Rhythms from Two Competing Periodic Sources Embedded in an Excitable Medium.

In an excitable medium, a stimulus generates a wave that propagates in space until it reaches the boundary or collides with another wave and annihilates. We study the dynamics generated by two periodic sources with different frequencies in excitable cardiac tissue culture using optogenetic techniques. The observed rhythms, which can be modeled using cellular automata and studied analytically, show unexpected regularities related to classic results in number theory. We apply the results to identify cardiac arrhythmias in people that are due to a putative mechanism of two competing pacemakers.

Universal mechanisms for self-termination of rapid cardiac rhythm.

Excitable media sustain circulating waves. In the heart, sustained circulating waves can lead to serious impairment or even death. To investigate factors affecting the stability of such waves, we have used optogenetic techniques to stimulate a region at the apex of a mouse heart at a fixed delay after the detection of excitation at the base of the heart. For long delays, rapid circulating rhythms can be sustained, whereas for shorter delays, there are paroxysmal bursts of activity that start and stop spontaneously. By considering the dependence of the action potential and conduction velocity on the preceding recovery time using restitution curves, as well as the reduced excitability (fatigue) due to the rapid excitation, we model prominent features of the dynamics including alternation of the duration of the excited phases and conduction times, as well as termination of the bursts for short delays. We propose that this illustrates universal mechanisms that exist in biological systems for the self-termination of such activities.

Double-wave reentry in excitable media.

A monolayer of chick embryo cardiac cells grown in an annular geometry supports two simultaneous reentrant excitation waves that circulate as a doublet. We propose a mechanism that can lead to such behavior. The velocity restitution gives the instantaneous velocity of a wave as a function of the time since the passage of the previous wave at a given point in space. Nonmonotonic restitution relationships will lead to situations in which various spacings between circulating waves are possible. In cardiology, the situation in which two waves travel in an anatomically defined circuit is referred to as double-wave reentry. Since double-wave reentry may arise as a consequence of pacing during cardiac arrhythmias, understanding the dynamic features of double-wave reentry may be helpful in understanding the physiological properties of cardiac tissue and in the design of therapy.

Caveolae in Rabbit Ventricular Myocytes: Distribution and Dynamic Diminution after Cell Isolation.

Caveolae are signal transduction centers, yet their subcellular distribution and preservation in cardiac myocytes after cell isolation are not well documented. Here, we quantify caveolae located within 100 nm of the outer cell surface membrane in rabbit single-ventricular cardiomyocytes over 8 h post-isolation and relate this to the presence of caveolae in intact tissue. Hearts from New Zealand white rabbits were either chemically fixed by coronary perfusion or enzymatically digested to isolate ventricular myocytes, which were subsequently fixed at 0, 3, and 8 h post-isolation. In live cells, the patch-clamp technique was used to measure whole-cell plasma membrane capacitance, and in fixed cells, caveolae were quantified by transmission electron microscopy. Changes in cell-surface topology were assessed using scanning electron microscopy. In fixed ventricular myocardium, dual-axis electron tomography was used for three-dimensional reconstruction and analysis of caveolae in situ. The presence and distribution of surface-sarcolemmal caveolae in freshly isolated cells matches that of intact myocardium. With time, the number of surface-sarcolemmal caveolae decreases in isolated cardiomyocytes. This is associated with a gradual increase in whole-cell membrane capacitance. Concurrently, there is a significant increase in area, diameter, and circularity of sub-sarcolemmal mitochondria, indicative of swelling. In addition, electron tomography data from intact heart illustrate the regular presence of caveolae not only at the surface sarcolemma, but also on transverse-tubular membranes in ventricular myocardium. Thus, caveolae are dynamic structures, present both at surface-sarcolemmal and transverse-tubular membranes. After cell isolation, the number of surface-sarcolemmal caveolae decreases significantly within a time frame relevant for single-cell research. The concurrent increase in cell capacitance suggests that membrane incorporation of surface-sarcolemmal caveolae underlies this, but internalization and/or micro-vesicle loss to the extracellular space may also contribute. Given that much of the research into cardiac caveolae-dependent signaling utilizes isolated cells, and since caveolae-dependent pathways matter for a wide range of other study targets, analysis of isolated cell data should take the time post-isolation into account.

Hypertension-induced remodelling: on the interactions of cardiac risk factors.

