Action Potential Plateau Amplitude Research Articles

Disrupted CaV1.2 selectivity causes overlapping long QT and Brugada syndrome phenotypes in the CACNA1C-E1115K iPS cell model

A missense mutation in the α1c subunit of voltage-gated L-type Ca2+ channel-coding CACNA1C-E1115K, located in the Ca2+ selectivity site, causes a variety of arrhythmogenic phenotypes. We aimed to investigate the electrophysiological features and pathophysiological mechanisms of CACNA1C-E1115K in patient-specific induced pluripotent stem cell (iPSC)-derived cardiomyocytes (CMs). We generated iPSCs from a patient carrying heterozygous CACNA1C-E1115K with overlapping phenotypes of long QT syndrome, Brugada syndrome, and mild cardiac dysfunction. Electrophysiological properties were investigated using iPSC-CMs. We used iPSCs from a healthy individual and an isogenic iPSC line corrected using CRISPR-Cas9-mediated gene editing as controls. A mathematical E1115K-CM model was developed using a human ventricular cell model. Patch-clamp analysis revealed that E1115K-iPSC-CMs exhibited reduced peak Ca2+ current density and impaired Ca2+ selectivity with an increased permeability to monovalent cations. Consequently, E1115K-iPSC-CMs showed decreased action potential plateau amplitude, longer action potential duration (APD), and a higher frequency of early afterdepolarization compared with controls. In optical recordings examining the antiarrhythmic drug effect, late Na+ channel current (INaL) inhibitors (mexiletine and GS-458967) shortened APDs specifically in E1115K-iPSC-CMs. The AP-clamp using a voltage command obtained from E1115K-iPSC-CMs with lower action potential plateau amplitude and longer APD confirmed the upregulation of INaL. An in silico study recapitulated the invitro electrophysiological properties. Our iPSC-based analysis in CACNA1C-E1115K with disrupted CaV1.2 selectivity demonstrated that the aberrant currents through the mutant channels carried by monovalent cations resulted in specific action potential changes, which increased endogenous INaL, thereby synergistically contributing to the arrhythmogenic phenotype.

Molecular mechanisms of ionic remodeling in L-type voltage dependent calcium channel of age-related change in atrial of canine

PurposeThe present study aims to investigate the molecular basis of L-type voltage dependent calcium channel (LVDCC) in adult and aged canine.MethodsThe action potential duration (APD90), amplitude of action potential plateau,...

Regional variations in action potentials and transient outward current in myocytes isolated from rabbit left ventricle.

1. Regional variations in the shape of early repolarization of the action potential have been correlated to differences in transient outward K+ current, I(t), in myocytes isolated from the epicardial surface, the endocardial trabeculae and the papillary muscles of rabbit left ventricles. Temperature was 35 degrees C during whole-cell, and 22-23 degrees C during cell-attached experiments. 2. Membrane resting potentials were very similar regionally. At 0.1 Hz stimulation the action potential plateau amplitude in papillary muscle cells was significantly higher (104.7 mV) than in epicardial cells (96.47 mV). Exposure to 4-aminopyridine or increases in the rate of stimulation from 0.1 Hz to 3.3 Hz increased plateau height and diminished the initial notch on repolarization. These effects were correlated to the magnitude of I(t) in these cells. At low rates of stimulation I(t) caused a 'spike and dome' morphology of the action potential. 3. Voltage clamp experiments confirmed a higher current density of I(t) in epicardial cells (7.66 pA/pF at +20 mV) than in endocardial (6.45 pA/pF) or papillary muscle cells (3.69 pA/pF). I(t) at 35 degrees C was faster and larger than previously reported and individual currents inactivated almost completely during 100 ms pulses to plateau potentials. No differences in the kinetics or voltage dependence of whole-cell currents were found. Thus, the half-inactivation potential was -37.8 mV in cells from all three regions. 4. Cell-attached recordings from endocardial and epicardial cells showed very similar single-channel amplitudes, burst open probabilities and ensemble averages. The peak channel open probability soon after the start of depolarizing voltage clamp pulses did not change between cell types (P approximately 0.8). The slope conductance of I(t) channels was 13.0 pS with an intercept near the resting potential of the cell. 5. We conclude that regional variations in the shape of initial repolarization in cells from rabbit left ventricle are caused by variations in the magnitude of the transient outward K+ current, I(t). Epicardial cells have the largest, and papillary muscle cells the smallest I(t). The differences are not explained by alterations in the whole-cell kinetics or single-channel kinetics and conductance. The most likely explanation for variations in whole-cell current density is therefore a decrease in channel density in endocardium and papillary muscle compared with epicardial tissue. We estimate the density of I(t) channels per cell to be 1495 (one per 3-4 micron2) in epicardium, 1175 (one per 4-5 micron2) in endocardium, and 875 (one per 6 micron2) in papillary muscle cells.

