Current In Heart Cells Research Articles

Dan Roden: learn, apply, evolve.

As a scientist, Dan Roden grew up as clinical electrophysiology did. Widely recognized for his research into mechanisms behind abnormal heart rhythms and drug responses, Roden was on the front lines in the 1980s and 1990s as new antiarrhythmic drugs were tested, the ability to use catheters to study drug actions evolved, and the specialty of clinical electrophysiology arose. From his first publication, appearing in the New England Journal of Medicine in 1980,1 Roden set a tone for a career focused on variable action of antiarrhythmic therapies and genetic determinants of that variation. Much of his current work focuses on individual cardiac ion channel mutations and their role in variable responses to drug therapy, and applications of genomics to healthcare. Among his achievements, Roden is credited with developing the idea of reduced repolarization reserve leading to acquired long QT intervals and arrhythmias2–4 and was one of the first to suggest that early afterdepolarizations cause long QT-related arrhythmias.5,6 As leader of Vanderbilt University’s pharmacogenomics project PREDICT (Pharmacogenomic Resource for Enhanced Decisions in Care & Treatment)—which pre-emptively enhances individuals’ electronic medical records with personalized information about genetic variants that could guide drug therapy—Roden and colleagues7 have demonstrated multiple advantages of preemptive genotyping. Meanwhile Vanderbilt’s massive BioVU DNA bank, which includes ≈175 000 human DNA samples and which Roden pioneered and directs, is similarly showing the power of incorporating genomic data into electronic medical records, for research purposes.7–11 Roden, who initially flirted with a future in journalism, described to Circulation Research some of his interests beginning as a child of immigrants in Montreal and leading to not just 1 career as an investigator, but a series of interlinked careers—in clinical pharmacology, cellular electrophysiology, molecular genetics, and population science. The paradigm for his progress, …

Critical Requirements for the Initiation of a Cardiac Arrhythmia in Heart: Cell Number

Many features of an intact functioning heart provide a margin of safety with respect to the generation of cardiac arrhythmias. We have been investigating Ca2+ dependent arrhythmias and the requirements for such pathological activity including features that focus on the margin of safety. One of the primary issues is the number of cells involved in the initiation process. If too few cells are needed to initiate the arrhythmia, arrhythmias will be rampant. If too many cells are needed, then it is difficult to explain how known arrhythmias occur. Thus the balance is one of the critical parameter to be estimated to validate the model and also to develop a useful set of computational tools to examine lethal cardiac arrhythmias. It has been suggested in published work that, a large number (∼800,000) of ventricular myocytes be depolarized to overcome the electrotonic load of the heart. In our recent work we have re-examined this question in light of a revised stochastic model of electrical and Ca2+ activity. We have found that as few as 12 myocytes may be needed to initiate an ectopic cardiac arrhythmia. This reduction of over 4 orders of magnitude reflects important improvements in the modeling. These changes include the stochasticity of the Ca2+ modeling, improved assumptions regarding the ionic currents in heart cells and updated geometries. We will present and discuss these critical changes and the justification for them. Our new findings regarding the initiation of spontaneous arrhythmias are in line with functional data from the heart. This new finding suggests a mechanism by which the generation of arrhythmias is possible while still maintaining a high safety margin.

The actions of mdivi-1, an inhibitor of mitochondrial fission, on rapidly activating delayed-rectifier K+ current and membrane potential in HL-1 murine atrial cardiomyocytes

Mdivi-1 is an inhibitor of dynamin related protein 1- (drp1)-mediated mitochondrial fission. However, the mechanisms through which this compound interacts directly with ion currents in heart cells remain unknown. In this study, its effects on ion currents and membrane potential in murine HL-1 cardiomyocytes were investigated. In whole-cell recordings, the addition of mdivi-1 decreased the amplitude of tail current (Itail) for the rapidly activating delayed-rectifier K+ current (IKr) in a concentration-dependent manner with an IC50 value at 11.6μM, a value that resembles the inhibition requirement for mitochondrial division. It shifted the activation curve of Itail to depolarized voltages with no change in the gating charge. However, mdivi-1 did not alter the rate of recovery from current inactivation. In cell-attached configuration, mdivi-1 inside the pipette suppressed the activity of acetylcholine-activated K+ channels without modifying the single-channel conductance. Mdivi-1 (30μM) slightly depressed the peak amplitude of Na+ current with no change in the overall current–voltage relationship. Under current-clamp recordings, addition of mdivi-1 resulted in prolongation for the duration of action potentials (APs) and to increase the firing of spontaneous APs in HL-1 cells. Similarly, in pituitary GH3 cells, mdivi-1 was effective in directly suppressing the amplitude of ether-à-go-go-related gene-mediated K+ current. Therefore, the lengthening of AP duration and increased firing of APs caused by mdivi-1 can be primarily explained by its inhibition of these K+ channels enriched in heart cells. The observed effects of mdivi-1 on ion currents were direct and not associated with its inhibition of mitochondrial division.

