Discovery and optimization of 5-fluoro-4,6-dialkoxypyrimidine GPR119 agonists

A series of 5-fluoro-4,6-dialkoxypyrimidine GPR119 modulators were discovered and optimized for in vitro agonist activity. A lead molecule was identified that has improved agonist efficacy relative to our clinical compound (APD597) and possesses reduced CYP2C9 inhibitory potential. This optimized lead was found to be efficacious in rodent models of glucose control both alone and in combination with a Dipeptidyl peptidase-4 (DPP-4) inhibitor.     

Effect of simulated I(to) on guinea pig and canine ventricular action potential morphology

The transient outward current (I(to)) is a major repolarizing current in the heart. Marked reduction of I(to) density occurs in heart failure and is accompanied by significant action potential duration (APD) prolongation. To understand the species-dependent role of I(to) in regulating the ventricular action potential morphology and duration, we introduced simulated I(to) conductance in guinea pig and canine endocardial ventricular myocytes using the dynamic clamp technique and perforated patch-clamp recordings. The effects of simulated I(to) in both types of cells were complex and biphasic, separated by a clear density threshold of approximately 40 pA/pF. Below this threshold, simulated I(to) resulted in a distinct phase 1 notch and had little effect on or moderately prolonged the APD. I(to) above the threshold resulted in all-or-none repolarization and precipitously reduced the APD. Qualitatively, these results agreed with our previous studies in canine ventricular cells using whole cell recordings. We conclude that 1) contrary to previous gene transfer studies involving the Kv4.3 current, the response of guinea pig ventricular myocytes to a fully inactivating I(to) is similar to that of canine ventricular cells and 2) in animals such as dogs that have a broad cardiac action potential, I(to) does not play a major role in setting the APD.     

蜜蜂表皮蛋白apidermin (apd)基因家族3个新成员的特性鉴定及昆虫APD家族序列特征的分析

Apidermin (APD)蛋白家族是一个新的昆虫结构性表皮蛋白家族。本研究结合生物信息学和RT-PCR扩增, 对意大利蜜蜂Apis mellifera ligustica（简称“意蜂”）的apd-1-like, apd-3-like和中华蜜蜂Apis cerena cerena（简称“中蜂”）的apd-2 等3个新的apd基因的结构特征和表达进行了分析, 并分析了昆虫APD蛋白家族的序列特征。结果显示, 在西方蜜蜂Apis mellifera（简称“西蜂”）中, apd基因家族的6个成员串联排列在基因组序列第4号连锁群上, 它们在A. m. ligustica雄蜂头部中的转录水平差异明显, 且其启动子序列所含顺式元件也不同。中蜂apd-2和意蜂apd-1-like都含有3个外显子和2个内含子, 而意蜂apd-3-like则由4个外显子和3个内含子组成。蛋白序列分析结果显示, 目前已知的10条APD蛋白序列N末端均具有相似的信号肽序列, 其成熟蛋白分子量为6.0~37.0 kD, pI为6.2~10.8。其中西蜂的APD1-3、APD-like和东方蜜蜂Apis cerena的APD-2等5条较短的多肽中疏水氨基酸残基达52%~67%, 且Ala含量最为丰富（占25%~34%）; 而丽蝇蛹集金小蜂Nasonia vitripennis的APD 1-3和西蜂APD-1-like, APD-3-like等另外5条APD多肽富含Gly（21%~30%）, 其序列中疏水氨基酸残基含量为35%~41%。多肽序列多重比对和系统进化分析结果显示, APD家族可划分为2个亚家族。亚家族Ⅰ含有西蜂APD 1-3和东方蜜蜂APD-2等4条较短的多肽序列, 其N末端为一个长33 aa的保守基序; 亚家族Ⅱ由另外6条相对较长的多肽序列组成, 其N末端保守基序长50 aa, C末端保守基序长16 aa。本文所描述的APD蛋白家族序列特征有助于以后从其他昆虫中鉴定新的apd基因。     

Mechanisms and determinants of ultralong action potential duration and slow rate-dependence in cardiac myocytes

