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. 2013 Jul 16;105(2):523–532. doi: 10.1016/j.bpj.2013.06.009

Diastolic Field Stimulation: the Role of Shock Duration in Epicardial Activation and Propagation

Marcella C Woods †, Ilija Uzelac ‡, Mark R Holcomb §, John P Wikswo †,‡,¶,‖, Veniamin Y Sidorov †,¶,∗
PMCID: PMC3714876  PMID: 23870273

Abstract

Detailed knowledge of tissue response to both systolic and diastolic shock is critical for understanding defibrillation. Diastolic field stimulation has been much less studied than systolic stimulation, particularly regarding transient virtual anodes. Here we investigated high-voltage-induced polarization and activation patterns in response to strong diastolic shocks of various durations and of both polarities, and tested the hypothesis that the activation versus shock duration curve contains a local minimum for moderate shock durations, and it grows for short and long durations. We found that 0.1–0.2-ms shocks produced slow and heterogeneous activation. During 0.8–1 ms shocks, the activation was very fast and homogeneous. Further shock extension to 8 ms delayed activation from 1.55 ± 0.27 ms and 1.63 ± 0.21 ms at 0.8 ms shock to 2.32 ± 0.41 ms and 2.37 ± 0.3 ms (N = 7) for normal and opposite polarities, respectively. The traces from hyperpolarized regions during 3–8 ms shocks exhibited four different phases: beginning negative polarization, fast depolarization, slow depolarization, and after-shock increase in upstroke velocity. Thus, the shocks of >3 ms in duration created strong hyperpolarization associated with significant delay (P < 0.05) in activation compared with moderate shocks of 0.8 and 1 ms. This effect appears as a dip in the activation-versus-shock-duration curve.

Introduction

Externally applied electric fields are known to cause changes in transmembrane potential (Vm) at locations distant from the stimulating electrodes, a phenomenon termed ”far-field stimulation”. These regions of positive and negative polarization are called virtual cathodes and virtual anodes, respectively. Many explanations for far-field stimulation have been suggested. Plonsey and Barr (1) hypothesized that intercellular resistance at gap junctions causes depolarization and hyperpolarization on opposite ends of each myocyte. However, attempts to experimentally verify the sawtooth effect in intact cardiac tissue have failed (2,3). Theoretical simulations using the bidomain model of cardiac tissue predict that fiber curvature and unequal anisotropy ratios between the intra- and extracellular spaces cause depolarization and hyperpolarization throughout the heart (4–7), whereas the transmural fiber rotation modulates the polarization gradient through the bulk of tissue (5). Ashihara et al. (8) hypothesize that mechanisms responsible for surface and bulk polarizations can cause tunnel propagation of postshock activation, which spreads through the shock-induced intramural excitable area and can underlie the isoelectric window and the induction of fibrillation.

Other heterogeneities have been hypothesized as contributors to the mechanism of far-field stimulation. Computer models incorporating syncytial heterogeneities of the bulk myocardium revealed islands of hyperpolarization and depolarization during an external electric shock (9,10). The contribution of local heterogeneities in the mechanism of the upper level of vulnerability was recently investigated in a theoretical study by Mazeh and Roth (11). Using a combination of bidomain and Beeler-Reuter models to represent passive and active properties of cardiac tissue, they found that local microscopic heterogeneities, represented with random fiber geometry, are crucial for determining the fate of the shock-induced wavefront at the edge of the virtual anode. Due to the higher threshold for microscale heterogeneities in comparison to macroscale ones, the local microscopic heterogeneities can determine the upper level of vulnerability.

At the scale of macroscopic heterogeneities whose dimensions are approximately a few cardiac-length constants, the heart can be modeled as a relatively insulated collection of fibers that, when exposed to an external electrical field, should produce hyperpolarization and depolarization in a patchy manner (12). In addition, cleavage planes between muscle layers may create secondary sources (13–15) of activation, causing nonuniform, anisotropic propagation, and provide a substrate for bulk resetting of the myocardium during defibrillation (13,16,17). Of potentially great practical importance is the recognition that the heterogeneities created by the coronary microvasculature are of critical importance to the defibrillation process (18–20).

The boundary conditions of the heart surfaces also affect how an electric field alters Vm. Latimer and Roth (21) modeled a slab of tissue with an intracavitary electrode in two conditions: the epicardium bounded by air, and the epicardium bounded by a conductive bath. They found that the conductive bath reverses the sign of Vm in some regions and dramatically increases the magnitude of the shock-induced changes in Vm at the epicardial surface. Additionally, Vm falls off rapidly with depth into the tissue, suggesting that the optically measured Vm would be much smaller than the true epicardial Vm. Entcheva et al. (22) modeled an ellipsoid bidomain heart with transmural fiber rotation using different boundary conditions. They found that the induced change in Vm partially results from the tissue boundary conditions and is not fully determined by tissue anisotropy.

Each of these hypotheses incorporates heterogeneities, from microscale to macroscale, as a mechanism for producing adjacent regions of positive and negative polarization during an external electric shock—i.e., cathodal regions that directly stimulate the tissue and accelerate propagation, and anodal regions that can delay propagation and also lead to tissue stimulation upon break (23). These adjacent areas of polarization are potential sources of wavefront generation that could cause the far-field stimulation observed during field shocks (20). The extent of virtual electrode polarization depends on the phase of the cardiac cycle at the time of stimulus application (24–27). During fibrillation, the various portions of the heart exist in states that range between fully refractory and fully recovered conditions; therefore, investigation of the cardiac response to electric stimulation delivered at different action potential (AP) phases is necessary to enable understanding of the mechanisms of defibrillation and for optimizing the shape and strength of defibrillation shock.

