(v)1.5, year after year, we are still learning new aspects regarding its roles in normal cardiac function and in diseased states. The clinical relevance of this channel cannot be understated. The cardiac I(Na) is the target of the class 1 anti-arrhythmic drugs(1), which are nowadays less frequently prescribed because of their well-documented pro-arrhythmic properties(2). In addition, since the first description in 1995 by Keating's group(3) of mutations in patients suffering from congenital long QT syndrome (LQTS) type 3, several hundred genetic variants in SCN5A, the gene coding for Na(v)1.5, have been reported and investigated(4). Interestingly, many of these genetic variants have been found in patients with diverse cardiac manifestations(5) such as congenital LQTS type 3, Brugada syndrome, conduction disorders, and more recently, atrial fibrillation and dilated cardiomyopathy. This impressive list underlines the importance of Na(v)1.5 in cardiac pathologies and raises the question about possible unknown roles and regulatory mechanisms of this channel in cardiac cells. Recent studies have provided experimental evidence that the function of Na(v)1.5, among many other described regulatory mechanisms(6), is also modulated by the mechanical stretch of the membrane in which it is embedded(7), thus suggesting that Na(v)1.5, like other ion channels, is "mechanosensitive". What does this mean? (SELECT FULL TEXT TO CONTINUE).

Stereoselective Inhibition of the hERG1 Potassium Channel

A growing number of drugs have been shown to prolong cardiac repolarization, predisposing individuals to life-threatening ventricular arrhythmias known as Torsades de Pointes. Most of these drugs are known to interfere with the human ether à-gogo related gene 1 (hERG1) channel, whose current is one of the main determinants of action potential duration. Prolonged repolarization is reflected by lengthening of the QT interval of the electrocardiogram, as seen in the suitably named drug-induced long QT syndrome. Chirality (presence of an asymmetric atom) is a common feature of marketed drugs, which can therefore exist in at least two enantiomers with distinct three-dimensional structures and possibly distinct biological fates. Both the pharmacokinetic and pharmacodynamic properties can differ between enantiomers, as well as also between individuals who take the drug due to metabolic polymorphisms. Despite the large number of reports about drugs reducing the hERG1 current, potential stereoselective contributions have only been scarcely investigated. In this review, we present a non-exhaustive list of clinically important molecules which display chiral toxicity that may be related to hERG1-blocking properties. We particularly focus on methadone cardiotoxicity, which illustrates the importance of the stereoselective effect of drug chirality as well as individual variations resulting from pharmacogenetics. Furthermore, it seems likely that, during drug development, consideration of chirality in lead optimization and systematic assessment of the hERG1 current block with all enantiomers could contribute to the reduction of the risk of drug-induced LQTS.

A missense mutation in the skeletal muscle chloride channel 1 (CLCN1) as candidate causal mutation for congenital myotonia in a New Forest pony

A 7-month-old New Forest foal presented for episodes of recumbency and stiffness with myotonic discharges on electromyography. The observed phenotype resembled congenital myotonia caused by CLCN1 mutations in goats and humans. Mutation of the CLCN1 gene was considered as possible cause and mutation analysis was performed. The affected foal was homozygous for a missense mutation (c.1775A>C, p.D592A) located in a well conserved domain of the CLCN1 gene. The mutation showed a recessive mode of inheritance within the reported pony family. Therefore, this CLCN1 polymorphism is considered to be a possible cause of congenital myotonia.

Cardiac sodium channel NaV1.5 distribution in myocytes via interacting proteins: the multiple pool model

The cardiac sodium current (INa) is responsible for the rapid depolarization of cardiac cells, thus allowing for their contraction. It is also involved in regulating the duration of the cardiac action potential (AP) and propagation of the impulse throughout the myocardium. Cardiac INa is generated by the voltage-gated Na(+) channel, NaV1.5, a 2016-residue protein which forms the pore of the channel. Over the past years, hundreds of mutations in SCN5A, the human gene coding for NaV1.5, have been linked to many cardiac electrical disorders, including the congenital and acquired long QT syndrome, Brugada syndrome, conduction slowing, sick sinus syndrome, atrial fibrillation, and dilated cardiomyopathy. Similar to many membrane proteins, NaV1.5 has been found to be regulated by several interacting proteins. In some cases, these different proteins, which reside in distinct membrane compartments (i.e. lateral membrane vs. intercalated disks), have been shown to interact with the same regulatory domain of NaV1.5, thus suggesting that several pools of NaV1.5 channels may co-exist in cardiac cells. The aim of this review article is to summarize the recent works that demonstrate its interaction with regulatory proteins and illustrate the model that the sodium channel NaV1.5 resides in distinct and different pools in cardiac cells. This article is part of a Special Issue entitled: Cardiomyocyte Biology: Cardiac Pathways of Differentiation, Metabolism and Contraction.