Muscular Thin Films for Label-Free Mapping of Excitation Propagation in Cardiac Tissue.

Muscular thin films (MTFs), have already found a variety of applications in cardiac tissue engineering and in building of lab-on-a-chip systems. Here we present a novel approach to label-free mapping of excitation waves in the cardiomyocyte cell cultures with the use of MTFs. Neonatal rat ventricular cardiomyocytes were cultured on polydimethylsiloxane (PDMS) thin films and observed by means of off-axis illumination. Inflexions of the membrane created by the contraction of cardiomyocytes led to formation of patterns of bright and dark areas on the surface of the membrane. These patterns were recorded and analyzed for the monitoring of the contraction propagation. The method was compared with a standard optical mapping technique based on the use of a Ca2+-sensitive fluorescent dye. A good consistency of the results obtained by these two methods was demonstrated. The proposed method is non-toxic and might be of particular interest for the purpose of continuous monitoring in test systems based on human induced pluripotent stem cells.

Self-organization of conducting pathways explains electrical wave propagation in cardiac tissues with high fraction of non-conducting cells.

Cardiac fibrosis occurs in many forms of heart disease and is considered to be one of the main arrhythmogenic factors. Regions with a high density of fibroblasts are likely to cause blocks of wave propagation that give rise to dangerous cardiac arrhythmias. Therefore, studies of the wave propagation through these regions are very important, yet the precise mechanisms leading to arrhythmia formation in fibrotic cardiac tissue remain poorly understood. Particularly, it is not clear how wave propagation is organized at the cellular level, as experiments show that the regions with a high percentage of fibroblasts (65-75%) are still conducting electrical signals, whereas geometric analysis of randomly distributed conducting and non-conducting cells predicts connectivity loss at 40% at the most (percolation threshold). To address this question, we used a joint in vitro-in silico approach, which combined experiments in neonatal rat cardiac monolayers with morphological and electrophysiological computer simulations. We have shown that the main reason for sustainable wave propagation in highly fibrotic samples is the formation of a branching network of cardiomyocytes. We have successfully reproduced the morphology of conductive pathways in computer modelling, assuming that cardiomyocytes align their cytoskeletons to fuse into cardiac syncytium. The electrophysiological properties of the monolayers, such as conduction velocity, conduction blocks and wave fractionation, were reproduced as well. In a virtual cardiac tissue, we have also examined the wave propagation at the subcellular level, detected wavebreaks formation and its relation to the structure of fibrosis and, thus, analysed the processes leading to the onset of arrhythmias.

Stilbene derivative as a photosensitive compound to control the excitability of neonatal rat cardiomyocytes.

Substances that can be used as photosensitizers for cardiac tissue are very helpful in modeling various excitation patterns in a cardiac tissue culture and may have prospective use in the temporary and permanent ablation of unwanted excitation sources in the heart.The aim of the present work is to study the effect of stilbene derivative c-TAB (2- {4- [(E) -2- (4-ethoxyphenyl) vinyl] phenoxy} ethyl) trimethylammonium bromide) on the cardiomyocyte layers and voltage-gated ion channels in cardiac cells. C-TAB is a structural analog to AzoTAB, reported previously as a photoswitch for cardiac and neural cells, in which the azobenzene moiety is replaced by a stilbene grouping. Such a replacement makes c-TAB less toxic to living cells. c-TAB has been shown to successfully inhibit excitation in cardiac cells in both trans- and cis- forms. The excitation inhibition of cardiac cells under c-TAB is reversible and can be overturned easily by washing out the c-TAB; however, not by light illumination. The irradiation of cardiac cells with near-UV, when the trans- form of c-TAB is applied, changes reversible inhibition to a permanent one that cannot be overturned by a washout.

Biocontractile microfluidic channels for peristaltic pumping.

Bio-actuated micro-pumps do not need any external power source and pose no risk of electrical or heat shock for the biological materials in lab-on-chip systems. Several different designs of bio-actuated micro-pumps based on the use of the contractile force of cultured cardiomyocites have been proposed earlier. Here we present a novel type of a bio-actuated micro-pump representing a microfluidic channel with a contractile wall. The flow inside the channel is generated by the peristaltic movement of its wall caused by the propagation of an excitation-contraction wave along the channels surface. The directional flow generated by the pump was demonstrated by tracking of polystyrene microspheres, moving in the direction of the propagation of the excitation-contraction wave with an average velocity of 6-8 µm/min. The suggested design of a micro-pump allows the control of pumping direction, which might be useful for targeted delivery of fluids and substances in lab-on-chip systems. Prospects of future development and implementation of this kind of bio-actuated peristaltic pumps are discussed.

Photocontrol of Voltage-Gated Ion Channel Activity by Azobenzene Trimethylammonium Bromide in Neonatal Rat Cardiomyocytes.

The ability of azobenzene trimethylammonium bromide (azoTAB) to sensitize cardiac tissue excitability to light was recently reported. The dark, thermally relaxed trans- isomer of azoTAB suppressed spontaneous activity and excitation propagation speed, whereas the cis- isomer had no detectable effect on the electrical properties of cardiomyocyte monolayers. As the membrane potential of cardiac cells is mainly controlled by activity of voltage-gated ion channels, this study examined whether the sensitization effect of azoTAB was exerted primarily via the modulation of voltage-gated ion channel activity. The effects of trans- and cis- isomers of azoTAB on voltage-dependent sodium (INav), calcium (ICav), and potassium (IKv) currents in isolated neonatal rat cardiomyocytes were investigated using the whole-cell patch-clamp technique. The experiments showed that azoTAB modulated ion currents, causing suppression of sodium (Na+) and calcium (Ca2+) currents and potentiation of net potassium (K+) currents. This finding confirms that azoTAB-effect on cardiac tissue excitability do indeed result from modulation of voltage-gated ion channels responsible for action potential.

Conditions for Waveblock Due to Anisotropy in a Model of Human Ventricular Tissue.

Waveblock formation is the main cause of reentry. We have performed a comprehensive numerical modeling study of block formation due to anisotropy in Ten Tusscher and Panfilov (2006) ionic model for human ventricular tissue. We have examined the border between different areas of myocardial fiber alignment and have shown that blockage can occur for a wave traveling from a transverse fiber area to a longitudinal one. Such blockage occurs for reasonable values of the anisotropy ratio (AR): from 2.4 to 6.2 with respect to propagation velocities. This critical AR decreases by the suppression of INa and ICa, slightly decreases by the suppression of IKr and IKs, and substantially increases by the suppression of IK1. Hyperkalemia affects the block formation in a complex, biphasic way. We provide examples of reentry formation due to the studied effects and have concluded that the suppression of IK1 should be the most effective way to prevent waveblock at the areas of abrupt change in anisotropy.