Hypertension induces considerable cardiac remodelling, such as hypertrophy, interstitial fibrosis, and abnormal activity of the cardiac sympathetic nervous system, which are established risk factors in several highly dangerous heart diseases, such as ventricular fibrillation and congestive heart failure. All these risk factors and heart diseases are studied extensively in isolation, but to our knowledge, there is no comprehensive review of their interactions. At the same time, there is growing evidence suggesting that such interactions are numerous and that a successful therapy against a particular condition may have unexpectedly weak effects on mortality, as treated patients may die of a different cause exacerbated by the therapy. In this article, we present a multiscale review of the literature focusing on the relationships between the above-mentioned risk factors and heart diseases, and introduce a framework that gives insight into their possible interactions. We use this framework to demonstrate that conditions such as fibrosis and elevated activity of the sympathetic nervous system may be compensatory, rather than purely pathological, mechanisms in certain contexts. Finally, we show why the described mechanisms are relevant not only in hypertension, but also in the case of healed myocardial infarction.

ß-Adrenergic receptor stimulation inhibits proarrhythmic alternans in postinfarction border zone cardiomyocytes: a computational analysis.

The border zone (BZ) of the viable myocardium adjacent to an infarct undergoes extensive autonomic and electrical remodeling and is prone to repolarization alternans-induced cardiac arrhythmias. BZ remodeling processes may promote or inhibit Ca2+ and/or repolarization alternans and may differentially affect ventricular arrhythmogenesis. Here, we used a detailed computational model of the canine ventricular cardiomyocyte to study the determinants of alternans in the BZ and their regulation by ß-adrenergic receptor (ß-AR) stimulation. The BZ model developed Ca2+ transient alternans at slower pacing cycle lengths than the control model, suggesting that the BZ may promote spatially heterogeneous alternans formation in an infarcted heart. ß-AR stimulation abolished alternans. By evaluating all combinations of downstream ß-AR stimulation targets, we identified both direct (via ryanodine receptor channels) and indirect [via sarcoplasmic reticulum (SR) Ca2+ load] modulation of SR Ca2+ release as critical determinants of Ca2+ transient alternans. These findings were confirmed in a human ventricular cardiomyocyte model. Cell-to-cell coupling indirectly modulated the likelihood of alternans by affecting the action potential upstroke, reducing the trigger for SR Ca2+ release in one-dimensional strand simulations. However, ß-AR stimulation inhibited alternans in both single and multicellular simulations. Taken together, these data highlight a potential antiarrhythmic role of sympathetic hyperinnervation in the BZ by reducing the likelihood of alternans and provide new insights into the underlying mechanisms controlling Ca2+ transient and repolarization alternans.NEW & NOTEWORTHY We integrated, for the first time, postmyocardial infarction electrical and autonomic remodeling in a detailed, validated computer model of ß-adrenergic stimulation in ventricular cardiomyocytes. Here, we show that ß-adrenergic stimulation inhibits alternans and provide novel insights into underlying mechanisms, adding to a recent controversy about pro-/antiarrhythmic effects of postmyocardial infarction hyperinnervation.Listen to this article's corresponding podcast at http://ajpheart.podbean.com/e/%CE%B2-ar-stimulation-and-alternans-in-border-zone-cardiomyocytes/.

Ccoffinn: Automated Wave Tracking in Cultured Cardiac Monolayers.

Cardiac arrhythmias are one of the most frequent causes of death worldwide. A popular biological model used to study arrhythmogenesis is the cultured cardiac cell monolayer, which provides a good trade-off between physiological relevance and experimental access. Excitation wave patterns are imaged using high-bandwidth detectors, producing large data sets that are typically analyzed manually. To make such analysis less time consuming and less subjective, we have designed and implemented a toolkit for segmentation and tracking of cardiac waves in optical mapping recordings. The toolkit is optimized for high-resolution detectors to accommodate the growing availability of inexpensive high-resolution detectors for life science imaging applications (e.g., scientific CMOS cameras). The software extracts key features of propagating waves, such as wavefront speed and entropy. The methods have been validated using synthetic data, and real-world examples are provided, showing a difference in conduction velocity between two different types of cardiac cell cultures.

All-optical control of cardiac excitation: combined high-resolution optogenetic actuation and optical mapping.