Although numerous studies have been dedicated to studying effects of field stimulation on systolic tissue, the effects of field stimulation during diastole have been less investigated. Sharifov and Fast (28) applied shocks transmurally across an isolated pig left ventricle. They found direct and rapid activation of the ventricular bulk over a wide range of shock strengths and delayed activation and multiphasic Vm responses at very strong shock strengths. However, because they were imaging the cut-end of a preparation, the extent to which damage, ischemia, and the angles of the cut fibers with the tissue surface contributed to their results is unknown (29). Recently Maleckar et al. (30) used high-speed optical mapping in conjunction with computer simulation to study the effect of polarity and stimulus strength on the activation time (AT) during diastole. They found that both polarity reversal and increasing shock-strength lower AT. They also indicated the important role of ventricular geometry in the location and size of shock-induced hyperpolarization.

Whereas our earlier work (30) focused primarily on the effect of shock strength on AT, this study examines the complementary role of stimulus duration on AT so that we can test hypotheses that:

  • 1.

    Hyperpolarization caused by long shock durations increases total AT; and

  • 2.

    The activation-versus-shock-duration curve has a local minimum in response to shocks of mild durations, and it rises in response to shocks of both short and long durations.

We operated with a high camera frame speed (5000 frames/s) and used a custom stimulator that could provide pulses as short as 100 μs (31) so that we could resolve for very fast time the transient virtual anodes at the very beginning of the shock. Because of the highly transient expected nature of shock-induced hyperpolarization, we utilized a field shock of strong intensity to investigate the duration-AT relationship during very short stimulation time. We kept this field strength, which at the shortest durations is just above the stimulus threshold, constant for all shock durations, just as we kept shock duration constant in the previous study (30). To our knowledge, this provides new insights into transient virtual anodes that, in typical experiments, are rapidly overrun by wavefronts propagating away from the virtual cathodes.

Methods

Experimental preparation

All experiments followed the guidelines of the National Institutes of Health (Bethesda, MD) for the ethical use of animals in research and were approved in advance by the Vanderbilt University Institutional Animal Care and Use Committee.

Langendorff-perfused hearts (N = 7) from New Zealand White rabbits (2.2–3.5 kg, 3–3.5 months old) were used in this study. A detailed description of the heart preparation has been previously published in Sidorov et al. (32,33). Briefly, the rabbit was preanesthetized with ketamine (50 mg/kg), heparinized (1000 units), and anesthetized by pentobarbital sodium injection (60 mg/kg). The heart was removed and Langendorff-perfused on a custom-built C-shaped glass arm (30). To minimize motion artifacts, the excitation-contraction uncoupler 2,3-butanedione monoxime (BDM; Sigma, St. Louis, MO) in a concentration of 15 mM was added to Tyrode’s solution. The temperature and pH were continuously maintained at 37°C and at 7.4, respectively. The heart was positioned in a temperature-controlled glass chamber such that the anterior side of the heart faced the optical system.

Optical imaging and data acquisition

The heart was stained with a gradual injection of 200 μL of the stock solution of the voltage-sensitive dye di-4-ANEPPS (0.5 mg/mL dimethyl sulfoxide; Molecular Probes, Eugene, OR) via injection port above the aorta. The fluorescence was excited by a diode-pumped, solid-state Verdi laser (Coherent, Santa Clara, CA) at 532 nm. The emitted light was passed through a cutoff filter (607 nm; Tiffen, Hauppauge, NY) and collected by means of a high temporal charge-coupled device camera (26 × 26 pixels, 5000 frames/s; RedShirt Imaging, Decatur, GA). The typical field of view was 5 × 5 cm2.

Stimulation protocol

The heart was paced at a basic cycle length of 300 ms (S1) with a computer-controlled current source (Bloom Associates, Narberth, PA) or a custom-designed physiological stimulator (34) via an insulated bipolar electrode located near the apex. The spatially uniform electric field, produced by means of a custom high-voltage stimulator (31), was applied through two titanium plates positioned at opposite ends of the rectangular bath to face the right and left ventricle. The dimension of the plates was 100 × 120 mm. The S2 field shocks of 50 V/cm in magnitude and of 0.1, 0.2, 0.4, 0.6, 0.8, 1, 1.5, 2, 3, 5, and 8 ms in duration were delivered to diastolic tissue at a S1-S2 coupling interval of 300 ms. Current and voltage waveforms were monitored on a digital phosphor oscilloscope (TDS5034B; Tektronix, Beaverton, OR) via a current probe (TCP305; Tektronix) and a high-voltage differential probe (P5205; Tektronix) connected to two Ag-AgCl electrodes of 2-mm diameter located in the bath (see the Supporting Material). The shocks for which the cathode faced the right ventricle and the anode faced the left ventricle were labeled RV-. Shocks of opposite polarity were referred to as LV- shocks (30).

Data processing and analysis

To preserve the shock timing, no temporal filtering was applied. We followed our previous image processing protocol (30). In summary, we digitally removed from the data pixels corresponding to atrial tissue for all subsequent analysis. The data were normalized on a pixel-by-pixel basis according to the change in fluorescence during S1 pacing response. The resulting normalized data (Fnorm) ranged from 0 (rest) to 1 (peak) for the S1 response. Field shock AT for each ventricular pixel was computed as the elapsed time from shock onset until 50% of the S1 action potential amplitude was reached. To estimate the activation dynamics for the whole heart, the sum of Fnorm for all ventricular pixels for each movie frame was computed and normalized by dividing by the number of pixels summed to yield averaged fluorescence (Favg).