Founder of cardiac cellular electrophysiology: Honouring Silvio Weidmann, 7 April 1921– 11 July 2005

Founder of cardiac cellular electrophysiology: Honouring Silvio Weidmann, 7 April 1921– 11 July 2005

Angiotensin II-induced increase of T-type Ca 2+ current and decrease of L-type Ca 2+ current in heart cells

The effect of angiotensin II (Ang II) on the T- and L-type calcium currents ( I Ca) in single ventricular heart cells of 18-week-old fetal human and 10-day-old chick embryos was studied using the whole-cell voltage clamp technique. Our results showed that in both, human and chick cardiomyocytes, Ang II (10 −7 M) increased the T-type calcium current and decreased the L-type I Ca. The effect of Ang II on both types of currents was blocked by the AT 1 peptidic antagonist, [Sar 1, Ala 8] Ang II (2 × 10 −7 M). Protein kinase C activator, phorbol 12,13-dibutyrate, mimicked the effect of Ang II on the T- and L-type calcium currents. These results demonstrate that in fetal human and chick embryo cardiomyocytes Ang II affects the T- and L-type Ca 2+ currents differently, and this effect seems to be mediated by the PKC pathway.

Bradykinin induced a positive chronotropic effect via stimulation of T- and L-type calcium currents in heart cells.

Using Fluo-3 calcium dye confocal microscopy and spontaneously contracting embryonic chick heart cells, bradykinin (10(-10) M) was found to induce positive chronotropic effects by increasing the frequency of the transient increase of cytosolic and nuclear free Ca2+. Pretreatment of the cells with either B1 or B2 receptor antagonists (R126 and R817, respectively) completely prevented bradykinin (BK) induced positive chronotropic effects on spontaneously contracting single heart cells. Using the whole-cell voltage clamp technique and ionic substitution to separate the different ionic current species, our results showed that BK (10(-6) M) had no effect on fast Na+ inward current and delayed outward potassium current. However, both L- and T-type Ca2+ currents were found to be increased by BK in a dose-dependent manner (10(-10)-10(-7) M). The effects of BK on T- and L-type Ca2+ currents were partially blocked by the B1 receptor antagonist [Leu8]des-Arg9-BK (R592) (10(-7) M) and completely reversed by the B2 receptor antagonist D-Arg[Hyp3,D-Phe7,Leu8]BK (R-588) (10(-7) M) or pretreatment with pertussis toxin (PTX). These results demonstrate that BK induced a positive chronotropic effect via stimulation of T- and L-type Ca2+ currents in heart cells mainly via stimulation of B2 receptor coupled to PTX-sensitive G-proteins. The increase of both types of Ca2+ current by BK in heart cells may explain the positive inotropic and chronotropic effects of this hormone.

Early fetal like slow Na+ current in heart cells of cardiomyopathic hamster.

Using the whole-cell voltage-clamp technique, early embryonic tetrodotoxin (TTX) and Mn(2+)-insensitive slow Na+ current was detected in 10-22 week old fetal human heart cells as well as in 1 day old and young cardiomyopathic hamster myocytes. This slow Na+ current in both heart cell preparations has the same kinetics and pharmacology. This type of slow Na+ current was absent in heart cells of newborn and young normal hamsters and became less present in myocytes of 19 and 22 week old human heart myocytes. Our results demonstrate that the slow Na+ channel does exist in early fetal human life and this type of channel continues to be functional after birth in myocytes of the hereditary cardiomyopathic hamster.

Phosphorylation-independent modulation of L-type calcium channels by magnesium-nucleotide complexes.

Free magnesium ions and magnesium-nucleotide complexes can exert opposite effects on many fundamental cellular processes. Although increases in the intracellular concentration of magnesium ions inhibit the L-type calcium current in heart cells, magnesium-adenosine triphosphate complexes (MgATP) would be expected to increase the current by promoting channel phosphorylation. Rapid increases in the intracellular concentration of MgATP induced by flash photolysis of caged magnesium or caged ATP resulted in enhanced calcium current. The increase in calcium current was not prevented by blocking phosphorylation, revealing a previously unrecognized direct regulatory action of the magnesium-nucleotide complex.