Cardiac tissue is an excitable system that can support complex spatiotemporal dynamics, including instabilities (arrhythmias) with lethal consequences. While over the last two decades optical mapping of excitation (voltage and calcium dynamics) has facilitated the detailed characterization of such arrhythmia events, until recently, no precise tools existed to actively interrogate cardiac dynamics in space and time. In this work, we discuss the combined use of new methods for space- and time-resolved optogenetic actuation and simultaneous fast, high resolution optical imaging of cardiac excitation waves. First, the mechanisms, limitations and unique features of optically induced responses in cardiomyocytes are outlined. These include the ability to bidirectionally control the membrane potential using depolarizing and hyperpolarizing opsins; the ability to induce prolonged sustained voltage changes; and the ability to control refractoriness and the shape of the cardiac action potential. At the syncytial tissue level, we discuss optogenetically enabled experimentation on cell-cell coupling, alteration of conduction properties and termination of propagating waves by light. Specific attention is given to space- and time-resolved application of optical stimulation using dynamic light patterns to perturb ongoing activation and to probe electrophysiological properties at desired tissue locations. The combined use of optical methods to perturb and to observe the system can offer new tools for precise feedback control of cardiac electrical activity, not available previously with pharmacological and electrical stimulation. These new experimental tools for all-optical electrophysiology allow for a level of precise manipulation and quantification of cardiac dynamics comparable in robustness to the computational setting, and can provide new insights into pacemaking, arrhythmogenesis and suppression or cardioversion.

Protection against ventricular fibrillation via cholinergic receptor stimulation and the generation of nitric oxide.

KEY POINTS: Animal studies suggest an anti-fibrillatory action of the vagus nerve on the ventricle, although the exact mechanism is controversial. Using a Langendorff perfused rat heart, we show that the acetylcholine analogue carbamylcholine raises ventricular fibrillation threshold (VFT) and flattens the electrical restitution curve. The anti-fibrillatory action of carbamylcholine was prevented by the nicotinic receptor antagonist mecamylamine, inhibitors of neuronal nitric oxide synthase (nNOS) and soluble guanylyl cyclase (sGC), and can be mimicked by the nitric oxide (NO) donor sodium nitroprusside. Carbamylcholine increased NO metabolite content in the coronary effluent and this was prevented by mecamylamine. The anti-fibrillatory action of both carbamylcholine and sodium nitroprusside was ultimately dependent on muscarinic receptor stimulation as all effects were blocked by atropine. These data demonstrate a protective effect of carbamylcholine on VFT that depends upon both muscarinic and nicotinic receptor stimulation, where the generation of NO is likely to be via a neuronal nNOS-sGC dependent pathway. ABSTRACT: Implantable cardiac vagal nerve stimulators are a promising treatment for ventricular arrhythmia in patients with heart failure. Animal studies suggest the anti-fibrillatory effect may be nitric oxide (NO) dependent, although the exact site of action is controversial. We investigated whether a stable analogue of acetylcholine could raise ventricular fibrillation threshold (VFT), and whether this was dependent on NO generation and/or muscarinic/nicotinic receptor stimulation. VFT was determined in Langendorff perfused rat hearts by burst pacing until sustained VF was induced. Carbamylcholine (CCh, 200 nmol l(-1) , n = 9) significantly (P < 0.05) reduced heart rate from 292 ± 8 to 224 ± 6 b.p.m. Independent of this heart rate change, CCh caused a significant increase in VFT (control 1.5 ± 0.3 mA, CCh 2.4 ± 0.4 mA, wash 1.1 ± 0.2 mA) and flattened the restitution curve (n = 6) derived from optically mapped action potentials. The effect of CCh on VFT was abolished by a muscarinic (atropine, 0.1 µmol l(-1) , n = 6) or a nicotinic receptor antagonist (mecamylamine, 10 µmol l(-1) , n = 6). CCh significantly increased NOx content in coronary effluent (n = 8), but not in the presence of mecamylamine (n = 8). The neuronal nitric oxide synthase inhibitor AAAN (N-(4S)-4-amino-5-[aminoethyl]aminopentyl-N'-nitroguanidine; 10 µmol l(-1) , n = 6) or soluble guanylate cyclase (sGC) inhibitor ODQ (1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; 10 µmol l(-1) , n = 6) prevented the rise in VFT with CCh. The NO donor sodium nitrprusside (10 µmol l(-1) , n = 8) mimicked the action of CCh on VFT, an effect that was also blocked by atropine (n = 10). These data demonstrate a protective effect of CCh on VFT that depends upon both muscarinic and nicotinic receptor stimulation, where the generation of NO is likely to be via a neuronal nNOS/sGC-dependent pathway.