In total, seven rabbit hearts were utilized to study diastolic tissue response to shocks of different durations. Group data are presented as mean values (SD). Statistical analysis was accomplished using the unpaired t-test. Differences were considered significant if P < 0.05.

Results

Effects of field shock duration on Vm distribution patterns during and soon after the shock

Fig. 1 illustrates typical Vm distribution resulting from 50 V/cm shocks of 0.2 ms (Fig. 1 A), 1 ms (Fig. 1 B), and 8 ms (Fig. 1 C) in duration. During a short shock, the negative polarization (blue pixels) of the anodal side and depolarization (red pixels) on the cathodal side of the heart is easily discernible at the time of shock termination (0.2 ms) for both RV- and LV- polarities. The hyperpolarization at both right and left ventricle ends immediately after shock cessation. At 4.2 ms, the fraction of activated tissue encompasses 90.5 and 93.2% for shocks of normal and reversed polarity, respectively. During 1-ms shock, the hyperpolarization is most prominent at approximately the middle of the shock for both polarities (Fig. 1 B, 0.6-ms frames). In contrast to the response to the short S2, in this case activation occurs more quickly and attains 96.1% at 2.2 ms after S2 onset, when the right ventricle faces the cathode (Fig. 1 B, RV-), and is 93.1% at 2.6 ms for the opposite polarity (Fig. 1 B, LV-). The response to an 8-ms shock results in strong positive and negative polarization, which is sustained for a longer period compared to the response to a 1-ms shock. When the left ventricle faces the cathode, the fraction of activated tissue reaches 91.4% at 3.8 ms (Fig. 1 C, LV-), and for the reverse polarity a similar fraction of activation of 91.1% takes place at the 4.2-ms frame. In all hearts the response to an 8-ms shock was much stronger and significantly faster compared to a 0.2-ms shock (P < 0.05, N = 7), and the response to a 1-ms shock was always faster than the response to both 0.2-ms and 8-ms shocks (P < 0.05, N = 7).

Figure 1.

Figure 1

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The Vm distribution as a function of the time in response to 50 V/cm shocks that were (A) short (0.2 ms), (B) intermediate (1.0 ms), or (C) long (8.0 ms). For RV-, the electric field is applied from right to left, and for LV-, from left to right. The data were normalized according to change in fluorescence during S1 response.

Effects of field-shock duration on the activation dynamics

Fig. 2 demonstrates whole heart activation dynamics in response to different S2 duration in one of the heart preparations. It is seen that for both polarities the shortest shocks (solid color, upper row) exhibit slower upstroke velocities than shocks with longer durations (lighter traces). The negative polarization is evident during S2 stimulation as a brief downward deflection at the time of S2. The fluorescence intensity in the time-duration plot (second row) represents the change of activation time with increasing shock duration. The horizontal solid line is hyperpolarization, which becomes thicker with S2 extension. The time-duration plot also indicates that faster activation is caused by S2 stimulation of intermediate duration for either shock polarity. To better show upstroke dynamics, the traces during and soon after shock termination were magnified and are presented in the third row. Hyperpolarization after shock onset can be clearly observed in each curve, including the traces resulting from RV- and LV- shocks of 0.1- and 0.2-ms durations, though in these cases hyperpolarization of the left and right ventricles is of small magnitude and therefore has a minimal contribution in the averaged signal. For all other shock durations and polarities, the negative polarization is strong and reveals itself in the whole heart activation dynamics. The slowest activations correspond to the shortest shock durations of 0.1 and 0.2 ms. As the duration of the shock grows longer, the stronger hyperpolarization dominates in averaged fluorescence and delays the onset of the response and ventricular activation.

Figure 2.

Figure 2

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The whole-heart integrated fluorescence, Favg, as a function of the shock duration and polarity. The shock strength is 50 V/cm. (Left and right columns) Analysis of activation times (ATs) in response to shocks of RV- and LV- polarities, respectively. (First row) Favg for each shock duration with the shortest shocks represented by the darkest text. (Second row) Time-duration plot. (Third row) Magnified view of the first 5 ms of Favg.

The change in the AT histogram from shock duration is displayed in Fig. 3. The corresponding AT maps are shown as insets. The S2 with 0.1-ms duration produces very slow and heterogeneous propagation. The increase in shock duration to 0.4 ms causes bimodal AT distribution with maxima at 0.9 and 2.5 ms for RV- shock and at 1.3 and 2.1 ms for shock of reverse polarity. The first fast peak reflects the activation of the virtual cathode area, whereas the second hump signifies activation of the rest tissue. The 1-ms shock generates fast and homogeneous propagation with maxima of AT distribution at 1.3 ms for both polarities. The successive increase of shock duration to 8 ms causes the histogram to become broader and smaller in amplitude. Such a change is due to hyperpolarization at the virtual anode region, which becomes stronger with S2 extension and impedes rapid activation of the entire heart. The behavior of the AT distribution change is similar for shocks of normal and reverse polarity.

Figure 3.

Figure 3

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Histograms of the AT distribution caused by the shock duration of 0.1, 0.4, 1, and 8 ms. (Insets) Corresponding AT maps.

