The transmembrane potential is recorded from small isopotential clusters of 2-4 embryonic chick ventricular cells spontaneously generating action potentials. We analyze the cycle-to-cycle fluctuations in the time between successive action potentials (the interbeat interval or IBI). We also convert an existing model of electrical activity in the cluster, which is formulated as a Hodgkin-Huxley-like deterministic system of nonlinear ordinary differential equations describing five individual ionic currents, into a stochastic model consisting of a population of ∼20 000 independently and randomly gating ionic channels, with the randomness being set by a real physical stochastic process (radio static). This stochastic model, implemented using the Clay-DeFelice algorithm, reproduces the fluctuations seen experimentally: e.g., the coefficient of variation (standard deviation/mean) of IBI is 4.3% in the model vs. the 3.9% average value of the 17 clusters studied. The model also replicates all but one of several other quantitative measures of the experimental results, including the power spectrum and correlation integral of the voltage, as well as the histogram, Poincaré plot, serial correlation coefficients, power spectrum, detrended fluctuation analysis, approximate entropy, and sample entropy of IBI. The channel noise from one particular ionic current (IKs), which has channel kinetics that are relatively slow compared to that of the other currents, makes the major contribution to the fluctuations in IBI. Reproduction of the experimental coefficient of variation of IBI by adding a Gaussian white noise-current into the deterministic model necessitates using an unrealistically high noise-current amplitude. Indeed, a major implication of the modelling results is that, given the wide range of time-scales over which the various species of channels open and close, only a cell-specific stochastic model that is formulated taking into consideration the widely different ranges in the frequency content of the channel-noise produced by the opening and closing of several different types of channels will be able to reproduce precisely the various effects due to membrane noise seen in a particular electrophysiological preparation.

Purpose Teduglutide is a recombinant analogue of human glucagon-like peptide-2 that has recently been approved for the treatment of short bowel syndrome in adults. This study was designed to study the influence of renal function and age on teduglutide pharmacokinetics. Methods This was an open-label study with six parallel groups (6 subjects each). Three groups with renal impairment (moderate, severe and end-stage renal disease) were compared to healthy subjects with normal renal function, which were matched to the renal-impaired subjects with respect to demographics. At least two elderly subjects (≥65 years) were enrolled per group. A single dose of 10 mg teduglutide was subcutaneously administered to each subject. Teduglutide plasma concentrations were measured using a validated liquid chromatography method with tandem mass spectrometric detection, and the primary pharmacokinetic variables (AUC inf and C max ) were calculated. Results Area under the concentration versus time curve extrapolated to infinity (AUC inf ) and maximum plasma concentration (C max ) of teduglutide in subjects with end-stage renal disease were approximately 2.59- and 2.08-fold higher, respectively, than those of healthy subjects. The AUC inf and C max were also slightly higher in subjects with moderate and severe renal impairment. Comparison of healthy subjects aged <65 years with healthy elderly subjects revealed very similar pharmacokinetics in both subgroups. Conclusions In our study population, the primary pharmacokinetic parameters of teduglutide increased with increased severity of renal impairment. These results suggest that the daily dose of teduglutide should be reduced by 50 % in patients with moderate and severe renal impairment and end-stage disease. We found no effect of age on the pharmacokinetics of teduglutide in healthy subjects. The treatment was well tolerated, and there were no safety concerns.

Objective: It was our aim to investigate effects of human LDL, copper-, or AAPH-oxidized over different periods of time to different degrees (ox-LDL), on viability and electrophysiological parameters of isolated ventricular myocytes of guinea pigs.Methods: Guinea pig ventricular myocytes were incubated with ox-LDL or native LDL (at 0.5 mg/ml) for 12 h, and afterwards myocyte damage, action potentials, and transmembrane ion currents were studied (at 37 degrees C).Results: Ox-LDL was found to induce severe myocyte damage, whereas native LDL had no effect. Myocyte damage was dependent on the content of total lipid hydroperoxides in both copper-oxidized and AAPH-oxidized LDL. Incubation with ox-LDL led to intense contractile and electrophysiological effects including prolongation of action potential duration, depolarization of resting membrane potential, spontaneous activity, generation of afterdepolarizations, and modification of transmembrane ion currents (e.g. inward rectifier, calcium, and background currents).Conclusions: Ox-LDL induced cell damage and irregular electrical activity in ventricular myocytes. These effects were dependent on the lipid hydroperoxide content of ox-LDL and were similar to oxidative stress (OS) induced by various OS-generating systems. The observed effects may play a role for functional cardiac abnormalities in patients with increased ox-LDL levels. (c) 2004 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.