Human-based approaches to pharmacology and cardiology: an interdisciplinary and intersectorial workshop.

Both biomedical research and clinical practice rely on complex datasets for the physiological and genetic characterization of human hearts in health and disease. Given the complexity and variety of approaches and recordings, there is now growing recognition of the need to embed computational methods in cardiovascular medicine and science for analysis, integration and prediction. This paper describes a Workshop on Computational Cardiovascular Science that created an international, interdisciplinary and inter-sectorial forum to define the next steps for a human-based approach to disease supported by computational methodologies. The main ideas highlighted were (i) a shift towards human-based methodologies, spurred by advances in new in silico, in vivo, in vitro, and ex vivo techniques and the increasing acknowledgement of the limitations of animal models. (ii) Computational approaches complement, expand, bridge, and integrate in vitro, in vivo, and ex vivo experimental and clinical data and methods, and as such they are an integral part of human-based methodologies in pharmacology and medicine. (iii) The effective implementation of multi- and interdisciplinary approaches, teams, and training combining and integrating computational methods with experimental and clinical approaches across academia, industry, and healthcare settings is a priority. (iv) The human-based cross-disciplinary approach requires experts in specific methodologies and domains, who also have the capacity to communicate and collaborate across disciplines and cross-sector environments. (v) This new translational domain for human-based cardiology and pharmacology requires new partnerships supported financially and institutionally across sectors. Institutional, organizational, and social barriers must be identified, understood and overcome in each specific setting.

Macro-micro imaging of cardiac-neural circuits in co-cultures from normal and diseased hearts.

The autonomic nervous system plays an important role in the modulation of normal cardiac rhythm, but is also implicated in modulating the heart's susceptibility to re-entrant ventricular and atrial arrhythmias. The mechanisms by which the autonomic nervous system is pro-arrhythmic or anti-arrhythmic is multifaceted and varies for different types of arrhythmia and their cardiac substrates. Despite decades of research in this area, fundamental questions related to how neuron density and spatial organization modulate cardiac wave dynamics remain unanswered. These questions may be ill-posed in intact tissues where the activity of individual cells is often experimentally inaccessible. Development of simplified biological models that would allow us to better understand the influence of neural activation on cardiac activity can be beneficial. This Symposium Review summarizes the development of in vitro cardiomyocyte cell culture models of re-entrant activity, as well as challenges associated with extending these models to include the effects of neural activation.

A molecular signature of tissues with pacemaker activity in the heart and upper urinary tract involves coexpressed hyperpolarization-activated cation and T-type Ca2+ channels.

Renal pacemakers set the origin and frequency of the smooth muscle contractions that propel wastes from the kidney to the bladder. Although congenital defects impairing this peristalsis are a leading cause of pediatric renal failure, the mechanisms underlying renal pacemaker activity remain unknown. Using ratiometric optical mapping and video microscopy, we discovered that hyperpolarization-activated cation (HCN) channel block with the specific anatagonist ZD7288 (30 µm; IC50) abolished the pacemaker depolarizations that initiate murine upper urinary tract peristalsis. Optical mapping and immunohistochemistry indicate that pacemaker potentials are generated by cells expressing HCN isoform-3, and that HCN3(+) cells are coupled to definitive smooth muscle via gap junctions. Furthermore, we demonstrate that HCN3(+) cells coexpress T-type Ca(2+) (TTC) channels and that TTC channel inhibition with R(-)efonidipine or NNC55-0396 decreased contractile frequency in a dose-dependent manner. Collectively, these data demonstrate that HCN3(+)/TTC(+) cells are the pacemakers that set the origin and rate of upper urinary tract peristalsis. These results reveal a conserved mechanism controlling autorhythmicity in 2 distinct muscle types, as HCN and TTC channels also mediate cardiac pacemaker activity. Moreover, these findings have translational applications, including the development of novel diagnostics to detect fetal urinary tract motility defects prior to renal damage.-Hurtado, R., Bub, G., Herzlinger, D. A molecular signature of tissues with pacemaker activity in the heart and upper urinary tract involves coexpressed hyperpolarization-activated cation and T-type Ca(2+) channels.

Fast measurement of sarcomere length and cell orientation in Langendorff-perfused hearts using remote focusing microscopy.