Fig. 4 shows the total mean AT as a function of shock duration and polarity measured in seven separate experiments. Because of high shock strength (50 V/cm), total mean AT decreases rapidly with increasing duration until 0.8 ms. The fastest total mean AT is 1.55 ± 0.27 ms and 1.63 ± 0.21 ms (N = 7) and corresponds to shock duration of 0.8 ms for RV- and LV- shocks, respectively. Further extension of the shock duration results in delay in activation in both normal and reverse polarities. The total mean AT measured for 3-, 5-, and 8-ms shocks is significantly longer than for 0.8- and 1-ms shocks for both polarities. The increasing S2 duration to 8 ms delays total mean AT to 2.32 × 0.41 ms and 2.37 ± 0.3 ms (N = 7) for RV- and LV- correspondingly. There is no significant difference in total mean AT between two polarities when compared for the same S2 duration.

Figure 4.

Figure 4

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Mean AT, measured from the onset of the shock, as a function of shock duration. The data represent seven separate experiments. Error bars indicate the SD of the mean. For cathode faced to right ventricle: ‡P < 0.05 compared with mean ATs in response to shock duration of 0.1, 0.2, 0.8, and 1 ms, and #P < 0.05 compared with mean ATs for shock duration of 0.8, 1, 3, 5, and 8 ms. For cathode faced to left ventricle: †P < 0.05 compared with mean ATs in response to shock duration of 0.1, 0.2, 0.8, and 1; and ∗P < 0.05 compared with mean ATs for shock duration of 0.8, 1, 3, 5, and 8 ms.

Analysis of the right- and left-ventricle regions of interest

Because two ventricular chambers activate differently depending on the field direction, we analyzed separately the regions of interest during LV- and RV- shocks. Fig. 5 displays mean AT in individual regions on the right and left ventricle as a result of shock duration and polarity. The regions of interest in the right and left ventricles are depicted in activation maps with shaded and solid rectangles, respectively. Two curves exhibit similar dynamics: abrupt descent when shock duration starts to increase, local minimum positioned between 0.4 and 0.8 ms S2, and then ascent of the curve with subsequent extending of the shock. The local minimum and ascending phase is more evident in curves generated from hyperpolarized chambers, whereas for chambers facing the cathode there is little change in mean AT for shocks longer than 1 ms. The dip appears earlier in the curves created from depolarized chambers and indicates very fast activation, which corresponds to 0.91 ± 0.16 ms and 0.84 ± 0.17 ms (N = 36) at shock duration of 0.4 ms for normal and reverse polarity, respectively. In the curves generated from hyperpolarized chambers, the dip occurs later and corresponds to AT of 2.02 ± 0.38 ms at 1-ms shock and 2.07 ± 0.39 ms (N = 36) at 0.8-ms shock for RV- and LV- shock, respectively. It is also evident that activation of negative polarized regions is significantly delayed for all shock durations compared with those for positive polarized regions of interest. The activation maps presented in the lower row clearly demonstrate the difference in activation pattern depending on the shock duration and polarity.

Figure 5.

Figure 5

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AT of right- and left-ventricle regions of interest as a function of shock duration and polarity. (Lower row) Whole heart AT maps for shock duration of 0.2, 1, and 8 ms. (Shaded and solid rectangles) Maps depict the right- and left-ventricle regions of interest. ∗P < 0.01 compared AT in right- and left-ventricle regions of interest.

To characterize further the differences in right- and left-ventricle activation in response to different shock duration and polarity, Favg was computed for regions of interest indicated in Fig. 5 and displayed in Fig. 6. The traces were aligned according to time of shock termination and represent activation during and immediately after the shock. Fig. 6 clearly illustrates dramatically different activation dynamics of the regions in the right and left ventricles caused by change in field direction. In the upper row for RV- shock, when the right ventricle faces the cathode, Favg traces for the right-ventricle region reveal slow activation for shock durations of 0.1 and 0.2 ms, but for all longer shock durations, the activation is rapid and starts at the shock onset. Negative polarization is very prominent in Favg traces computed from the left ventricle facing the anode, especially at long and intermediate shock durations. In general, activation of the left ventricle is much slower in comparison with the right ventricle. The traces from the left ventricle region of interest in response to shock durations of 3, 5, and 8 ms exhibit complex dynamics, which has four phases: negative polarization with shock onset, fast depolarization, slow depolarization, and sharp increase in upstroke velocity after shock termination. The hyperpolarization in the Favg traces lasts for ∼1.5 ms for shock durations of 1.5 ms and greater. The increase in shock duration did not lengthen the time of hyperpolarization. In the lower row, the field direction was reversed so that negative polarization is observed in all of the right-ventricle traces and four-phase behavior is evident for shocks of long duration. In this case, activation for the left ventricle is stronger and faster than for the right ventricle.

Figure 6.

Figure 6

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Favg for right- and left-ventricle regions of interest as a function of shock duration and polarity. (Vertical arrow) Termination of the shock, and all waveforms were aligned by their S2 termination to emphasize both the hyperpolarization of the virtual anodes that occurs at the beginning of each shock, and the rapid activation that occurs immediately at the end of the shock, particularly for the shorter shocks. The timescale was chosen to show activation dynamics during and immediately after the shocks. (Left column) Favg for the right-ventricle region of interest. (Right column) Favg for the left-ventricle region of interest. (Upper row) RV- stimulation. (Lower row) LV- stimulation.