We recorded transmembrane potential in whole cell recording mode from small clusters (2-4 cells) of spontaneously beating 7-day embryonic chick ventricular cells after 1-3 days in culture and investigated effects of the blockers D-600, diltiazem, almokalant, and Ba2+. Electrical activity in small clusters is very different from that in reaggregates of several hundred embryonic chick ventricular cells, e.g., TTX-sensitive fast upstrokes in reaggregates vs. TTX-insensitive slow upstrokes in small clusters (maximum upstroke velocity approximately 100 V/s vs. approximately 10 V/s). On the basis of our voltage- and current-clamp results and data from the literature, we formulated a Hodgkin-Huxley-type ionic model for the electrical activity in these small clusters. The model contains a Ca2+ current (ICa), three K+ currents (IKs, IKr, and IK1), a background current, and a seal-leak current. ICa generates the slow upstroke, whereas IKs, IKr, and IK1 contribute to repolarization. All the currents contribute to spontaneous diastolic depolarization, e.g., removal of the seal-leak current increases the interbeat interval from 392 to 535 ms. The model replicates the spontaneous activity in the clusters as well as the experimental results of application of blockers. Bifurcation analysis and simulations with the model predict that annihilation and single-pulse triggering should occur with partial block of ICa. Embryonic chick ventricular cells have been used as an experimental model to investigate various aspects of spontaneous beating of cardiac cells, e.g., mutual synchronization, regularity of beating, and spontaneous initiation and termination of reentrant rhythms; our model allows investigation of these topics through numerical simulation.

Objective: The left human atrium plays an important role in initiation of atrial fibrillation (AF) and the hyperpolarization activated cation current (I-f) is a candidate for contributing to abnormal automaticity. However, electrophysiological data concerning I-f are not available in this cardiac region and we therefore investigated I-f in human left atrial tissue.Methods: Human atrial myocytes were isolated from the left atrial appendage (LAA) and the left atrial wall (LAW) obtained from patients undergoing open heart surgery. I-f was measured with the whole-cell patch-clamp technique.Results: I-f densities between -70 and -110 mV were found to be significantly higher in LAA than in LAW cells. Furthermore, in the group of LAA cells the half maximal activation potential (V-1/2) was found to be less negative (V-1/2 of -84.3 +/- 1.9 mV, n = 14/9) compared to LAW Cells (V-1/2 of -97.8 +/- 2.1 mV, n = 28/9). Beta-adrenergic receptor Stimulation with isoproterenol (1 muM) caused an acceleration of current activation and a V-1/2 shift to more positive potentials in cells of both regions (LAA: 8.8 +/- 2.3 mV, n = 6/4 and LAW: 8.9 +/- 2.6 mV, n = 6/4). Simulations using a mathematical model of the human atrial myocyte demonstrated that I-f was able to induce spontaneous activity in the model at a regular rhythm due to the interplay of I-f, Na+/Ca2+ exchange current and Ca2+ release of the sarcoplasmic reticulum (SR).Conclusions: Our study revealed the presence of I-f in left atrial myocytes and showed that I-f parameters depend on atrial region. I-f current densities were sufficient to convert the mathematical model of a quiescent human atrial cell into a "pacemaker cell". These data support the hypothesis of I-f as a contributor to abnormal automaticity in human atrial tissue. (C) 2004 European Society of Cardiology. Published by Elsevier B.V. All rights reserved.

