1 We studied the effects of ranolazine, an antianginal agent with promise as an antiarrhythmic drug, on wild-type (WT) and long QT syndrome variant 3 (LQT-3) mutant Na þ channels expressed in human embryonic kidney (HEK) 293 cells and knock-in mouse cardiomyocytes and used site-directed mutagenesis to probe the site of action of the drug. 2 We find preferential ranolazine block of sustained vs peak Na þ channel current for LQT-3 mutant (DKPQ and Y1795C) channels (IC 50 ¼ 15 vs 135 mM) with similar results obtained in HEK 293 cells and knock-in myocytes. 3 Ranolazine block of both peak and sustained Na þ channel current is significantly reduced by mutation (F1760A) of a single residue previously shown to contribute critically to the binding site for local anesthetic (LA) molecules in the Na þ channel. 4 Ranolazine significantly decreases action potential duration (APD) at 50 and 90% repolarization by 2375 and 2773%, respectively, in DKPQ mouse ventricular myocytes but has little effect on APD of WT myocytes. 5 Computational modeling of human cardiac myocyte electrical activity that incorporates our voltage-clamp data predicts marked ranolazine-induced APD shortening in cells expressing LQT-3 mutant channels. 6 Our results demonstrate for the first time the utility of ranolazine as a blocker of sustained Na þ channel activity induced by inherited mutations that cause human disease and further, that these effects are very likely due to interactions of ranolazine with the receptor site for LA molecules in the sodium channel.
Defects of the SCN5A gene encoding the cardiac sodium channel ␣-subunit are associated with both the long QT-3 (LQT-3) subtype of long-QT syndrome and Brugada syndrome (BrS). One previously described SCN5A mutation (1795insD) in the C terminus results in a clinical phenotype combining QT prolongation and ST segment elevation, indicating a close interrelationship between the two disorders. Here we provide additional evidence that these two disorders are closely related. We report the analysis of two novel mutations on the same codon, Y1795C (LQT-3) and Y1795H (BrS), expressed in HEK 293 cells and characterized using whole-cell patch clamp procedures. We find marked and opposing effects on channel gating consistent with activity associated with the cellular basis of each clinical disorder. Y1795H speeds and Y1795C slows the onset of inactivation. The Y1795H, but not the Y1795C, mutation causes a marked negative shift in the voltage dependence of inactivation, and neither mutation affects the kinetics of the recovery from inactivation. Interestingly, both mutations increase the expression of sustained Na ؉ channel activity compared with wild type (WT) channels, although this effect is most pronounced for the Y1795C mutation, and both mutations promote entrance into an intermediate or a slowly developing inactivated state. These data confirm the key role of the C-terminal tail of the cardiac Na ؉ channel in the control of channel gating, illustrate how subtle changes in channel biophysics can have significant and distinct effects in human disease, and, additionally, provide further evidence of the close interrelationship between BrS and LQT-3 at the molecular level.Mutations of SCN5A, the gene coding for the ␣-subunit of the cardiac sodium channel, have been linked to the following four human syndromes: congenital long QT syndrome type 3 (LQT-3), 1 Brugada syndrome (BrS), different types of conduction block (2), and sudden infant death syndrome (3-7). Despite this classification based on clinical characteristics such as ECG findings, age of first presentation, triggers of events, and others, significant clinical and pathophysiological overlap between these different disorders have been noted recently. This observation has led to the proposition that these syndromes are in fact only different appearances of a "unique" SCN5A disease. The most significant overlap exists between LQT-3 and the BrS (8). The most frequent presentation in both disorders is a history of syncope or cardiac arrests with a very high lethality, occurring mainly during rest or sleep and with a poor response to anti-adrenergic treatment such as  receptor blockade (9 -12). However, the typical electrocardiographic manifestations of long-QT syndrome (QT interval prolongation) and BrS (ST segment elevation in leads V1 through V3) may coexist in the same patients, which raises questions about the actual differences between LQT3 and BrS (8). Moreover, in some LQT-3 patients, exposure to the sodium channel blocker flecainide generates an ECG pattern th...
Electrical activity in nerve, skeletal muscle, and heart requires finely tuned activity of voltage-gated Na+ channels that open and then enter a nonconducting inactivated state upon depolarization. Inactivation occurs when the gate, the cytoplasmic loop linking domains III and IV of the α subunit, occludes the open pore. Subtle destabilization of inactivation by mutation is causally associated with diverse human disease. Here we show for the first time that the inactivation gate is a molecular complex consisting of the III-IV loop and the COOH terminus (C-T), which is necessary to stabilize the closed gate and minimize channel reopening. When this interaction is disrupted by mutation, inactivation is destabilized allowing a small, but important, fraction of channels to reopen, conduct inward current, and delay cellular repolarization. Thus, our results demonstrate for the first time that physiologically crucial stabilization of inactivation of the Na+ channel requires complex interactions of intracellular structures and indicate a novel structural role of the C-T domain in this process.
Na+ channel blockers such as flecainide have found renewed usefulness in the diagnosis and treatment of two clinical syndromes arising from inherited mutations in SCN5A, the gene encoding the α subunit of the cardiac voltage–gated Na+ channel. The Brugada syndrome (BrS) and the LQT-3 variant of the Long QT syndrome are caused by disease-linked SCN5A mutations that act to change functional and pharmacological properties of the channel. Here we have explored a set of SCN5A mutations linked both to BrS and LQT-3 to determine what disease-modified channel properties underlie distinct responses to the Na+ channel blocker flecainide. We focused on flecainide block that develops with repetitive channel activity, so-called use-dependent block (UDB). Our results indicate that mutation-induced changes in the voltage-dependence of channel availability (inactivation) may act as determinants of flecainide block. The data further indicate that UDB by flecainide requires channel opening, but is not likely due to open channel block. Rather, flecainide appears to interact with inactivation states that follow depolarization-induced channel opening, and mutation-induced changes in channel inactivation will alter flecainide block independent of the disease to which the mutation is linked. Analysis of flecainide block of mutant channels linked to these rare disorders has provided novel insight into the molecular determinants of drug action.
This paper introduces an approach to performance animation that employs a small number of motion sensors to create an easy-to-use system for an interactive control of a full-body human character.Our key idea is to construct a series of online local dynamic models from a prerecorded motion database and utilize them to construct full-body human motion in a maximum a posteriori framework (MAP). We have demonstrated the effectiveness of our system by controlling a variety of human actions, such as boxing, golf swinging, and table tennis, in real time. Given an appropriate motion capture database, the results are comparable in quality to those obtained from a commercial motion capture system with a full set of motion sensors (e.g., XSens [2009]); however, our performance animation system is far less intrusive and expensive because it requires a small of motion sensors for full body control. We have also evaluated the performance of our system by leave-one-out-experiments and by comparing with two baseline algorithms.
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