Discussion

The existence of virtual electrodes in the heart as a result of point stimulation was first demonstrated in 1991 (35) and was fully consistent with the predictions of the cardiac bidomain model (36). Subsequently, it became clear that virtual cathodes and anodes supported four different modes of cardiac activation (23). As the significance of virtual electrodes was more widely appreciated, it became clear that they provided the long-sought mechanism for large-scale electrical defibrillation of the heart (4–7). Most recently, an analysis of global cardiac activation time versus shock strength (30) provided insights into how the effect of distributed virtual electrodes depended upon shock strength, but did not address the source of these distributed virtual electrodes. This result contributed to the development of low-energy electrical control of cardiac rhythm disturbances, where several small shocks were shown to be more effective in halting fibrillation than one large one because of the contributions of virtual electrodes that arise from cardiac heterogeneities produced by the branching structure of the cardiac vasculature (19,20).

However, none of these studies resolved the long-running question as to why imaging of activation of the entire heart by field stimulation demonstrated strong virtual cathodes but not the corresponding virtual anodes. We hypothesized that these virtual anodes were transient and were very quickly overrun by activation from anodal break or by wavefronts propagating into them from adjacent virtual cathodes. As the speed of charge-coupled device cameras increased to 5000 frames/s and we developed a custom defibrillator capable of strong defibrillation shocks (∼100 J at 500 volts) with durations as short as 100 μs (31), we began to search for these missing virtual anodes. This article represents a significant extension of our prior work (30) by exploring global activation of the heart not as a function of shock strength but as a function of shock duration, with an emphasis on very short shocks. The need to probe with short shocks then required the use of a strong field, because the amount of charge delivered to the cardiac membrane that is required for activation is proportional to the product of local field strength and duration (37).

In this study we investigated, for the first time to our knowledge, the effects of duration of a strong uniform field shock on polarization pattern and activation dynamics in a diastolic whole heart preparation and tested the hypothesis that:

  • 1.

    Hyperpolarization caused by long shock durations increases AT; and

  • 2.

    The activation-versus-shock-duration curve contains a dip.

To examine this hypothesis, we utilized our custom defibrillator, which could deliver shocks as short as 100 μs (31) and a high-speed optical mapping system capable of imaging 5000 frames/s to image the anterior surface of the isolated rabbit heart. In previous work we have shown that hyperpolarization is an important factor, which, if of sufficient size, can halt propagation, causing a paradoxical increase in AT for stronger shocks (30). The existence of hyperpolarization inside the bulk muscle adds complexity to this phenomenon (8).

Our experimental evidence confirms the important role of hyperpolarization in cardiac tissue activation. Here we demonstrated complex behavior of the AT shock duration relationship, which has a local minimum for S2 duration of 0.8 and 1 ms. The activation of the heart corresponding to these shocks occurred significantly faster than during shorter or longer durations. We did not observe any statistically significant difference in AT for the same shock duration caused by change in field orientation. The separate analysis of right and left ventricles revealed origination of the dip in the AT shock duration curve predominantly in hyperpolarized areas of either right or left ventricle, and the activation of the depolarized chamber was significantly faster than the hyperpolarized.

During long shock duration we observed four different phases in Favg from hyperpolarized areas: initial hyperpolarization at shock onset, rapid depolarization, slow depolarization, and fast depolarization after shock cessation. This complex morphology is similar to the morphology of the optical traces obtained from porcine LV wedge preparations during transmural field stimulation (28). In particular, the second, third, and fourth phases we observed correspond to three phases detected by Sharifov and Fast (28). The difference is that they did not analyze hyperpolarization. However, careful examination of one of the curves in their article (in Fig. 4 C) reveals transient, small magnitude hyperpolarization at shock onset in response to a shock of 38 V/cm of 10 ms in duration. This disagrees with our results, wherein we detect very prominent negative polarization for shocks with these approximate parameters. Such a discrepancy could be explained by a difference between our whole heart and their wedge preparations. In response to strong shocks, the whole heart preparation exhibits rather uniform surface polarization of the right and left ventricle, so that hyperpolarization is not much obstructed by intermittent virtual cathodes, as happens during mapping wedge preparations.

Sharifov and Fast (28) hypothesized that the complex upstroke is a result of electrotonic interaction and spatial averaging of virtual electrode polarization at microscopic heterogeneities. The activation is very rapid at the virtual cathode, while at virtual anodes, negative polarization can occur for the duration of the shock. Spatial averaging by the imaging modality of adjacent virtual cathodes and anodes would lead to observations of multiphasic upstrokes. When a homogeneous electric field affects the whole heart, basically two types of the membrane polarization affect the heart:

  • 1.

    The surface polarization occupies a thin layer that extends to a few length constants (6) and is affected by tissue structure and fiber orientation with respect to the field lines (5).

  • 2.

    The bulk polarization occurs throughout the ventricular wall, mostly due to the specific fiber architecture, in particular the fiber curvature and rotation (5,11).

In addition, the fibers are organized in layers with clefts between them, which together with the heart vascular system increase intramural tissue heterogeneity and enhance the impact of the tissue bulk in shock outcome (17,20). Therefore, the surface polarization at the tissue-bath interface in the field-induced response more likely relates to the fourth phase in upstroke of the optical signal.