In normal cardiac myocytes, the action potential duration (APD) is several hundred milliseconds. However, experimental studies showed that under certain conditions, APD could be excessively long (or ultralong), up to several seconds. Unlike the normal APD, the ultralong APD increases sensitively with pacing cycle length even when the pacing rate is very slow, exhibiting a sensitive slow rate-dependence. In addition, these long action potentials may or may not exhibit early afterdepolarizations (EADs). Although these phenomena are well known, the underlying mechanisms and ionic determinants remain incompletely understood. In this study, computer simulations were performed with a simplified action potential model. Modifications to the L-type calcium current (I(Ca,L)) kinetics and the activation time constant of the delayed rectifier K current were used to investigate their effects on APD. We show that: 1) the ultralong APD and its sensitive slow rate-dependence are determined by the steady-state window and pedestal I(Ca,L) currents and the activation speed and the recovery of the delayed rectifier K current; 2) whether an ultralong action potential exhibits EADs or not depends on the kinetics of I(Ca,L); 3) increasing inward currents elevates the plateau voltage, which in general prolongs APD, however, this can also shorten APD when the APD is already ultralong under certain conditions; and 4) APD alternans occurs at slow pacing rates due to the sensitive slow rate-dependence and the ionic determinants are different from the ones causing APD alternans at fast heart rates.     

Short-term cardiac memory and mother rotor fibrillation

Short-term cardiac memory refers to the effects of pacing history on action potential duration (APD). Although the ionic mechanisms for short-term memory occurring over many heartbeats (also called APD accommodation) are poorly understood, they may have important effects on reentry and fibrillation. To explore this issue, we incorporated a generic memory current into the Phase I Luo and Rudy action potential model, which lacks short-term memory. The properties of this current were matched to simulate quantitatively human ventricular monophasic action potential accommodation. We show that, theoretically, short-term memory can resolve the paradox of how mother rotor fibrillation is initiated in heterogeneous tissue by physiological pacing. In simulated heterogeneous two-dimensional tissue and three-dimensional ventricles containing an inward rectifier K(+) current gradient, short-term memory could spontaneously convert multiple wavelet fibrillation to mother rotor fibrillation or to a mixture of both fibrillation types. This was due to progressive acceleration and stabilization of rotors as accumulation of memory shortened APD and flattened APD restitution slope nonuniformly throughout the tissue.     

铬对大鼠心电图及心肌细胞的电生理影响

应用心电图及细胞内微电极技术观察铬对心肌电生理的影响。大鼠腹腔内注射铬,９周后心电图显示各剂量组ＱＴ间期均缩短,细胞内微电极检查显示动作电位时程（ＡＰＤ５０、ＡＰＤ９０）于２周后随剂量增加而逐渐缩短,０．４ｍｇ组显著缩短,９周后各剂量组ＡＰＤ５０、ＡＰＤ９０均缩短。心率、静息电位（ＲＰ）与动作电位（ＡＰＡ）幅度及动作电位最大上升速率（Ｖｍａｘ）无变化。铬影响了心肌复极引起ＱＴ间期缩短,而ＡＰＤ５０、ＡＰＤ９０缩短可能是铬影响了钙内流及钾外流的结果。     

Contrasting effects of ischemia on the kinetics of membrane voltage and intracellular calcium transient underlie electrical alternans

Repolarization alternans has been considered a strong marker of electrical instability. The objective of this study was to investigate the hypothesis that ischemia-induced contrasting effects on the kinetics of membrane voltage and intracellular calcium transient (Ca(i)T) can explain the vulnerability of the ischemic heart to repolarization alternans. Ischemia-induced changes in action potential (AP) and Ca(i)T resulting in alternans were investigated in perfused Langendorff guinea pig hearts subjected to 10-15 min of global no-flow ischemia followed by 10-15 min of reperfusion. The heart was stained with 100 microl of rhod-2 AM and 25 microl of RH-237, and AP and Ca(i)T were simultaneously recorded with an optical mapping system of two 16 x 16 photodiode arrays. Ischemia was associated with shortening of AP duration (D) but delayed upstroke, broadening of peak, and slowed decay of Ca(i)T resulting in a significant increase of Ca(i)T-D. The changes in APD were spatially heterogeneous in contrast to a more spatially homogeneous lengthening of Ca(i)T-D. Ca(i)T alternans could be consistently induced with the introduction of a shorter cycle when the upstroke of the AP occurred before complete relaxation of the previous Ca(i)T and generated a reduced Ca(i)T. However, alternans of Ca(i)T was not necessarily associated with alternans of APD, and this was correlated with the degree of spatially heterogeneous shortening of APD. Sites with less shortening of APD developed alternans of both Ca(i)T and APD, whereas sites with greater shortening of APD could develop a similar degree of Ca(i)T alternans but slight or no APD alternans. This resulted in significant spatial dispersion of APD. The study shows that the contrasting effects of ischemia on the duration of AP and Ca(i)T and, in particular, on their spatial distribution explain the vulnerability of ischemic heart to alternans and the increased dispersion of repolarization during alternans.     

