Pacing Frequency Research Articles

Temperature dependence of cardiac sarcoplasmic reticulum and sarcolemma in the ventricle of catfish (Clarias gariepinus)

The present study was undertaken to examine the relative contribution of the SR-Ca2+ release and sarcolemmal Ca2+ channels in developing the cardiac force at two different temperatures (20 and 30°C) in the catfish (Clarias gariepinus).The sarcolemmal Ca2+ contribution of activator Ca2+ was greater at a test temperature of 30°C as assessed by verapamil. Whereas the SR-Ca2+ contribution was higher at 20 and 30°C and a frequency rate of 0.2 and 0.4Hz as assessed by caffeine and adrenaline, respectively. Bradykinin potentiating factor (BPF7) which was isolated from jelly fish (Cassiopea andromeda) decreased the cardiac force developed at a frequency rate of 0.2Hz and a temperature of 20°C, whereas it increased the force developed at frequency rates of 0.2 and 0.4Hz at 30°C. These results indicate that BPF7 may act like verapamil in reducing the cardiac force through blocking the sarcolemmal Ca2+ channels at low temperature and like adrenaline in an increase of the cardiac force developed at warm temperature and the high frequency rate through stimulation of SR-Ca2+ activator. Therefore, this study indicates that the sarcolemmal Ca2+ influx and the SR-Ca2+ release contributors of activator Ca2+ for cardiac force development in the catfish heart were significantly greater at warm temperature and at the pacing frequency rates of 0.2 and 0.4Hz as assessed by verapamil, adrenaline, caffeine and BPF7. However, the relative contribution of the sarcolemmal Ca2+ influx in the development of cardiac force in the catfish heart was greater than that of SR-Ca2+ release.

A Multiscale Tridomain Model for Simulating Bioelectric Gastric Pacing.

Gastric motility disorders have been associated with abnormal slow wave electrical activity (gastric dysrhythmias). Gastric pacing is a potential therapy for gastric dysrhythmias; however, new pacing protocols are required that can effectively modulate motility patterns, while being power efficient. This study presents a novel comprehensive 3-D multiscale modeling framework of the human stomach, including anisotropic conduction, capable of evaluating pacing strategies. A high-resolution anatomically realistic mesh was generated from CT images taken from a human stomach. Principal conduction axes were calculated and embedded within this model based on a modified Laplace-Dirichlet rule-based algorithm. A continuum-based tridomain formulation was implemented and evaluated for performance and used to model the slow-wave propagation, which takes into account the two main cell types present in gastric musculature. Model parameters were found by matching predicted normal slow-wave activity to experimental observation and data. These simulation parameters were applied while modeling an external pacing event to entrain slow-wave patterns. The proposed formulation was found to be two times more efficient than a previous formulation for a normal slow-wave simulation. Convergence analysis showed that a mesh resolution of [Formula: see text] is required for an accurate solution process. The effect of different pacing frequencies on entrainment demonstrated that the pacing protocols are limited by the frequency of the native propagation and the refractory period of the cellular activity. The model is expected to become an important tool in studying pacing protocols for both efficiency and effectiveness.

Distinct Functional Roles of Cardiac Mitochondrial Subpopulations Revealed by a 3D Simulation Model

Experimental characterization of two cardiac mitochondrial subpopulations, namely, subsarcolemmal mitochondria (SSM) and interfibrillar mitochondria (IFM), has been hampered by technical difficulties, and an alternative approach is eagerly awaited. We previously developed a three-dimensional computational cardiomyocyte model that integrates electrophysiology, metabolism, and mechanics with subcellular structure. In this study, we further developed our model to include intracellular oxygen diffusion, and determined whether mitochondrial localization or intrinsic properties cause functional variations. For this purpose, we created two models: one with equal SSM and IFM properties and one with IFM having higher activity levels. Using these two models to compare the SSM and IFM responses of [Ca2+], tricarboxylic acid cycle activity, [NADH], and mitochondrial inner membrane potential to abrupt changes in pacing frequency (0.25–2 Hz), we found that the reported functional differences between these subpopulations appear to be mostly related to local [Ca2+] heterogeneity, and variations in intrinsic properties only serve to augment these differences. We also examined the effect of hypoxia on mitochondrial function. Under normoxic conditions, intracellular oxygen is much higher throughout the cell than the half-saturation concentration for oxidative phosphorylation. However, under limited oxygen supply, oxygen is mostly exhausted in SSM, leaving the core region in an anoxic condition. Reflecting this heterogeneous oxygen environment, the inner membrane potential continues to decrease in IFM, whereas it is maintained to nearly normal levels in SSM, thereby ensuring ATP supply to this region. Our simulation results provide clues to understanding the origin of functional variations in two cardiac mitochondrial subpopulations and their differential roles in maintaining cardiomyocyte function as a whole.