Effects of caffeine and ryanodine on depression of post‐rest tension development produced by Bay K 8644 in canine ventricular muscle

1. Post-rest inotropy in canine ventricular myocardium has proved a useful indicator of sarcoplasmic reticular calcium release. This phenomenon is converted to rest depression by the calcium channel activator (agonist), Bay K 8644 as well as other chemically diverse agents such as caffeine and ryanodine. 2. Rapid cooling contractures and post-rest contraction amplitude were used as independent measures of sarcoplasmic reticular calcium content and release. Simultaneous recordings of transmembrane action potentials and their accompanying contractions were obtained to determine the association between electrophysiological and mechanical events. The present study was designed to elucidate the mechanism by which Bay K 8644, caffeine and ryanodine alter force production after variable periods of rest. 3. Bay K 8644 (1 microM) increased steady state contraction in response to a constant train of stimulation, caused rest-depression after 2 and 8 min rest, prolonged action potential duration and increased action potential plateau amplitude. Augmented steady state tension was not accompanied by any change in time to peak tension or rapid cooling contracture amplitude. However, the post-rest rapid cooling contracture was greatly diminished compared to that observed prior to Bay K 8644 treatment. 4. Caffeine (3 and 5 mM) caused rest-depression with an increase in steady state contraction amplitude. Along with this there was a slight decrease in action potential duration and plateau amplitude and an increase in time to peak tension. The rapid cooling contractures were virtually abolished at all conditioning intervals. The effect of caffeine on twitch tension and cooling contracture is consistent with the ability of this compound to inhibit calcium sequestration by the sarcoplasmic reticulum. 5. A combination of Bay K 8644 and caffeine caused significantly less rest-depression than that seen with Bay K 8644 alone. The augmented twitch tension was accompanied by a long time to peak tension and action potential duration. However, there was no increase in the amplitude of the rapid cooling contracture, either after a regular train of stimulation or after rest, compared to that seen after Bay K 8644. 6. Ryanodine (10 nM), produced rest-depression, reduced steady state twitch tension and augmented the rest-depression produced by Bay K 8644. The steady state rapid cooling contracture remained unchanged when both agents were present simultaneously, while the post-rest rapid cooling contracture was significantly depressed compared to that observed with Bay K 8644 alone. 7. Bay K 8644 and ryanodine appear to have similar actions with respect to promoting diastolic loss of calcium from the sarcoplasmic reticulum. Although caffeine also decreases post-rest potentiation, it antagonizes rest-depression caused by Bay K 8644. The data from these experiments suggest that this reversal is a result of depressed intracellular calcium buffering and enhanced myofilament sensitivity produced by caffeine in the presence of increased transmembrane calcium influx promoted by Bay K 8644.

Impairment of Ca release from mammalian ventricular sarcoplasmic reticulum by the calcium channel agonist Bay K 8644

Positive inotropic concentration of the Ca-channel agonist, Bay K 8644, depressed contraction of canine right ventricular trabecula immediately after a rest period of 8 min, without decreasing action potential plateau amplitude. In contrast, high external Ca and ouabagenin caused only a slight decrease in post-rest contraction. Bay K 8644-induced post-rest depression was inversely related to the extracellular Ca concentration. Hence it could not be due to cellular Ca overload. Since post-rest potentiation is due to increased contribution of Ca from the sarcoplasmic reticulum, these results suggest that Bay K 8644 decrease the amount of releasable Ca from this structure during rest.

Excitation-contraction coupling in myocardium of nonhibernating and hibernating chipmunks: effects of isoprenaline, a high calcium medium, and ryanodine.

The electromechanical responses to isoprenaline and a high calcium medium on cardiac muscles from nonhibernating and hibernating chipmunks were studied in the presence and the absence of ryanodine. In nonhibernating animal preparations, isoprenaline (5 X 10(-8) M) caused a marked positive inotropic effect with an increase in the amplitude of the action potential plateau and augmented the slow action potential in muscle depolarized with 26 mM K+. In hibernating animal preparations, isoprenaline failed to cause a positive inotropic effect in spite of an increase in the amplitude of action potential plateau and slow action potentials. Similar inotropic effects were caused in the two preparations when extracellular calcium was raised to 6 mM, but action potential plateau and slow action potentials of the two preparations were less affected by this procedure; only in nonhibernating animal preparations was the amplitude of the slow action potential slightly increased. Ryanodine (2 X 10(-6) M) partially inhibited the contraction but augmented the action potential plateau and slow action potentials in nonhibernating animal preparations, while in hibernating animal preparations, it eliminated the contraction and severely inhibited the action potential plateau and slow action potentials. The electrical effects of isoprenaline on the two preparations and of a high calcium medium on nonhibernating animal preparations were more pronounced in the presence of ryanodine than in the absence of it. However, the electrical activity on hibernating animal preparations was unaffected by a high calcium medium in either the presence or absence of ryanodine.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcium channel block and recovery from block in mammalian ventricular muscle treated with organic channel inhibitors.