Tetrahydronaphthalene-derived amino alcohols and amino ketones as potent and selective inhibitors of the delayed rectifier potassium current IKs

Class III anti-arrhythmic drugs (e.g., dofetilide) prolong cardiac action potential duration (APD) by blocking the fast component of the delayed rectifier potassium current (I(Kr)). The block of I(Kr) can result in life threatening ventricular arrhythmias (i.e., torsades de pointes). Unlike I(Kr), the role of the slow component of the delayed rectifier potassium current (I(Ks)) becomes significant only at faster heart rate. Therefore selective blockers of I(Ks) could prolong APD with a reduced propensity to cause pro-arrhythmic side effects. This report describes structure-activity relationships (SARs) of a series of I(Ks) inhibitors derived from 6-alkoxytetralones with good in vitro activity (IC(50) > or =30 nM) and up to 40-fold I(Ks)/I(Kr) selectivity.     

Role of maximum rate of depolarization in predicting action potential duration during ventricular fibrillation

During ventricular fibrillation (VF) only 39% of the variation in action potential duration (APD) is accounted for by the previous diastolic interval [DI((n-1))], i.e., restitution, and the previous APD [APD((n-1))], i.e., memory. We tested the hypothesis that a characteristic of the AP upstroke, the maximum rate of depolarization (V(max)), also helps account for its APD. A floating microelectrode was used to make transmembrane recordings at 16,000 samples/s from the anterior left ventricular wall during four 20-s episodes of VF in each of six pigs. V(max), time from V(max) to 60% repolarization (APD(60)), and DI were calculated throughout all episodes. Stepwise linear regression was used to determine how well each APD(60) (APD(60n)) was predicted by V(max) of that AP, the four previous DIs (n-1, n - 2, n - 3, n - 4), and the three previous APD(60)s (n-1, n - 2, n - 3). V(max) entered in the regression equation significantly more often (86% of VF episodes) than either APD((n-1)) (47% of episodes) or DI((n-1)) (58% of episodes). When these three variables entered first or second, their coefficients were almost always positive, consistent with a longer APD associated with 1) a larger V(max), 2) a longer APD((n-1)), and 3) a longer DI((n-1)). R(2) of the regression for all entered variables was 0.51 +/- 0.01 (mean +/- SD). During the first 20 s of VF in swine, V(max) is a more important determinant of APD than the previous DI (restitution) or the previous APD (memory). All variables together account for only one-half of APD variation during VF.     

Post-rest adaptation of electrical and mechanical activity in the isolated guinea-pig papillary muscle

In guinea-pig papillary muscle the time course of the changes in contractile force and action potential duration (APD) were studied after periods of rest of variable duration. After a long period of rest, the force of contraction adapted to pre-rest control values in a monophasic manner whereas the time-course of the APD was clearly biphasic. The post-rest adaptation of the APD could be described mathematically by a simple model, which considers the action potential duration during steady state as the sum of a resting value (APDR) plus a lengthening effect of activation (LEA) minus a shortening effect of activation (SEA). LEA and SEA are assumed to occur immediately, with each excitation and to decay continuously. During repetitive stimulation, both effects will accumulate. Using the constants found for the post-rest adaptation of the APD, the steady-state frequency-dependence of the APD could also be described with this model.     