0179 : Specific down-regulation of cardiac connexin and pro-arrhythmic impulse propagation under electrical pacing

The rhythmic and coordinated propagation cardiac of the action potential is ensured by gap junction channels (GJCs) that are composed of connexin40 (Cx40), Cx43, Cx45 and Cx30.2, which present specific patterns of expression and electrical properties in different functionally specialized cardiac tissues. This study aims to determine the distinct contribution of Cxs in regulating the action potential propagation in mimicked healthy and diseased cardiac rates. The mouse atrial phenotypic HL-1 cell line that co-expresses Cx40, Cx43 and Cx45 was seeded on microelectrodes arrays to characterize the intrinsic electrical conduction and during pacing frequency from 2 to 30Hz. Specific SiRNA transfection of each Cx is used to determine the contribution of Cxs on regulating the conduction. We previously showed that the higher the frequency, the slower the conduction velocity (CV), up to a pro-arrhythmic frequency (10Hz). After downregulation of Cx40 (controlled by immunofluorescence), cells display a similar intrinsic coordinated activity to non-transfected cells (≈1Hz). However, the action potential propagates slower than non-transfected cells (≈60% at maximum downregulation of Cx40). Importantly, we observed that the arrhythmic conduction occurs at lower frequency after down-regulation of Cx40. These results indicate that Cx40 ensures a positive regulation of the action potential propagation under an increase of the cardiac rates, which suggests a specific GJCs make-up and electrical properties. Ongoing siRNA transfection for Cx43 and Cx45 will permit to distinguish the contribution of each Cxs in such regulation. Complementary biochemical characterization and patch clamp recordings on cell pairs will correlate the GJCs make-up and electrical properties to the changes of CV. This will improve our understanding on the specific role of each type of Cxs in regulating the impulse propagation in the healthy heart and their pro-arrhythmic dysfunction.

Phosphoregulation of Cardiac Inotropy via Myosin Binding Protein-C During Increased Pacing Frequency or β1-Adrenergic Stimulation.

Mammalian hearts exhibit positive inotropic responses to β-adrenergic stimulation as a consequence of protein kinase A-mediated phosphorylation or as a result of increased beat frequency (the Bowditch effect). Several membrane and myofibrillar proteins are phosphorylated under these conditions, but the relative contributions of these to increased contractility are not known. Phosphorylation of cardiac myosin-binding protein-C (cMyBP-C) by protein kinase A accelerates the kinetics of force development in permeabilized heart muscle, but its role in vivo is unknown. Such understanding is important because adrenergic responsiveness of the heart and the Bowditch effect are both depressed in heart failure. The roles of cMyBP-C phosphorylation were studied using mice in which either WT or nonphosphorylatable forms of cMyBP-C [ser273ala, ser282ala, ser302ala: cMyBP-C(t3SA)] were expressed at similar levels on a cMyBP-C null background. Force and [Ca(2+)]in measurements in isolated papillary muscles showed that the increased force and twitch kinetics because increased pacing or β1-adrenergic stimulation were nearly absent in cMyBP-C(t3SA) myocardium, even though [Ca(2+)]in transients under each condition were similar to WT. Biochemical measurements confirmed that protein kinase A phosphorylated ser273, ser282, and ser302 in WT cMyBP-C. In contrast, CaMKIIδ, which is activated by increased pacing, phosphorylated ser302 principally, ser282 to a lesser degree, and ser273 not at all. Phosphorylation of cMyBP-C increases the force and kinetics of twitches in living cardiac muscle. Further, cMyBP-C is a principal mediator of increased contractility observed with β-adrenergic stimulation or increased pacing because of protein kinase A and CaMKIIδ phosphorylations of cMyB-C.