RATIONALE: Sarcomere length (SL) is a key indicator of cardiac mechanical function, but current imaging technologies are limited in their ability to unambiguously measure and characterize SL at the cell level in intact, living tissue. OBJECTIVE: We developed a method for measuring SL and regional cell orientation using remote focusing microscopy, an emerging imaging modality that can capture light from arbitrary oblique planes within a sample. METHODS AND RESULTS: We present a protocol that unambiguously and quickly determines cell orientation from user-selected areas in a field of view by imaging 2 oblique planes that share a common major axis with the cell. We demonstrate the effectiveness of the technique in establishing single-cell SL in Langendorff-perfused hearts loaded with the membrane dye di-4-ANEPPS. CONCLUSIONS: Remote focusing microscopy can measure cell orientation in complex 2-photon data sets without capturing full z stacks. The technique allows rapid assessment of SL in healthy and diseased heart experimental preparations.

Mechanism of reentry induction by a 9-V battery in rabbit ventricles.

Although the application of a 9-V battery to the epicardial surface is a simple method of ventricular fibrillation induction, the fundamental mechanisms underlying this process remain unstudied. We used a combined experimental and modelling approach to understand how the interaction of direct current (DC) from a battery may induce reentrant activity within rabbit ventricles and its dependence on battery application timing and duration. A rabbit ventricular computational model was used to simulate 9-V battery stimulation for different durations at varying onset times during sinus rhythm. Corresponding high-resolution optical mapping measurements were conducted on rabbit hearts with DC stimuli applied via a relay system. DC application to diastolic tissue induced anodal and cathodal make excitations in both simulations and experiments. Subsequently, similar static epicardial virtual electrode patterns were formed that interacted with sinus beats but did not induce reentry. Upon battery release during diastole, break excitations caused single ectopics, similar to application, before sinus rhythm resumed. Reentry induction was possible for short battery applications when break excitations were slowed and forced to take convoluted pathways upon interaction with refractory tissue from prior make excitations or sinus beats. Short-lived reentrant activity could be induced for battery release shortly after a sinus beat for longer battery applications. In conclusion, the application of a 9-V battery to the epicardial surface induces reentry through a complex interaction of break excitations after battery release with prior induced make excitations or sinus beats.

Electrotonic suppression of early afterdepolarizations in the neonatal rat ventricular myocyte monolayer.

Pathologies that result in early afterdepolarizations (EADs) are a known trigger for tachyarrhythmias, but the conditions that cause surrounding tissue to conduct or suppress EADs are poorly understood. Here we introduce a cell culture model of EAD propagation consisting of monolayers of cultured neonatal rat ventricular myocytes treated with anthopleurin-A (AP-A). AP-A-treated monolayers display a cycle length dependent prolongation of action potential duration (245 ms untreated, vs. 610 ms at 1 Hz and 1200 ms at 0.5 Hz for AP-A-treated monolayers). In contrast, isolated single cells treated with AP-A develop prominent irregular oscillations with a frequency of 2.5 Hz, and a variable prolongation of the action potential duration of up to several seconds. To investigate whether electrotonic interactions between coupled cells modulates EAD formation, cell connectivity was reduced by RNA silencing gap junction Cx43. In contrast to well-connected monolayers, gap junction silenced monolayers display bradycardia-dependent plateau oscillations consistent with EADs. Further, simulations of a cell displaying EADs electrically connected to a cell with normal action potentials show a coupling strength-dependent suppression of EADs consistent with the experimental results. These results suggest that electrotonic effects may play a critical role in EAD-mediated arrhythmogenesis.

Temporal pixel multiplexing for simultaneous high-speed, high-resolution imaging.

We introduce an imaging modality that, by offsetting pixel-exposure times during capture of a single image frame, embeds temporal information in each frame. This allows simultaneous acquisition of full-resolution images at native detector frame rates and high-speed image sequences at reduced resolution, without increasing bandwidth requirements. We demonstrate this method using macroscopic and microscopic examples, including imaging calcium transients in heart cells at 250 Hz using a 10-Hz megapixel camera.