The effects of a bradycardic agent, AQA 39, on the action potential, tension and membrane current of cat ventricular muscle have been compared with the effects of 1. AQA 39 and D600 both lengthened the action potential duration; neither drug affected the resting potential (−83 to −85 mV). Both drugs depressed the action potential plateau amplitude and tension when the stimulation rate was 120 min−1; ED50's for the negative inotropic effect were 7×10−8 M for D600 and 3×10−5 M for AQA 39. At 20 min−1, D600 was only slightly less depressant (ED50 for negative inotropy 4×10−7 M) but 5×10−5 M AQA 39 had no effect, neither on tension nor on the action potential plateau amplitude. 2. In voltage clamp experiments (holding potential −50 mV), 2×10−5 M AQA 39 and 2×10−6 M D600 were equipotent. With depolarizing pulses (250 ms steps to −10 mV at 20 min−1), they depressed slow inward calcium current (Isi) by 70% and peak tension by 80% in 30 min. 3. At a membrane potential of −50 mV, Isi in muscle exposed to either drug for 30 min without stimulation was not depressed during the first post-rest clamp but the block of calcium channels was rapid upon resumption of stimulation (i.e. a “use-dependent” block). 4. Following steady-state block, Isi could be restored to control amplitude with hyperpolarizing pre-pulses, whereas depolarizing pre-pulses enhanced the block of Isi. 5. It seems that both drugs block calcium channels only after the channels have been activated (opened) and that the degree of activation and the degree of block are linked. The drugs dissociate from a blocking site at a rate dependent upon the diastolic membrane potential; the more negative the membrane potential inbetween subsequent depolarizations, the quicker the dissociation. While the dissociation of AQA 39 at −50 mV is only about twice as fast as that of D600, it is nearly 100 times faster at −80 mV and more than 150 times faster at potentials more negative than −90 mV. These differences might be related to the different lipophilic properties of the two drugs.

AQA 39*, a new bradycardic agent which blocks myocardial calcium (Ca) channels in a frequency- and voltage-dependent manner.

AQA 39 is a new drug which resembles verapamil and D600 in chemical structure. However, under normal conditions (i.e., extracellular potassium of 5.4 mmol/l, 0.33 Hz stimulation rate, fully polarized healthy preparations), the effects of this agent on ventricular muscle bear remarkable differences to those of the classic Ca channel blocking agents under the same conditions, since AQA 39 does not affect either calcium-dependent electrical or mechanical activity. Therefore, the effects of this drug on rabbit ventricular muscle have been investigated in detail: 1. The action potential duration lengthened with concentration (10−6–10−4 mol/l) at stimulation rates of 0.33, 1, and 2 Hz. There was no change in the resting potential. 2. At 0.33 Hz, concentrations up to 10−4 mol/l had no effect on the action potential plateau amplitude or peak force of contraction. At 1 and 2 Hz depression was apparent at 5×10−6 mol/l and above. 3. The plateau amplitude of the slow action potential (20 mmol/l K+) was markedly depressed by AQA 39 (2×10−5 mol/l) at 0.33 Hz; peak force of contraction declined to about 30% control. 4. When voltage clamp steps were imposed from a holding potential of −50 mV, there was a large reduction in slow inward calcium current (Isi) and peak force of contraction at voltages between −40 and +40mV. The outward current was also depressed. 5. Changes in the reversal potential of Isi or in the voltage dependence of the activation kinetics were ruled out as explanation for the depression of Isi. 6. The block of Ca channels was dependent on the level of the diastolic potential. When Isi was depressed to about 30% control with 0.33 Hz stimulation from −50 mV, a 10 s rest at −50 mV was without effect but a 5 s rest at −85 mV restored Isi to the pre-drug amplitude. 7. It is concluded that AQA 39 at a critical concentration ought to allow normal rhythm but supress tachyarrhythmic episodes or early extrasystolic activity in the S-A node, the A-V node or depolarized myocardial foci without affecting electrical activity and contractility in other regions.

Electrophysiological and inotropic effects of natural amino-deoxyglyco-cardenolides

The electrophysiological, inotropic and chronotropic effects of the two first natural amino-deoxyglyco-cardenolides, mitiphylline and holarosine B, were studied. Mitiphylline has qualitatively the same properties as those of common cardiac glycosides, but is more potent; holarosine B has a similar action on cardiac electrophysiological parameters of the guinea pig heart but is less potent. The difference in activity between these two compounds could be attributed to the configuration in 5- and 17-positions of the genins. Holarisone B has the particularity of increasing the action potential plateau amplitude and its duration in rat heart; this could be part of the role of the aminosugar moiety.