The aim of this investigation was to study L-type and T-type Ca(2+) current (I(CaL) and I(CaT)) in short-term cultured adult guinea pig ventricular myocytes. The isolated myocytes were suspended in serum-supplemented medium up to 5 days. Using whole-cell patch clamp techniques ICaL and ICaT were studied by applying voltage protocols from different holding potentials (-40 and -90 mV). After 5 days in culture the myocytes still showed their typical rod shaped morphology but a decline in cell membrane capacitance (26 %). The peak density of ICaT was reduced significantly between day 0 (-1.6+/-0.37 pA/pF, n=9) and day 5 (-0.4+/-0.13 pA/pF, n=11), whereas peak ICaL density revealed no significant differences during culturing. The I(CaT)/I(CaL) ratio dropped from 0.13 at day 0 to 0.05 at day 5. Compared with day 0 I(CaL) the steady state inactivation curve of day 1, day 3 and day 5 myocytes was slightly shifted to more negative potentials. Our data indicate that guinea pig ventricular L-type and T-type Ca(2+) channels are differently regulated in culture.

The aim of this study was to investigate the effect of NADH-supplementation on the metabolic condition of isolated guinea pig ventricular cardiomyocytes using the pinacidil-primed IK(ATp) as an indicator of subsarcolemmal ATP concentration. Membrane currents were studied using the patch-clamp technique in the whole-cell recording mode at 36-37C. Under physiological conditions (4.3 mM ATP in the pipette solution, ATPj) IK(ATP) did not contribute to basal electrical activity The K(ATP) channel opener pinacidil activated IK(ATP) dependent on [ATP]i showing a significantly more pronounced activation at lower (1 mM) [ATP]j. Incubation of cardiomyocytes with 300 μg/mI NADH (4-6 h) resulted in a significantly reduced IK(ATp) activation by pinacidil compared to control cells. Equimolar amounts of the related compounds nicotinamide and NAD were not able to achieve a similar effect like NADH. These data show that incubation of guinea pig ventricular cardiomyocytes with NADH results in a decreased activation Of IK(ATp) by pinacidil compared to control myocytes indicating a higher subsarcolemmal ATP concentration due to NADH -supplementation. Measurement of adenine nucleotides by HPLC revealed a significant increase in intracellular ATP (NADH supplementation: 45.59 ± 1,88 nmol/mg protein versus control: 35.35 ± 2.57 nmol/mg protein, P < 0.000005).

. I f was shown to be present in adult human atrial and ventricular myocytes but data obtained from infant myocytes are lacking. We have studied I f in isolated ventricular myocytes from children undergoing surgical correction of tetralogy of Fallot (TOF; n = 5; mean age: 15.3 months). All recordings were made with the patch clamp technique in the whole cell mode at a temperature of 36–37 °C. A modified Tyrode solution containing 25 mM KCl was used to amplify I f . Considering I f to be present when its current density at –120 mV was greater than 0.5 pA/pF, I f could be found in 28 out of 32 myocytes (88%). The mean current density was –2.01 ± 0.3 pA/pF (mean ± S.E.M.). First current activation occurred at –70 mV and I f could be reversibly inhibited by superfusing the myocytes with CsCl (2 mM). Half maximal activation (V 1/2 ) of I f was at –80.3 ± 1.0 mV (n = 28). Beta-adrenergic receptor stimulation with isoproterenol (1 µM) caused an acceleration of current activation and a shift of V 1/2 by 7.88 ± 1.8 mV (n = 10) to less negative potentials. This study provides first evidence that the hyperpolarization-activated pacemaker current I f is present in infant human ventricular myocytes. Our results suggest that I f in ventricle of infants suffering from TOF has similar properties as I f in adult ventricle.