A long lasting Ca2+ -activated outward current in guinea-pig atrial myocytes

Among other characteristics, the steady-state current-voltage relationship of patch-clamped single atrial myocytes from guinea-pig hearts is defined by an outward current hump in the potential region -15 to +40 mV. This hump was reversibly suppressed by Co2+ (3 mM) or nitrendipine (5 microM) and enhanced by Bay K 8644 (5 microM). The maintained outward current component suppressed by Co2+ extended between -15.2 +/- 1.9 mV and +39.5 +/- 1.7 mV (mean +/- SEM of 14 cells) and has an amplitude of 95.7 +/- 9.4 pA at +10 mV. In isochronal I-V curves, the hump was already visible at 400 ms with essentially the same amplitude as at 1500 ms. The Co2+-sensitive outward current underlying the hump was poorly time-dependent during 1.5 s voltage pulses but slowly relaxed upon repolarization. Tail currents reversed near the K+ equilibrium potential under our experimental conditions. The current hump of the steady-state I-V curve was also abolished by caffeine (10 mM) or ryanodine (3 microM), both drugs that interfere with sarcoplasmic reticulum function. Apamin (1 microM) or quinine (100 microM) but not TEA (5-50 mM) markedly reduced its amplitude. However, at similar concentrations as required to inhibit the hump, both apamin and quinine appeared to be poorly specific for Ca2+-activated K+ currents in heart cells since they also inhibited the L-Type Ca2+ current. It is concluded that a long lasting Ca2+-activated outward current, probably mainly carried by K+ ions but not sensitive to TEA, exists in atrial myocytes which is responsible for the current hump of the background I-V curve.

Intracellular bretylium blocks Na + and K + currents in heart cells

The effects of intracellular and extracellular application of bretylium tosylate on outward potassium current and fast inward Na + current have been studied in single ventricular cells of chick embryo (18-day-old) using the whole-cell voltage clamp technique. Extracellular superfusion with bretylium (3 × 10 −4 M) decreased the K + current (I K) whereas the fast I Na remained unaffected. Introducing bretylium intracellularly also decreased I K, but rapidly inhibited the fast I Na. The addition of 3 × 10 −4 M bretylium to the superfusion medium in the presence of intracellular bretylium further decreased I K and caused the fast inward sodium current (I Na) to recover. The decrease of I K by bretylium may account for the antifibrillatory properties of this drug in heart.

Separation of Na-Ca exchange and transient inward currents in heart cells.

Enzymatically dispersed single cells from rabbit ventricle were voltage clamped using the suction pipette method to investigate whether in isolated cardiac cells a recently described slow inward current (IEX) due to the electrogenic Na+-dependent Ca2+ extrusion also underlies a transient inward current (ITI), which can trigger certain cardiac arrhythmias. The cells were held at -40 mV to inactivate the fast sodium current. After depolarizing pulses (to 0 or +10 mV for 50 to 200 ms), slow inward "tail" currents were consistently recorded. Previous results indicate that this tail current IEX is generated by the Na+-Ca2+ exchanger. After loading the cells with Ca2+ by blocking the Na+-K+ pump [either with strophanthidin (10(-5) M) treatment or by reducing external K+ to 1 mM or less], ITIS appeared. These were usually spontaneous but occasionally were time locked to the clamp pulses. It was possible to separate IEX and ITI by a variety of methods. These include the following. 1) Different stimulation protocols; repolarizing to more negative potentials augmented IEX and decreased or eliminated ITI. Increasing the rate of stimulation diminished IEX and increased ITI. 2) Pharmacological methods; adding BaCl2 (0.5-2.0 mM) or caffeine (5-10 mM) decreased IEX but abolished ITI. The findings suggest that different mechanisms regulate these two currents.

Voltage-dependent modulation of Ca channel current in heart cells by Bay K8644.

We have investigated the voltage-dependent effects of the dihydropyridine Bay K8644 on Ca channel currents in calf Purkinje fibers and enzymatically dispersed rat ventricular myocytes. Bay K8644 increases the apparent rate of inactivation of these currents, measured during depolarizing voltage pulses, and shifts both channel activation and inactivation in the hyperpolarizing direction. Consequently, currents measured after hyperpolarizing conditioning pulses are larger in the presence of drug compared with control conditions, but are smaller than control if they are measured after positive conditioning pulses. Most of our experimental observations on macroscopic currents can be explained by a single drug-induced change in one rate constant of a simple kinetic model. The rate constant change is consistent with results obtained by others with single channel recordings.