The Power Output of Intact, Isolated Rat Cardiomyocytes

A method is presented to measure the power output of isolated intact cardiac myocytes. The mycoytes were attached to a very sensitive and fast force transducer on one side (OptiForce form Ionoptix) and a direct drive piezo-translator on the other side to stretch or shorten the myocyte. This allowed feedback based force control. Force clamps were used to impose a minimum ‘pre-load’ level and maximum ‘after-load’ level between which the myocyte was allowed to operate, mimicking the aortic and atrial pressure respectively. As long as the myocyte reaches the after load level in systole, the loaded shortening results in a force versus length curve that forms a work loop, analogous to the pressure volume curve in the whole heart. Within the measured range (0-1.5 µNewton pre-load, corresponding to sarcomere lengths of 1.86±0.07µm through 1.96±0.08µm) the end-systolic and end-diastolic force-length relations were linear. Measuring the area within each work loop provided the external work performed by the mycoyte for each contraction. A power curve was created by varying the after load for a given pre-load and multiplying the work per contraction with the pacing frequency. For rat myocytes at 37°C the peak power output increased linearly with pre-load within the range measured, and at a pre-load of 1.5 µNewton varied between 25 and 90 pJ.s-1. The large variation was mostly due to differences in cell size. Maximum power output was achieved at pacing frequencies between 6-8 Hz.

Spatial Symmetry Breaking Determines Spiral Wave Rotation Direction in Simplified Cardiac Systems

Chirality (“rotation direction”) represents a fundamental property of spiral waves, self-sustaining vortices of electrical activity thought to underlie a number of re-entrant cardiac arrhythmias. However, the key regulators of spiral wave chirality remain unclear. We introduced inexcitable obstacles into cardiac monolayers composed of embryonic cardiac chick cells, which gave rise to clockwise-rotating, counterclockwise-rotating, and pairs of spiral waves. Numerical simulations of the experiments showed that the precise location of the obstacle in combination with the pacing frequency determined spiral wave chirality [1]. In particular, we observed that obstacle position controls wavefront curvature, which makes different regions of the substrate more susceptible to propagation block and spiral wave formation. To quantify this observation, we found that instabilities predicted by curves relating the action potential duration and the pacing frequency at different spatial locations predict sites of wave break initiation, and, hence, spiral wave chirality. [1] T. Quail, A. Shrier, L. Glass, Spatial Symmetry Breaking Determines Spiral Wave Chirality, Phys. Rev. Lett., In Press, (2014).

Formula for Equivalent Impulsive Force to Predict Vibrational Response of High-frequency Staircases

High-frequency staircases are widely used nowadays to meet aesthetics and functionality needed in modern architecture. Unfortunately, no design guide is available in domestic practice to predict response or evaluate the vibration performance of high-frequency staircases. SCI-P354 published by the Steel Construction Institute of UK provides the formula for effective impulsive force. However, this formula was shown to overestimate the response of high-frequency staircases excited by fast ascending and descending over 2.2Hz pace frequency because it was developed based on the walking test in a slow pace frequency. This study proposes a semi-analytical formula to predict the response of stiff staircases based on analytical and experimental studies of response acceleration for various walking frequencies covering 1.4~4.5Hz.

Development of Ca Alternans in Atrial Myocytes is Modulated by Action Potential Morphology

Cardiac alternans is defined as cyclic, beat-to-beat variations in contraction force, action potential duration (APD) and cytosolic Ca transient (CaT) amplitude at constant stimulation rate.The bidirectional coupling of beat-to-beat regulation of cytosolic [Ca]i and membrane voltage (Vm) is a key causative factor for the development of alternans. The majority of previous studies have emphasized the role of [Ca]i→Vm coupling and disturbances in [Ca]i regulation as the primary cause of APD alternans. In this study we investigated how alternation in AP morphology feeds back on intracellular Ca handling (Vm→[Ca]i coupling) in single rabbit atrial myocytes. From AP recordings during pacing induced simultaneous CaT and APD alternans two AP waveforms were selected that represent a typical short and long AP of APD alternans. These AP waveforms were used as command voltages in subsequent voltage clamp experiments (AP voltage clamp). At pacing frequencies below alternans induction, using a series of short or long APs voltage clamp pulses, APD affected CaT amplitude where short APs caused slightly smaller amplitude CaTs. At higher frequencies CaT alternans could be induced with AP voltage clamp protocols in the absence of APD alternans, however the pacing frequency threshold where CaT alternans could be induced decreased with increasing APD, i.e. a long APD was capable of triggering CaT alternans at lower frequencies than a short AP. When cells were stimulated with an APD alternans protocol (APs of alternating duration as command voltage) the CaT alternans threshold was significantly lower. The data identify a contribution of Vm→[Ca]i coupling to the development and severity of CaT alternans and demonstrate that Vm→[Ca]i coupling affects Ca handling through changes in diastolic SR Ca load and voltage-dependent regulation of L-type Ca channels and Na/Ca exchange.