The aim of this study was to investigate the effect of nicotinamide‐adenine dinucleotide (NADH) supplementation on the metabolic condition of isolated guinea‐pig ventricular cardiomyocytes. The pinacidil‐primed ATP‐dependent potassium current I K(ATP) was used as an indicator of subsarcolemmal ATP concentration and intracellular adenine nucleotide contents were measured. Membrane currents were studied using the patch‐clamp technique in the whole‐cell recording mode at 36–37°C. Adenine nucleotides were determined by HPLC. Under physiological conditions (4.3 m M ATP in the pipette solution, ATP i ) I K(ATP) did not contribute to basal electrical activity. The ATP‐dependent potassium (K (ATP) ) channel opener pinacidil activated I K(ATP) dependent on [ATP] i showing a significantly more pronounced activation at lower (1 m M ) [ATP] i . Supplementation of cardiomyocytes with 300 μ g ml −1 NADH (4–6 h) resulted in a significantly reduced I K(ATP) activation by pinacidil compared to control cells. The current density was 13.8±3.78 ( n =6) versus 28.9±3.38 pA pF −1 ( n =19; P <0.05). Equimolar amounts of the related compounds nicotinamide and NAD + did not achieve a similar effect like NADH. Measurement of adenine nucleotides by HPLC revealed a significant increase in intracellular ATP (NADH supplementation: 45.6±1.88 nmol mg −1 protein versus control: 35.4±2.57 nmol mg −1 protein, P <0.000005). These data show that supplementation of guinea‐pig ventricular cardiomyocytes with NADH results in a decreased activation of I K(ATP) by pinacidil compared to control myocytes, indicating a higher subsarcolemmal ATP concentration. Analysis of intracellular adenine nucleotides by HPLC confirmed the significant increase in ATP. British Journal of Pharmacology (2003) 139 , 749–754. doi: 10.1038/sj.bjp.0705300

The aim of this study was to investigate the effects of the potassium channel openers (PCOs) cromakalim and pinacidil on the ATP-dependent potassium current I K(ATP) in human atrial myocytes.

It was the aim of our study to investigate the effects of the sulphonylurea glibenclamide on voltage dependent potassium currents in human atrial myocytes. The drug blocked a fraction of the quasi steady state current (ramp response) which was activated positive to −20 mV, was sensitive to 4‐aminopyridine (500 μM) and was different from the ATP dependent potassium current IK(ATP). Glibenclamide dose dependently inhibited both, the peak as well as the late current elicited by step depolarization positive to −20 mV. The IC50 for reduction in charge area of total outward current was 76 μM. The double‐exponential inactivation time‐course of the total outward current was accelerated in the presence of glibenclamide with a τfast of 12.7±1.5 ms and a τslow of 213±25 ms in control and 5.8±1.9 ms (P<0.001) and 101±20 ms (P<0.05) under glibenclamide (100 μM). Our data suggest, that both repolarizing currents in human atrial myocytes, the transient outward current (Ito1) and the ultrarapid delayed rectifier current (IKur) were inhibited by glibenclamide. In human ventricular myocytes glibenclamide inhibited Ito1 without affecting the late current. Our data suggest that glibenclamide inhibits human voltage dependent cardiac potassium currents at concentrations above 10 μM. British Journal of Pharmacology (1999) 128, 1175–1180; doi:10.1038/sj.bjp.0702904

Objective: It was the aim of our study to describe repolarizing currents in ventricular myocytes isolated from children with tetralogy of Fallot. This is the first report on outward currents in ventricular myocytes from children. Methods: Ventricular myocytes were isolated from tissue samples of the outflow tract of the right ventricle which were obtained during corrective surgery of tetralogy of Fallot. Action potentials and whole-cell currents were recorded with the patch clamp technique at a temperature of 36-37 degrees C. Results: The mean resting potential was -71.7+/-1.92 mV, action potential amplitude was 110+/-2.96 mV and action potential duration at 90% repolarization was 794+/-99.5 ms (n = 12). In four out of 12 myocytes early afterdepolarizations (EADs) were observed. Upon hyperpolarization Ba2+-sensitive inward currents similar to the inward rectifier current (I-K1) could be observed. The current density at -120 mV was -22.8+/-2.47 pA/pF (n = 14). A transient outward current (I-to 1) could be recorded in all myocytes studied, the current density varied from 0.3 to 8.6 pA/pF with a mean of 3.77+/-0.47 pA/pF at +40 mV (n=38). Recovery of I-to 1 from inactivation was fast (70% recovery within 100 ms), rate-dependent reduction amounted to 38.2% at 4 Hz. A delayed rectifier current was seen in only two out of 38 myocytes (rapid component I-Kr). Conclusions: The electrophysiological characteristics of right ventricular myocytes isolated from children with tetralogy of Fallot resemble in most cases subendocardial myocytes from adults. The most prominent difference is a fast recovery from inactivation as well as a small rate dependent reduction of I-to 1. The observed EADs may have clinical implications. (C) 1999 Elsevier Science B.V. All rights reserved.