Simultaneous Widefield Voltage and Dye-Free Optical Mapping Quantifies Electromechanical Waves in Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

Coupled electromechanical waves define a heart's function in health and diseases. Optical mapping of electrical waves using fluorescent labels offers mechanistic insights into cardiac conduction abnormalities. Dye-free/label-free mapping of mechanical waves presents an attractive non-invasive alternative. In this study, we developed a simultaneous widefield voltage and interferometric dye-free optical imaging methodology that was used as follows: (1) to validate dye-free optical mapping for quantification of cardiac wave properties in human iPSC-cardiomyocytes (CMs); (2) to demonstrate low-cost optical mapping of electromechanical waves in hiPSC-CMs using recent near-infrared (NIR) voltage sensors and orders of magnitude cheaper miniature industrial CMOS cameras; (3) to uncover previously underexplored frequency- and space-varying parameters of cardiac electromechanical waves in hiPSC-CMs. We find similarity in the frequency-dependent responses of electrical (NIR fluorescence-imaged) and mechanical (dye-free-imaged) waves, with the latter being more sensitive to faster rates and showing steeper restitution and earlier appearance of wavefront tortuosity. During regular pacing, the dye-free-imaged conduction velocity and electrical wave velocity are correlated; both modalities are sensitive to pharmacological uncoupling and dependent on gap-junctional protein (connexins) determinants of wave propagation. We uncover the strong frequency dependence of the electromechanical delay (EMD) locally and globally in hiPSC-CMs on a rigid substrate. The presented framework and results offer new means to track the functional responses of hiPSC-CMs inexpensively and non-invasively for counteracting heart disease and aiding cardiotoxicity testing and drug development.

Predicting discrete-time bifurcations with deep learning.

Many natural and man-made systems are prone to critical transitions-abrupt and potentially devastating changes in dynamics. Deep learning classifiers can provide an early warning signal for critical transitions by learning generic features of bifurcations from large simulated training data sets. So far, classifiers have only been trained to predict continuous-time bifurcations, ignoring rich dynamics unique to discrete-time bifurcations. Here, we train a deep learning classifier to provide an early warning signal for the five local discrete-time bifurcations of codimension-one. We test the classifier on simulation data from discrete-time models used in physiology, economics and ecology, as well as experimental data of spontaneously beating chick-heart aggregates that undergo a period-doubling bifurcation. The classifier shows higher sensitivity and specificity than commonly used early warning signals under a wide range of noise intensities and rates of approach to the bifurcation. It also predicts the correct bifurcation in most cases, with particularly high accuracy for the period-doubling, Neimark-Sacker and fold bifurcations. Deep learning as a tool for bifurcation prediction is still in its nascence and has the potential to transform the way we monitor systems for critical transitions.

KHz-rate volumetric voltage imaging of the whole Zebrafish heart.

Fast volumetric imaging is essential for understanding the function of excitable tissues such as those found in the brain and heart. Measuring cardiac voltage transients in tissue volumes is challenging, especially at the high spatial and temporal resolutions needed to give insight to cardiac function. We introduce a new imaging modality based on simultaneous illumination of multiple planes in the tissue and parallel detection with multiple cameras, avoiding compromises inherent in any scanning approach. The system enables imaging of voltage transients in situ, allowing us, for the first time to our knowledge, to map voltage activity in the whole heart volume at KHz rates. The high spatiotemporal resolution of our method enabled the observation of novel dynamics of electrical propagation through the zebrafish atrioventricular canal.

The role of photon scattering in voltage-calcium fluorescent recordings of ventricular fibrillation.

Recent optical mapping studies of cardiac tissue suggest that membrane voltage (V(m)) and intracellular calcium concentrations (Ca) become dissociated during ventricular fibrillation (VF), generating a proarrhythmic substrate. However, experimental methods used in these studies may accentuate measured dissociation due to differences in fluorescent emission wavelengths of optical voltage/calcium (V(opt)/Ca(opt)) signals. Here, we simulate dual voltage-calcium optical mapping experiments using a monodomain-Luo-Rudy ventricular-tissue model coupled to a photon-diffusion model. Dissociation of both electrical, V(m)/Ca, and optical, V(opt)/Ca(opt), signals is quantified by calculating mutual information (MI) for VF and rapid pacing protocols. We find that photon scattering decreases MI of V(opt)/Ca(opt) signals by 23% compared to unscattered V(m)/Ca signals during VF. Scattering effects are amplified by increasing wavelength separation between fluorescent voltage/calcium signals and respective measurement-location misalignment. In contrast, photon scattering does not affect MI during rapid pacing, but high calcium dye affinity can decrease MI by attenuating alternans in Ca(opt) but not in V(opt). We conclude that some dissociation exists between voltage and calcium at the cellular level during VF, but MI differences are amplified by current optical mapping methods.