Due to the three-dimensional origin of the recorded optical signal, the photon scattering can affect optical AP upstroke and influence accuracy of AT measuring (38–42). Walton et al. (43) have shown that surface AT determined based on maximal slope of upstroke was more precise than that measured at 50% amplitude. Due to deeper optical penetration, this discrepancy was more pronounced for longer wavelength excitation when near-infrared dye was utilized. The upstroke distortion also depends on front orientation with respect to epicardial surface, so that photon scattering is most profound when the wavefront propagates toward or away from the recording surface (41,42). In particular, the maximal slope of upstroke is located near the peak of optical AP if the wave propagates toward the surface, and it is close to the resting value of optical AP when the wave spreads from the surface of the heart (41).

In general, the influence of the photon scattering on the upstroke is most evident when the heart is paced locally from a particular site (43) and during field stimulation with near-threshold intensities (17). Under these circumstances, the majority of the cardiac tissue is activated by propagating excitation waves. In our experiments, a similar condition takes place during stimulation with very short shocks, resulting in the most heterogeneous activation. On the contrary, if shocks are longer than 0.2 ms, a substantial part of the tissue is excited directly via largely extended virtual cathode polarization, and optical AP upstroke would be less distorted. However, when shock duration lengthens for >3 ms and causes an increase in total AT, the virtual anode-mediated strong hyperpolarization precludes earlier activation at these regions and thereby could favor propagation of excitation from intramural virtual cathodes toward surface-hyperpolarized layers. The complex activation dynamics could result in the multiphasic AP upstroke displayed in Fig. 6, where the second and third phases of fast and slow depolarization could reflect intramural excitation. In such manner, the criterion of 50% of amplitude that was utilized in our work more likely shifts AT at the virtual anode regions to be faster and therefore underestimates the hyperpolarization-mediated delay in total AT.

Conclusion

The results of our work are potentially relevant to mechanisms responsible for defibrillation shock failure. It is considered that negative polarization creates excitable gaps and thus produces postshock excitable areas that can serve as substrates for existing or new wavefront precipitation (4). In this context, the time of propagation through these areas is critical for defibrillation shock outcome. If this time is long enough for adjacent tissue to recover, the reentrant electrical activity can revive (44). In conditions where virtual electrode-induced hyperpolarization affects tissue with lower Vm, the hyperpolarization could cause a delay in excitation and thereby hamper the success of the defibrillation.

Limitations

Along with known limitations of the use of BDM (45,46), in this study we did not image the posterior surface of the heart. This information would allow us to analyze and represent the fractional contribution of negative and positive polarization in activation time of the whole heart. However, we believe that it would not change the main results and conclusion of our work relating to the role of hyperpolarization and shock duration in activation dynamics of cardiac tissue.

Acknowledgments

The authors thank Allison Price for assistance with the preparation of this manuscript.

This work was supported by the National Institutes of Health (grant No. R01 HL58241-11) through the American Recovery and Reinvestment Act of 2009 and by the Vanderbilt Institute for Integrative Biosystems Research and Education.

Supporting Material

Document S1. One figure
mmc1.pdf (95KB, pdf)
Document S2. Article plus Supporting Material
mmc2.pdf (1.4MB, pdf)