Targeting the late component of the cardiac L-type Ca2+ current to suppress early afterdepolarizations

Early afterdepolarizations (EADs) associated with prolongation of the cardiac action potential (AP) can create heterogeneity of repolarization and premature extrasystoles, triggering focal and reentrant arrhythmias. Because the L-type Ca²⁺ current (I_Ca,L) plays a key role in both AP prolongation and EAD formation, L-type Ca²⁺ channels (LTCCs) represent a promising therapeutic target to normalize AP duration (APD) and suppress EADs and their arrhythmogenic consequences. We used the dynamic-clamp technique to systematically explore how the biophysical properties of LTCCs could be modified to normalize APD and suppress EADs without impairing excitation–contraction coupling. Isolated rabbit ventricular myocytes were first exposed to H₂O₂ or moderate hypokalemia to induce EADs, after which their endogenous I_Ca,L was replaced by a virtual I_Ca,L with tunable parameters, in dynamic-clamp mode. We probed the sensitivity of EADs to changes in the (a) amplitude of the noninactivating pedestal current; (b) slope of voltage-dependent activation; (c) slope of voltage-dependent inactivation; (d) time constant of voltage-dependent activation; and (e) time constant of voltage-dependent inactivation. We found that reducing the amplitude of the noninactivating pedestal component of I_Ca,L effectively suppressed both H₂O₂- and hypokalemia-induced EADs and restored APD. These results, together with our previous work, demonstrate the potential of this hybrid experimental–computational approach to guide drug discovery or gene therapy strategies by identifying and targeting selective properties of LTCC.     

Effects of quinidine and lidocaine on action potential and membrane currents of frog ventricles

The effects of quinidine and lidocaine on frog ventricle were studied by using a single sucrose gap voltage clamp technique. In Ca2+-Ringer, quinidine (80 microM) caused slight prolongation of action potential duration (APD50) and significant inhibition of twitch tension. Lidocaine (40 microM) shortened APD50 without significant effect on twitch tension. In tetrodotoxin (TTX)-treated preparations, quinidine caused significant prolongation of APD50 from 529 +/- 19 msec to 597 +/- 11 msec, (n = 9) and inhibition of twitch tension, but lidocaine did not affect APD50 and twitch tension. Under voltage clamp condition, quinidine reduced peak inward current in the absence of TTX, but enhanced peak inward current in the presence of TTX. The steady state outward current was increased by quinidine. Lidocaine didn't affect peak inward current in the absence or in the presence of TTX. Membrane current through the inward rectifier (IK1) was slightly increased by lidocaine, but significantly inhibited by quinidine. The enhancement of peak inward current by quinidine was retarded or reversed in preparation bathed with Sr2+-Ringer. When Ni2+ was added to a preparation bathed in Ca2+-Ringer, an inhibition of calcium inward current and action potential plateau was observed. The spike amplitude of the action potential was, however, unaffected by Ni2+. In this Ni2+-treated preparation, lidocaine (20 microM) caused significant shortening of APD50 without significant effect on action potential amplitude. The shortening of APD50 was associated with a slight increase of steady state outward current. The increase of steady state outward current by lidocaine was absent in the TTX-treated preparation.(ABSTRACT TRUNCATED AT 250 WORDS)     

Focal extracellular potential: a means to monitor electrical activity in single cardiac myocytes

The focal extracellular potential (FEP) described in this study is an electrophysiological signal related to the transmembrane potential (V(m)) of cardiac myocytes that avoids the mechanical fragility, interference with contraction, and intracellular contact associated with conventional whole cell recording. One end of a frog ventricular myocyte was secured into a glass holding pipette. The FEP was measured differentially between this pipette and a bath pipette while the cell was voltage- or current-clamped by a third whole cell pipette. The FEP appeared as an amplitude-truncated action potential, while FEP duration accurately reflected the action potential duration (APD) at 90% repolarization (APD(90)). FEP magnitude increased as the holding pipette K(+) concentration ([K(+)]) was increased. The FEP-voltage relation was quasi-linear at negative V(m) with a slope that increased with elevated holding pipette [K(+)]. Increasing the membrane conductance inside the holding pipette by adding amphotericin B or cromakalim linearized the FEP-voltage relation across all V(m). The FEP accurately reported electrical activation and APD(90) during changes of stimulation frequency and episodes of cellular stretch.     