Excitation-Metabolism Coupling in Mouse Heart

A beating heart must balance energy consumption with production. This balance requires cellular ATP production to respond dynamically to changes in cardiac output and energy demand. While it is well understood that mitochondrial oxidative phosphorylation serves as the primary pathway for ATP production in heart, the mechanisms that govern the regulation of mitochondrial metabolism remain unclear. Calcium (Ca2+), specifically Ca2+ in the mitochondrial matrix ([Ca2+]m), has been implicated as a likely signaling pathway for this regulation. [Ca2+]m has been suggested to regulate nearly every stage of mitochondrial metabolism including the activity of the tricarboxylic acid cycle (TCA) via Ca2+-sensitive dehydrogenases (DHOs), critical proteins in the electron transport chain (ETC), and even the F1/F0 ATP synthase itself. Here we expand our local control mathematical model of excitation-contraction coupling in mouse heart to investigate excitation-metabolism coupling. The model features mechanistic initiation and termination of Ca2+ sparks and cytosolic Ca2+ ([Ca2+]i) transients in a system that maintains sarcoplasmic reticulum (SR) Ca2+ pump/leak balance. Mitochondrial Ca2+ uptake and export is simulated using experimentally constrained formulations of the mitochondrial Ca2+ uniporter (MCU) and the mitochondrial sodium/Ca2+ exchanger (NCLX). The model employs key mechanisms of ATP consumption (i.e. SR Ca2+ ATPase and myosin ATPase) and buffering (phosphocreatine) as well as a Ca2+ dependent model for mitochondrial ATP generation. We investigated mitochondrial Ca2+ dynamics at physiological pacing frequencies for mouse heart and show that changes in ATP consumption can be translated to increased energy production through [Ca2+]m signals. Our model provides new insights into mitochondrial Ca2+ dynamics and how these [Ca2+]m signals may function to preserve energy homeostasis in the face of increased demand.

Quantification of human-structure interaction

In lightweight structural systems there is increasing evidence that the presence of humans influences the dynamics characteristics of the system. In the past, most effort on determining the footfall-induced vertical force to the walking surface has been conducted using rigid or non-flexible surfaces such as treadmills. However, should the walking surface be vibrating, the characteristics of human walking could change to maximize comfort. This interaction between the structure and human may account for the discrepancy between the levels of vibration predicted by theory and those observed in practice. Indeed, many design rules can be seen to be conservative, perhaps partly because knowledge of this human-structure interaction is limited. This work aims to address this problem by quantifying the magnitude of human-structure interaction through a comprehensive experimental programme. Novel experimental techniques are used to measure the human-imparted force on the walking surface. Both rigid and flexible (vibrating) surfaces are used, and we measure the imparted vibration response on a lively footbridge (the Warwick Bridge) which acts as the flexible surface. A range of test subjects is considered, walking at a range of pacing frequencies. Comparison is made between a notional vibration response from the footfall force imparted to the rigid surface and the actual vibration response caused by the footfall force imparted to the flexible surface. Key aspects of the experimental regime are also explained. Finally, some comparisons are made using footfall force models from the literature. It is concluded that human-structure interaction is a key phenomenon that should be taken into account in the design and assessment of vibration-sensitive structures.

