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A new open-hardware bioreactor capable of applying electrical field stimulation in conjunction with static or cyclic stretch is presented. Stretch is applied to cells by a specially designed elastomeric membrane with a central seeding region. The main interest of our approach is the fine control of the characteristics of stimulations in regard to timing and amplitude in a simple design based on affordable, easy to find components and 3D printable parts. Our approach opens the way to more complex protocols for electrical and/or mechanical stimulations, which are known important regulators of cardiac phenotypes.

Background— The development of effective and safe antiarrhythmic drugs for atrial fibrillation (AF) rhythm control is an unmet clinical need. Multichannel blockers are believed to have advantages over single-channel blockers for AF, but their development has been completely empirical to date. We tested the hypothesis that adding K + -channel blockade improves the atrium-selective electrophysiological profile and anti-AF effects of optimized Na + -channel blockers. Methods and Results— Realistic cardiomyocyte-, tissue-, and state-dependent Na + -channel block mathematical models, optical mapping, and action potential recording were used to study the effect of Na + -current ( I Na ) blockade with or without concomitant inhibition of the rapid or ultrarapid delayed-rectifier K + currents ( I Kr and I Kur , respectively). In the mathematical model, maximal AF selectivity was obtained with an inactivated-state Na + -channel blocker. Combining optimized Na + -channel blocker with I Kr block increased rate-dependent and atrium-selective peak I Na reduction, increased AF selectivity, and more effectively terminated AF compared with optimized Na + -channel blocker alone. Combining optimized Na + -channel blocker with I Kur block had similar effects but without I Kr block–induced ventricular action potential prolongation. Consistent with the mathematical model, in coronary-perfused canine hearts, the addition of dofetilide (selective I Kr blocker) to pilsicainide (selective I Na blocker) produced enhanced atrium-selective effects on maximal phase 0 upstroke and conduction velocity. Furthermore, pilsicainide plus dofetilide had higher AF termination efficacy than pilsicainide alone. Pilsicainide alone had no statistically significant effect on AF inducibility, whereas pilsicainide plus dofetilide rendered AF noninducible. Conclusions— K + -channel block potentiates the AF-selective anti-AF effects obtainable with optimized Na + -channel blockade. Combining optimized Na + -channel block with blockade of atrial K + currents is a potentially valuable AF-selective antiarrhythmic drug strategy.

The species-specific determinants of repolarization are poorly understood. This study compared the contribution of various currents to cardiac repolarization in canine and human ventricle. Conventional microelectrode, whole-cell patch-clamp, molecular biological and mathematical modelling techniques were used. Selective IKr block (50-100 nmol l(-1) dofetilide) lengthened AP duration at 90% of repolarization (APD90)3-fold more in human than dog, suggesting smaller repolarization reserve in humans. Selective IK1 block (10 μmol l(-1) BaCl2) and IKs block (1 μmol l(-1) HMR-1556) increased APD90 more in canine than human right ventricular papillary muscle. Ion current measurements in isolated cardiomyocytes showed that IK1 and IKs densities were 3- and 4.5-fold larger in dogs than humans, respectively. IKr density and kinetics were similar in human versus dog. ICa and Ito were respectively ~30% larger and ~29% smaller in human, and Na(+)-Ca(2+) exchange current was comparable. Cardiac mRNA levels for the main IK1 ion channel subunit Kir2.1 and the IKs accessory subunit minK were significantly lower, but mRNA expression of ERG and KvLQT1 (IKr and IKs α-subunits) were not significantly different, in human versus dog. Immunostaining suggested lower Kir2.1 and minK, and higher KvLQT1 protein expression in human versus canine cardiomyocytes. IK1 and IKs inhibition increased the APD-prolonging effect of IKr block more in dog (by 56% and 49%, respectively) than human (34 and 16%), indicating that both currents contribute to increased repolarization reserve in the dog. A mathematical model incorporating observed human-canine ion current differences confirmed the role of IK1 and IKs in repolarization reserve differences. Thus, humans show greater repolarization-delaying effects of IKr block than dogs, because of lower repolarization reserve contributions from IK1 and IKs, emphasizing species-specific determinants of repolarization and the limitations of animal models for human disease.

Mechanical stretch of cardiomyocytes modulates the action potential (AP) via stretch-activated channels. The electrical activity is also modulated by the parasympathetic nervous system via the acetylcholine (ACh)-dependent potassium current. The ACh effect is however heterogeneous throughout the atria thus facilitating arrhythmic events. Simultaneous activation of both systems could occur and may facilitate atrial arrhythmias. Simulations of a canine atrial ionic model in an isolated cell and linear tissue strand were computed with varying stretch and ACh levels. Pacing at 1Hz, AP duration (APD) is increased (see APs in panel A) by ∼47 ms with 20% stretch compared to control (CTL). However, stretch did not increase APD in presence of 25 nmol/L of ACh (ACh vs. ACh+stretch in panel A). Restitution curves calculated with the S1-S2 protocol (S1=1 Hz) are plotted in panel B. Stretch (20%) results in an upshift of ∼47 ms of the restitution curve compared to CTL. ACh almost eliminate atrial restitution compared to CTL with no important changes with stretch (ACh vs. ACh+stretch curves). Preliminary results obtained in a cable with heterogeneous ACh distribution showed an increased interval for block of electrical propagation with tissue stretch.View Large Image | View Hi-Res Image | Download PowerPoint Slide

Problematic. The biological pacemaker is a new therapeutic approach that could lead to optimized treatment of bradycardia. A possibility is the development of a thin sheet of cardiomyocytes, cultured to obtain a target activation rate. Fundamental research, often conducted with neonatal rat ventricular myocytes (NRVMs), partially revealed two basic coupled mechanisms of automaticity termed Voltage Clock (synergy of membrane currents) and Calcium Clock (internal oscillations of calcium concentration). To date, no ionic model is able to reproduce in silico the autonomous activity found in cultured NRVMs. The present project aims to fill this gap.Methods. A non-automatic NRVM ionic model (Korhonen-Tavi, 2009) is modified according to documented Voltage and Calcium Clocks mathematical formulations. The myocytes are cultured for 48 hours at low density, allowing cells to remain single on dishes. Autonomous action potentials (APs) are measured with patch clamp method, and calcium transients (CTs) with Fluo-4 AM fluorescence imaging. Experimental variables (spontaneous rate, AP amplitude and duration, CT decay time course) are extracted and compared to the output of the model. Results. Insertion of Voltage Clock (upregulation of If and ICaL, and downregulation of IK1 and Ito) into the model showed the presence of automaticity with the characteristic slow late diastole depolarization. The maximum activation frequencies (0,7Hz) reached only the lower frequency range documented experimentally and found in the litterature (0,4 to 6Hz). Conclusion. Limitations in the rate of automaticity found emphasise the importance of exploring the Calcium Clock mechanism whose insertion may lead to higher spontaneous rates. In the future, a bifurcation analysis will give some insights on the dynamical mechanisms behind pacemaker generation in cultured NRVMs.