腺苷抗豚鼠室性心律失常的电生理研究

实验用全细胞膜片钳技术在单个豚鼠心室肌细胞上研究了腺苷 (Ado)对正常及异丙肾上腺素 (Iso)致豚鼠心室肌细胞动作电位、迟后除极 (DAD)、L 型钙电流 (ICa.L)和短暂内向电流 (Iti)的作用。结果表明 :(1)Ado在2 0～ 10 0 μmol/L时对豚鼠心室肌细胞动作电位和ICa .L无明显直接作用 ,但却可明显降低Iso所致的动作电位时程(APD)延长和ICa .L峰值增大 ,Iso (10nmol/L)使细胞APD50 从 3 40± 2 1ms延长到 486± 2 8ms (P <0 0 1) ,APD90从 3 61± 17ms延长至 5 0 1± 2 9ms (P <0 0 1) ;ICa .L峰值从 - 6 5 3± 1 4pA/pF增大到 - 18 2 8± 2 4pA/pF (P <0 0 1) ,电流电压曲线明显左移和下移 ;Ado (5 0 μmol/L)使APD50 和APD90 降至 40 3± 19ms和 419± 2 6ms ,但并不影响动作电位其它参数 ,使ICa.L峰值降低至 - 10 2± 1 5pA/pF (P <0 0 1)。 (2 )Iso (3 0nmol/L)可诱发心室肌细胞产生DADs,其发生率为 10 0 % ;Ado (5 0 μmol/L)可完全抑制Iso引发DADs;细胞经 - 40～ +2 0mV、时程 2s的除极电压 ,Iso (3 0nmol/L)诱导出Iti,其发生率为 10 0 % ;Ado (5 0 μmol/L)可明显抑制Iso致Iti的发生 ,其发生率降为 14 3 %。研究结果提示 ,Ado对豚鼠心室肌细胞动作电位和ICa.L无明显直接作用 ,但却可显著降低Is     

Cardiac mechano-electric feedback and electrical restitution in humans

Electrical restitution in the heart is the property whereby the action potential duration and conduction velocity of a beat of altered cycle length vary according to its immediacy to the preceding basic beat--the coupling interval, usually the diastolic interval. In general, action potential duration (APD) increases with increasing coupling interval, and the relation between action potential duration and the preceding diastolic interval describes the APD restitution curve. The latter has recently been the focus of considerable interest since the steepness of the initial part of the restitution curve plays an important role in electrical stability and arrhythmogenesis. Mechanical stretch has been shown to alter APD and hence refractoriness either through stretch activated channels or by influencing calcium cycling. Such an effect on refractoriness has been proposed as a mechanism of arrhythmogenesis particularly if spatially inhomogeneities manifest within the heart. Here, we review (1) the spatial and temporal characteristics of APD restitution in humans; (2) previously reported work showing that mechanical loading differentially effects APD of interpolated beats of altered cycle length, and hence alters the slope of the APD restitution curve; and (3) evidence that inhomogeneity of APD restitution slope may be an important factor in arrhythmogenesis.     

Overexpression of the Large-Conductance,Ca2+-Activated K+ (BK) Channel Shortens Action Potential Duration in HL-1 Cardiomyocytes