Saxagliptin Preserves Cardiomyocyte Function and Morphology in Aortic-Banded Mini-Swine

Impaired cGMP-PKG signaling may contribute to cardiomyocyte remodeling and cardiac dysfunction in heart failure with preserved ejection fraction (HFpEF) patients. The purpose of this study was to assess cardiomyocyte function and morphology in aortic-banded mini-swine displaying a HFpEF phenotype following manipulation of cGMP signaling via two mechanisms: 1) driving cGMP synthesis with the DPP4 inhibitor saxagliptin; and 2) preventing cGMP catabolism via the PDE5 inhibitor tadalafil. We hypothesized that preserving cGMP-PKG signaling would prevent cardiomyocyte dysfunction and remodeling. Contractile function was measured in enzymatically isolated cardiomyocytes electrically stimulated at three frequencies (0.25, 0.5, 1.0 Hz)from four groups: control (CON), aortic-banded (AB), AB saxagliptin-treated (AB-SAX), and AB tadalafil-treated (AB-TAD). Increased cGMP activity was only observed in AB-TAD animals, however, neither drug treatment increased PKG activity. A significant pacing-induced decrease in diastolic sarcomere length was observed in the AB and AB-TAD groups compared to CON. This finding, indicative of impaired diastolic relaxation, was prevented by saxagliptin treatment. Shortening amplitude decreased in AB cardiomyocytes with increasing pacing frequency, revealing a negative shortening-frequency relationship that was attenuated by both drug treatments. Length:width ratio was decreased in AB and AB-TAD animals with commensurate decreases in left ventricular (LV) end diastolic and end systolic volumes. Normal cardiomyocyte size and LV volumes were preserved in the AB-SAX group. Interestingly, all AB groups exhibited similar gross hypertrophic remodeling (heart weight:body weight ratio) despite differences in cardiomyocyte morphology. In conclusion, saxagliptin appears superior for preserving normal cardiomyocyte morphology and overall function versus tadalafil, independent of changes in cGMP-PKG activity.

Species-dependent adaptation of the cardiac Na+/K+ pump kinetics to the intracellular Na+ concentration.

The Na(+)/K(+) ATPase (NKA) plays a critical role in maintaining ionic homeostasis and dynamic function in cardiac myocytes, within both the in vivo cell and in silico models. Physiological conditions differ significantly between mammalian species. However, most existing formulations of NKA used to simulate cardiac function in computational models are derived from a broad range of experimental sources spanning many animal species. The resultant inability of these models to discern species-specific features is a significant obstacle to achieving a detailed quantitative and comparative understanding of physiological behaviour in different biological contexts. Here we present a framework for characterising the steady-state NKA current using a biophysical mechanistic model specifically designed to provide a mechanistic explanation of the NKA flux supported by self-consistent species-specific data. We thus compared NKA kinetics specific to guinea- pig and rat ventricular myocytes. We observe that the apparent binding affinity for sodium in the rat is significantly lower, whereas the overall pump cycle rate is doubled, in comparison to the guinea pig. This sensitivity of NKA to its regulatory substrates compensates for the differences in Na(+) concentrations between the cell types. NKA is thereby maintained within its dynamic range over a wide range of pacing frequencies in these two species, despite significant disparities in sodium concentration. Hence, by replacing a conventional generic NKA model with our rat-specific NKA formula into a whole-cell simulation, we have, for the first time, been able to accurately reproduce the action potential duration and the steady-state sodium concentration as functions of pacing frequency.

Abstract 12646: Oxidation-Dependent Phosphomimetic Effect of a Human Heart Failure Mutation of Phospholamban

Rationale: A naturally-occurring, missense Arg9-to-Cys (R9C) mutation of phospholamban (PLB) triggers cardiomyopathy and premature death in humans. However, the fundamental molecular mechanism underlying the cardiotoxic role of R9C-PLB in sarco/endoplasmic reticulum Ca 2+ -ATPase (SERCA) regulation and cardiomyocyte Ca 2+ handling is not clear. Objective: The goal of this study was to investigate the acute physiological consequences of R9C-PLB mutation on cardiomyocyte Ca 2+ kinetics and contractility and identify the molecular mechanism underlying R9C pathology. Methods and Results: We measured the physiological consequences of R9C-PLB mutation on Ca 2+ transients and sarcomere shortening in adult cardiomyocytes at increasing pacing frequencies. In contrast to studies of chronic R9C-PLB expression in transgenic mice, we found that acute expression of R9C-PLB exerts a positively inotropic and lusitropic effect in cardiomyocytes. Importantly, R9C-PLB exhibited blunted sensitivity to frequency potentiation and β-adrenergic stimulation, two major physiological mechanisms for the regulation of cardiac performance. To identify the molecular mechanism of R9C pathology, we fused fluorescent protein tags to PLB and SERCA, and compared the effect of R9C and pentamer-destabilizing mutation (SSS) on PLB oligomerization and PLB-SERCA interaction. Fluorescence resonance energy transfer (FRET) measurements in live cells revealed that R9C exhibited an increased affinity of PLB oligomerization, and a decreased binding affinity to SERCA due to an oxidative modification which mimics phosphorylation. Real-time FRET analysis in cardiomyocytes revealed that R9C-PLB exhibits enhanced sensitivity to oxidative stress, which is a prevailing condition in heart failure. Conclusions: We conclude that the primary mechanism of R9C pathology is a phosphomimetic effect of PLB cysteine oxidation, manifested as increased oligomerization and a change in the structure of the PLB-SERCA regulatory complex.

