28 IEEE TRANSACTIONS ON BIOMEDICAL ENGINEERING, VOL. 60, NO. 1, JANUARY 2013 Greater Cardiac Cell Excitation Efficiency With Rapidly Switching Multidirectional Electrical Stimulation Alexandra V. S. Fonseca, Rosana A. Bassani ∗ , Pedro X. Oliveira, and Jos´ e W. M. Bassani Abstract—Electric field (E) stimulation is widely used in experi- ments with myocardial preparations and in the clinical setting (e.g., defibrillation). As a rule, stimuli are applied in a single direction, which limits excitatory cell recruitment because myocytes are dis- posed in different directions and their sensitivity to E depends on the stimulus orientation with respect to the cell major axis. Here, we propose a stimulatory approach, namely rapidly switching mul- tidirectional stimulation (RSMS), in which stimuli are delivered in three directions within the electric refractory period. In popu- lations of randomly oriented isolated rat cardiomyocytes, RSMS doubled the percentage of cells excited by near-threshold E (P< 0.001), which was more than the increase in recruitment in a single direction achieved by doubling E intensity. This effect was similar for monophasic and biphasic pulses, but for the latter, a given per- cent recruitment was obtained with 20–30% lower E intensity (P < 0.01), so that RSMS with biphasic pulses allowed at least 60% re- duction of E intensity for recruitment of >70% of the cells. RSMS can be applied to improve stimulation efficiency in experiments with isolated cardiac myocytes, and may be a promising alterna- tive for decreasing shock intensity requirements for cardioversion and defibrillation. Index Terms—Cardiology, electric stimulation, electrical field, method. I. INTRODUCTION M YOCARDIAL electric stimulation is widely used not only for clinical purposes (pacemaking, cardioversion, defibrillation, and cardiac electrophysiological tests), but also experimentally, for in vitro pacing of isolated preparations (my- ocytes, cardiac muscle, and perfused heart), and in the devel- opment of models of cardiovascular disease (e.g., rapid pacing- induced heart failure). During stimulation with electric fields (E), as it is typically the case of stimulation of isolated cells and transthoracic de- Manuscript received July 18, 2012; revised September 17, 2012; accepted September 18, 2012. Date of publication September 28, 2012; date of current version December 14, 2012. This work was supported in part by the Con- selho Nacional de Desenvolvimento Cient´ ıfico e Tecnol´ ogico (CNPq, Proc N. 300632/2005-3), Brazil. The work of A. V. S. Fonseca was supported by a scholarship from Coordenac ¸˜ ao de Aperfeic ¸oamento de Pessoal de N´ ıvel Supe- rior, Brazil. Asterisk indicates corresponding author. A. V. S. Fonseca is with the Boston Scientific, 05804-900 S˜ ao Paulo-SP, Brazil (e-mail: Alexandra.Fonseca@bsci.com). ∗ R. A. Bassani is with the Center for Biomedical Engineering, University of Campinas, 13083-881 Campinas-SP, Brazil (e-mail: rosana@ceb.unicamp.br). P. X. Oliveira and J. W. M. Bassani are with the Department of Biomedi- cal Engineering, School of Electrical and Computer Engineering, University of Campinas, 13083-881 Campinas-SP, Brazil (e-mail: pedrox@ceb.unicamp.br; bassani@ceb.unicamp.br). Digital Object Identifier 10.1109/TBME.2012.2220766 fibrillation, the threshold intensity is highly dependent on the direction of E application. Several studies have shown that sen- sitivity to stimulation is the greatest when E is oriented longitu- dinally to the cell major axis, and the lowest for the transversal orientation [1]–[4]. Nevertheless, in the whole heart, fibers are arranged in different directions [5], [6]. Likewise, isolated my- ocytes plated on a typical perfusion chamber tend to adhere to the bottom at random orientations. This implies that stimulus in- tensity must be relatively high to produce electrical recruitment of a significant percentage of cells. One could hypothesize that if stimulation could be applied in different directions, it would be possible to achieve a large recruitment with comparatively lower stimulus intensity. However, alternating stimulus direc- tion would not be the best solution. Because different sets of cells would respond to stimulation in each direction, and some cells might respond to more than one direction, it would not be possible to maintain a regular pacing rate (which is important during stimulation of isolated cells) or achieve synchronization (which is crucial for cardioversion/defibrillation). In the present report, we propose a procedure for multi- directional E stimulation (rapidly switching multidirectional stimulation—RSMS) that allows synchronized excitation by switching the stimulus direction during the refractory period, so that cells that respond in a given direction cannot be reex- cited by the next stimulus. The aims of this study were 1) to develop instrumentation for RSMS, in which stimuli applied in three directions are switched within a few tens of milliseconds; 2) to test whether this approach enhances the efficiency of stim- ulation of isolated rat ventricular myocytes; and 3) to investigate whether the stimulus waveform modifies the response to RSMS. II. MATERIALS AND METHODS A. Myocardial Preparation Myocytes were isolated from the left ventricle of adult male Wistar rats by collagenase digestion during coronary perfusion at 37 ◦ C [7]. Rats were maintained at 23 ± 2 ◦ C, under a 12 h:12 h light–dark cycle, and were offered water and pelleted chow ad libitum. Animals were killed by exsanguination fol- lowing cerebral concussion. The protocols of animal care and euthanasia were in agreement with the principles of the Brazil- ian Society of Laboratory Animal Science (SBCAL) and were approved by the institutional Committee of Ethics in Animal Use (CEUA/IB/UNICAMP, protocol numbers P1186-1, P1517- 1, and P1737-1). 0018-9294/$31.00 © 2012 IEEE