References

  • 1.Plonsey R., Barr R.C. Effect of microscopic and macroscopic discontinuities on the response of cardiac tissue to defibrillating (stimulating) currents. Med. Biol. Eng. Comput. 1986;24:130–136. doi: 10.1007/BF02443925. [DOI] [PubMed] [Google Scholar]
  • 2.Gillis A.M., Fast V.G., Kléber A.G. Spatial changes in transmembrane potential during extracellular electrical shocks in cultured monolayers of neonatal rat ventricular myocytes. Circ. Res. 1996;79:676–690. doi: 10.1161/01.res.79.4.676. [DOI] [PubMed] [Google Scholar]
  • 3.Zhou X., Knisley S.B., Ideker R.E. Spatial changes in the transmembrane potential during extracellular electric stimulation. Circ. Res. 1998;83:1003–1014. doi: 10.1161/01.res.83.10.1003. [DOI] [PubMed] [Google Scholar]
  • 4.Efimov I.R., Aguel F., Trayanova N. Virtual electrode polarization in the far field: implications for external defibrillation. Am. J. Physiol. Heart Circ. Physiol. 2000;279:H1055–H1070. doi: 10.1152/ajpheart.2000.279.3.H1055. [DOI] [PubMed] [Google Scholar]
  • 5.Entcheva E., Trayanova N.A., Claydon F.J. Patterns of and mechanisms for shock-induced polarization in the heart: a bidomain analysis. IEEE Trans. Biomed. Eng. 1999;46:260–270. doi: 10.1109/10.748979. [DOI] [PubMed] [Google Scholar]
  • 6.Trayanova N.A., Roth B.J., Malden L.J. The response of a spherical heart to a uniform electric field: a bidomain analysis of cardiac stimulation. IEEE Trans. Biomed. Eng. 1993;40:899–908. doi: 10.1109/10.245611. [DOI] [PubMed] [Google Scholar]
  • 7.Wikswo J.P., Roth B.J. Virtual electrode theory of pacing. In: Efimov I., Kroll M., Tchou P., editors. Cardiac Bioelectric Therapy: Mechanisms and Practical Implications. Springer; New York: 2009. pp. 283–330. [Google Scholar]
  • 8.Ashihara T., Constantino J., Trayanova N.A. Tunnel propagation of postshock activations as a hypothesis for fibrillation induction and isoelectric window. Circ. Res. 2008;102:737–745. doi: 10.1161/CIRCRESAHA.107.168112. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 9.Fishler M.G. Syncytial heterogeneity as a mechanism underlying cardiac far-field stimulation during defibrillation-level shocks. J. Cardiovasc. Electrophysiol. 1998;9:384–394. doi: 10.1111/j.1540-8167.1998.tb00926.x. [DOI] [PubMed] [Google Scholar]
  • 10.Pumir A., Nikolski V., Krinsky V. Wave emission from heterogeneities opens a way to controlling chaos in the heart. Phys. Rev. Lett. 2007;99:208101. doi: 10.1103/PhysRevLett.99.208101. [DOI] [PubMed] [Google Scholar]
  • 11.Mazeh N., Roth B.J. A mechanism for the upper limit of vulnerability. Heart Rhythm. 2009;6:361–367. doi: 10.1016/j.hrthm.2008.11.010. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 12.Krassowska W., Kumar M.S. The role of spatial interactions in creating the dispersion of transmembrane potential by premature electric shocks. Ann. Biomed. Eng. 1997;25:949–963. [PubMed] [Google Scholar]
  • 13.Fast V.G., Rohr S., Kléber A.G. Activation of cardiac tissue by extracellular electrical shocks: formation of ‘secondary sources’ at intercellular clefts in monolayers of cultured myocytes. Circ. Res. 1998;82:375–385. doi: 10.1161/01.res.82.3.375. [DOI] [PubMed] [Google Scholar]
  • 14.Plonsey R., Barr R.C. Inclusion of junction elements in a linear cardiac model through secondary sources: application to defibrillation. Med. Biol. Eng. Comput. 1986;24:137–144. doi: 10.1007/BF02443926. [DOI] [PubMed] [Google Scholar]
  • 15.Sobie E.A., Susil R.C., Tung L. A generalized activating function for predicting virtual electrodes in cardiac tissue. Biophys. J. 1997;73:1410–1423. doi: 10.1016/S0006-3495(97)78173-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 16.Hooks D.A., Tomlinson K.A., Hunter P.J. Cardiac microstructure: implications for electrical propagation and defibrillation in the heart. Circ. Res. 2002;91:331–338. doi: 10.1161/01.res.0000031957.70034.89. [DOI] [PubMed] [Google Scholar]
  • 17.Zemlin C.W., Mironov S., Pertsov A.M. Near-threshold field stimulation: intramural versus surface activation. Cardiovasc. Res. 2006;69:98–106. doi: 10.1016/j.cardiores.2005.08.012. [DOI] [PubMed] [Google Scholar]
  • 18.Bishop M.J., Boyle P.M., Vigmond E.J. Modeling the role of the coronary vasculature during external field stimulation. IEEE Trans. Biomed. Eng. 2010;57:2335–2345. doi: 10.1109/TBME.2010.2051227. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 19.Gray R.A., Wikswo J.P. Cardiovascular disease: several small shocks beat one big one. Nature. 2011;475:181–182. doi: 10.1038/475181a. [DOI] [PubMed] [Google Scholar]
  • 20.Luther S., Fenton F.H., Bodenschatz E. Low-energy control of electrical turbulence in the heart. Nature. 2011;475:235–239. doi: 10.1038/nature10216. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 21.Latimer D.C., Roth B.J. Effect of a bath on the epicardial transmembrane potential during internal defibrillation shocks. IEEE Trans. Biomed. Eng. 1999;46:612–614. doi: 10.1109/10.759063. [DOI] [PubMed] [Google Scholar]
  • 22.Entcheva E., Eason J., Claydon F. Virtual electrode effects in transvenous defibrillation-modulation by structure and interface: evidence from bidomain simulations and optical mapping. J. Cardiovasc. Electrophysiol. 1998;9:949–961. doi: 10.1111/j.1540-8167.1998.tb00135.x. [DOI] [PubMed] [Google Scholar]
  • 23.Wikswo J.P., Jr., Lin S.-F., Abbas R.A. Virtual electrodes in cardiac tissue: a common mechanism for anodal and cathodal stimulation. Biophys. J. 1995;69:2195–2210. doi: 10.1016/S0006-3495(95)80115-3. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 24.Cheng D.K., Tung L., Sobie E.A. Nonuniform responses of transmembrane potential during electric field stimulation of single cardiac cells. Am. J. Physiol. 1999;277:H351–H362. doi: 10.1152/ajpheart.1999.277.1.H351. [DOI] [PubMed] [Google Scholar]