Blood pressure response to renal nerve stimulation in patients undergoing renal denervation: a feasibility study.

During renal sympathetic denervation (RDN), no mapping of renal nerves is performed and there is no clear end point of RDN. We hypothesized high-frequency renal nerve stimulation (RNS) may increase blood pressure (BP), and this increase is significantly blunted after RDN. The aim of this study was to determine the feasibility of RNS in patients undergoing RDN. Eight patients with resistant hypertension undergoing RDN were included. A quadripolar catheter was positioned at four different sites in either renal artery. RNS was performed during 1 min with a pacing frequency of 20 Hz. After all patients successfully underwent RDN, RNS was repeated at the site of maximum BP response before RDN in either renal artery. Mean age was 66 years. During RNS, BP increased significantly from 108/55 to 132/68 mm Hg (P < 0.001). After RDN, systolic BP response at the site of maximum response to RNS was significantly blunted (+43.1 vs +9.3 mm Hg, P = 0.002). In three patients, a systolic BP increase >10 mm Hg was observed after RDN. In conclusion, RNS resulted in an acute temporary BP increase. This response was significantly blunted after RDN. RNS may potentially serve as an end point for RDN.

Spatial symmetry breaking determines spiral wave chirality.

Chirality represents a fundamental property of spiral waves. Introducing obstacles into cardiac monolayers leads to the initiation of clockwise-rotating, counterclockwise-rotating, and pairs of spiral waves. Simulations show that the precise location of the obstacle and the pacing frequency determine spiral wave chirality. Instabilities predicted by curves relating the action potential duration and the pacing frequency at different spatial locations predict sites of wave break initiation and, hence, spiral wave chirality.

Electrophysiological properties under heart failure conditions in a human ventricular cell: a modeling study.

Heart failure (HF) is one of the major diseases across the world. During HF the electrophysiology of the failing heart is remodeled, which renders the heart more susceptible to ventricular arrhythmias. In this study, we quantitatively analyze the effects of electrophysiological remodeling of the major currents of human ventricular myocytes on the dynamics of the failing heart. We develop a HF model using a modified version of a recently published model of the human ventricular action potential, the O'Hara-Virag-Varro-Rudy (OVVR) model. The proposed HF model incorporates recently available HF clinical data. It can reproduce most of the action potential (AP) properties of failing myocytes, including action potential duration (APD), amplitude (APA), notch (APN), plateau (APP), resting membrane potential (RMP), and maximum upstroke velocity (dV/dtmax). In addition, the model reproduces the behavior of the [Na+], concentration and [Ca(2)+]i dynamics. Moreover, the HF model exhibits alternans with a fast pacing frequency and can induce early afterdepolarizations (EADs). Additionally, blocking the late sodium current shortens the APD and suppresses EADs, in agreement with experimental findings. The dynamics of the proposed model are assessed through investigating the rate dependence of the AP and the dynamics of the major currents. The steady-state (S-S) and S1-S2 restitution curves along with accommodation to an abrupt change in cycle length were evaluated. Our study should help to elucidate the roles of alterations in electrophysiological properties during HF. Also, this HF cellular model could be used to study HF in a realistic geometry and could be embedded into a model of HF electromechanics to investigate electrical and mechanical properties simultaneously during HF.

Experimental identification of the lateral human–structure interaction mechanism and assessment of the inverted-pendulum biomechanical model

Within the context of crowd-induced lateral bridge vibration, human–structure interaction (HSI) is a widely studied phenomenon. Central to this study is the self-excited component of the ground reaction force (GRF). This force harmonic, induced by a walking pedestrian, resonates with lateral deck motion, irrespective of the pedestrian׳s pacing frequency. Its presence can lead to positive feedback between pedestrian GRFs and structural motion. Characterisation of the self-excited force as equivalent structural mass and damping has greatly improved the understanding of HSI and its role in developing lateral dynamic instability. However, despite this evolving understanding, a key question has remained unanswered; what are the features of a pedestrian׳s balance response to base motion that gives rise to the self-excited force? The majority of the literature has focussed on the effects of HSI with the underlying mechanism receiving comparatively little attention. This paper presents data from experimental testing in which 10 subjects walked individually on a laterally oscillating treadmill. Lateral deck motion as well as the GRFs imposed by the subject was recorded. Three-dimensional motion capture equipment was used to track the position of visual markers mounted on the subject. Thus whole body response to base motion was captured in addition to the GRFs generated. The data presented herein supports the authors’ previous findings that the self-excited force is a frequency sideband harmonic resulting from amplitude modulation of the lateral GRF. The gait behaviour responsible for this amplitude modulation is a periodic modulation of stride width in response to a sinusoidally varying inertia force induced by deck motion. In a separate analysis the validity of the passive inverted pendulum model, stabilised by active control of support placement was confirmed. This was established through comparison of simulated and observed frontal plane CoM motion. Despite the relative simplicity of this biomechanical model, remarkable agreement was observed.

Behavioral and physiological responses in felids to exhibit construction.

Despite the growing body of literature examining the welfare of zoo-housed animals, little standardized work has been published on the effect of construction and environmental disruption on the physiology and behavior of affected animals. When Lincoln Park Zoo (Chicago, IL), embarked on a renovation project for its Kovler Lion House, the opportunity was taken to perform a scientific study of behavioral and physiological markers in the resident felids to determine the effect of construction and environmental disruption. Fecal samples and behavioral observations were collected on four felid species (five individuals) before, during, and after the period of construction. As a group, the average z-score for fecal glucocorticoid metabolite concentration increased during construction relative to baseline. Levels remained elevated after construction, but trended toward baseline. All individuals demonstrated a significant decrease in the frequency of pacing and time spent visible during construction. Overall activity levels also showed a significant decrease relative to baseline measures. As zoological institutions continue to recognize the importance of habitat design, construction and renovation become inevitable. It is important to be aware of the potential consequences this can have on animals in the vicinity and to work toward minimizing negative effects. One recommendation is the availability of ample retreat and hiding space for felids during disruption to their environment.

P778Physiological and pharmacological characterization of human engineered heart tissue

Objective: Human induced pluripotent stem cell (iPS cell)-derived cardiomyocytes represent a valuable tool in cardiovascular research of tremendous potential. However, these cells are still immature and characterized by poor sarcomeric organization and cellular orientation in 2D cell culture. Therefore, the measurement of contractile force, the most important and best understood function of cardiomyocytes in vivo, is not established for these cells. This study describes the generation of three-dimensional, strip-format, force generating engineered heart tissues (EHTs) from human iPS cell-derived cardiomyocytes and presents a characterization of physiological and pharmacological parameters based on force development. Methods and results: Cardiomyocyte differentiation of human induced pluripotent cells was achieved by a growth factor-based three stage protocol. Strip-format EHTs were generated from dissociated cardiomyocytes in fibrin matrix between flexible silicone posts. Within two weeks after casting coherently beating human EHTs were formed and EHTs displayed a regular beating pattern for several weeks. Histological analysis revealed a high degree of sarcomeric organization and alignment of cardiomyocytes in EHTs. Functional analysis was performed by measuring force response to calcium concentration, pre-load, pacing frequency, beta-adrenergic and muscarinic agonists, modulators of sodium, calcium and potassium channels and revealed concentration-dependent effects. Comparison with native human heart tissue suggests an overall high level of similarity and minor differences. Conclusions: This study demonstrates feasibility to characterize human iPS cell-derived cardiomyocytes in EHTs by measuring contractile force. The analysis suggests high levels of similarity between EHTs and native human heart tissue. Human EHTs are a promising platform for automated toxicology screens in future drug development and for in vitro experiments on human cardiomyocytes in general.