  • 25.Fast V.G., Rohr S., Ideker R.E. Nonlinear changes of transmembrane potential caused by defibrillation shocks in strands of cultured myocytes. Am. J. Physiol. Heart Circ. Physiol. 2000;278:H688–H697. doi: 10.1152/ajpheart.2000.278.3.H688. [DOI] [PubMed] [Google Scholar]
  • 26.Gray R.A., Huelsing D.J., Trayanova N.A. Effect of strength and timing of transmembrane current pulses on isolated ventricular myocytes. J. Cardiovasc. Electrophysiol. 2001;12:1129–1137. doi: 10.1046/j.1540-8167.2001.01129.x. [DOI] [PubMed] [Google Scholar]
  • 27.Zhou X., Smith W.M., Ideker R.E. Transmembrane potential changes caused by shocks in guinea pig papillary muscle. Am. J. Physiol. 1996;271:H2536–H2546. doi: 10.1152/ajpheart.1996.271.6.H2536. [DOI] [PubMed] [Google Scholar]
  • 28.Sharifov O.F., Fast V.G. Optical mapping of transmural activation induced by electrical shocks in isolated left ventricular wall wedge preparations. J. Cardiovasc. Electrophysiol. 2003;14:1215–1222. doi: 10.1046/j.1540-8167.2003.03188.x. [DOI] [PubMed] [Google Scholar]
  • 29.Efimov I.R., Nikolski V.P. Diastolic shocking experience: do virtual electrodes exist only during systole? J. Cardiovasc. Electrophysiol. 2003;14:1223–1224. doi: 10.1046/j.1540-8167.2003.03442.x. [DOI] [PubMed] [Google Scholar]
  • 30.Maleckar M.M., Woods M.C., Trayanova N.A. Polarity reversal lowers activation time during diastolic field stimulation of the rabbit ventricles: insights into mechanisms. Am. J. Physiol. Heart Circ. Physiol. 2008;295:H1626–H1633. doi: 10.1152/ajpheart.00706.2008. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Mashburn D.N., Hinkson S.J., Wikswo J.P. A high-voltage cardiac stimulator for field shocks of a whole heart in a bath. Rev. Sci. Instrum. 2007;78:104302. doi: 10.1063/1.2796832. [DOI] [PubMed] [Google Scholar]
  • 32.Sidorov V.Y., Woods M.C., Baudenbacher F. Examination of stimulation mechanism and strength-interval curve in cardiac tissue. Am. J. Physiol. Heart Circ. Physiol. 2005;289:H2602–H2615. doi: 10.1152/ajpheart.00968.2004. [DOI] [PubMed] [Google Scholar]
  • 33.Sidorov V.Y., Woods M.C., Wikswo J.P. Effects of elevated extracellular potassium on the stimulation mechanism of diastolic cardiac tissue. Biophys. J. 2003;84:3470–3479. doi: 10.1016/S0006-3495(03)70067-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 34.Holcomb M.R., Devine J.M., Sidorov V.Y. Continuous-waveform constant-current isolated physiological stimulator. Rev. Sci. Instrum. 2012;83:044303. doi: 10.1063/1.3700977. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 35.Wikswo J.P., Jr., Wisialowski T.A., Roden M. Virtual cathode effects during stimulation of cardiac muscle: two-dimensional in vivo measurements. Cardiovasc. Res. 1991;68:513–530. doi: 10.1161/01.res.68.2.513. [DOI] [PubMed] [Google Scholar]
  • 36.Sepulveda N.G., Roth B.J., Wikswo J.P., Jr. Current injection into a two-dimensional anisotropic bidomain. Biophys. J. 1989;55:987–999. doi: 10.1016/S0006-3495(89)82897-8. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 37.Malkin R.A., Guan D., Wikswo J.P. Experimental evidence of improved transthoracic defibrillation with electroporation-enhancing pulses. IEEE Trans. Biomed. Eng. 2006;53:1901–1910. doi: 10.1109/TBME.2006.881787. [DOI] [PubMed] [Google Scholar]
  • 38.Bishop M.J., Rodriguez B., Trayanova N.A. The role of photon scattering in optical signal distortion during arrhythmia and defibrillation. Biophys. J. 2007;93:3714–3726. doi: 10.1529/biophysj.107.110981. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Bishop M.J., Rodriguez B., Gavaghan D.J. Inference of intramural wavefront orientation from optical recordings in realistic whole-heart models. Biophys. J. 2006;91:3957–3958. doi: 10.1529/biophysj.106.092544. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 40.Hyatt C.J., Mironov S.F., Pertsov A.M. Optical action potential upstroke morphology reveals near-surface transmural propagation direction. Circ. Res. 2005;97:277–284. doi: 10.1161/01.RES.0000176022.74579.47. [DOI] [PubMed] [Google Scholar]
  • 41.Hyatt C.J., Mironov S.F., Pertsov A.M. Synthesis of voltage-sensitive fluorescence signals from three-dimensional myocardial activation patterns. Biophys. J. 2003;85:2673–2683. doi: 10.1016/s0006-3495(03)74690-6. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 42.Pertsov A.M., Zemlin C.W., Bernus O. What can we learn from the optically recorded epicardial action potential? Biophys. J. 2006;91:3959–3960. doi: 10.1529/biophysj.106.091835. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 43.Walton R.D., Smith R.M., Pertsov A.M. Extracting surface activation time from the optically recorded action potential in three-dimensional myocardium. Biophys. J. 2012;102:30–38. doi: 10.1016/j.bpj.2011.10.036. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 44.Trayanova N., Plank G., Rodríguez B. What have we learned from mathematical models of defibrillation and postshock arrhythmogenesis? Application of bidomain simulations. Heart Rhythm. 2006;3:1232–1235. doi: 10.1016/j.hrthm.2006.04.015. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 45.Biermann M., Rubart M., Zipes D.P. Differential effects of cytochalasin D and 2,3 butanedione monoxime on isometric twitch force and transmembrane action potential in isolated ventricular muscle: implications for optical measurements of cardiac repolarization. J. Cardiovasc. Electrophysiol. 1998;9:1348–1357. doi: 10.1111/j.1540-8167.1998.tb00110.x. [DOI] [PubMed] [Google Scholar]
  • 46.Liu Y., Cabo C., Jalife J. Effects of diacetyl monoxime on the electrical properties of sheep and guinea pig ventricular muscle. Cardiovasc. Res. 1993;27:1991–1997. doi: 10.1093/cvr/27.11.1991. [DOI] [PubMed] [Google Scholar]

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