Long QT syndrome is characterized by a prolongation of the interval between the Q wave and the T wave on the electrocardiogram. This abnormality reflects a prolongation of the ventricular action potential caused by a number of genetic mutations or a variety of drugs. Since effective treatments are unavailable, we explored the possibility of using cardiac expression of the large-conductance, Ca²⁺-activated K⁺ (BK) channel to shorten action potential duration (APD). We hypothesized that expression of the pore-forming α subunit of human BK channels (hBKα) in HL-1 cells would shorten action potential duration in this mouse atrial cell line. Expression of hBKα had minimal effects on expression levels of other ion channels with the exception of a small but significant reduction in Kv11.1. Patch-clamped hBKα expressing HL-1 cells exhibited an outward voltage- and Ca²⁺-sensitive K⁺ current, which was inhibited by the BK channel blocker iberiotoxin (100 nM). This BK current phenotype was not detected in untransfected HL-1 cells or in HL-1 null cells sham-transfected with an empty vector. Importantly, APD in hBKα-expressing HL-1 cells averaged 14.3 ± 2.8 ms (n = 10), which represented a 53% reduction in APD compared to HL-1 null cells lacking BKα expression. APD in the latter cells averaged 31.0 ± 5.1 ms (n = 13). The shortened APD in hBKα-expressing cells was restored to normal duration by 100 nM iberiotoxin, suggesting that a repolarizing K⁺ current attributed to BK channels accounted for action potential shortening. These findings provide initial proof-of-concept that the introduction of hBKα channels into a cardiac cell line can shorten APD, and raise the possibility that gene-based interventions to increase hBKα channels in cardiac cells may hold promise as a therapeutic strategy for long QT syndrome.     

I<Subscript>CaL</Subscript> and I<Subscript>to</Subscript> mediate rate-dependent repolarization in rabbit atrial myocytes

Rate-dependent repolarization (RDR) of action potential (AP) in cardiomyocyte plays a critical role in the genesis of arrhythmias and RDR in atrium has been linked with atrial fibrillation. However, detailed studies focusing on the role of RDR in rabbit atrium are scant. In this study, atrial cells were isolated from rabbit heart and rate-dependent property was explored in single atrial cell to elucidate the underlying mechanism. Our results indicated that rate-dependent prolongation was evident at the action potential duration at 20% (APD20) and 50% (APD50) repolarization but not at 90% repolarization (APD90) under control condition. Using transient outward potassium current (I_to) inhibitor 4-Aminopyridine (4-AP, 2 mM) effectively eliminated the changes in APD20 and APD50, and unmasked the rate-dependent reduction of APD90 which could be diminished by further adding L-type calcium current (I_CaL) inhibitor nifedipine (30 μM). However, using the selective late sodium current (I_NaL) inhibitor GS-458967 (GS967, 1 μM) caused minimal effect on APD90 of atrial cells both in the absence and presence of 4-AP. In consistence with results from APs, I_to and I_CaL displayed significant rate-dependent reduction because of their slow reactivation kinetics. In addition, the magnitude of I_NaL in rabbit atrium was so small that its rate-dependent changes were negligible. In conclusion, our study demonstrated that I_to and I_CaL mediate RDR of AP in rabbit atrium, while minimal effect of I_NaL was seen.     

Dynamical effects of diffusive cell coupling on cardiac excitation and propagation: a simulation study

Cell coupling is considered to be important for cardiac action potential propagation and arrhythmogenesis. We carried out computer simulations to investigate the effects of stimulation strength and cell-to-cell coupling on action potential duration (APD) restitution, APD alternans, and stability of reentry in models of isolated cell, one-dimensional cable, and two-dimensional tissue. Phase I formulation of the Luo and Rudy action potential model was used. We found that stronger stimulation resulted in a shallower APD restitution curve and onset of APD alternans at a faster pacing rate. Reducing diffusive coupling between cells prolonged APD. Weaker diffusive currents along the direction of propagation steepened APD restitution and caused APD alternans to occur at a slower pacing rate in tissue. Diffusive current due to curvature changed APD but had little effect on APD restitution slope and onset of instability. Heterogeneous cell coupling caused APD inhomogeneities in space. Reduction in coupling strength either uniformly or randomly had little effect on the rotation period and stability of a reentry, but random cell decoupling slowed the rotation period and, thus, stabilized the reentry, preventing it from breaking up into multiple waves. Therefore, in addition to its effects on action potential conduction velocity, diffusive cell coupling also affects APD in a rate-dependent manner, causes electrophysiological heterogeneities, and thus modulates the dynamics of cardiac excitation. These effects are brought about by the modulation of ionic current activation and inactivation.     

Modeling ventricular repolarization: effects of transmural and apex-to-base heterogeneities in action potential durations

Heterogeneities in the densities of membrane ionic currents of myocytes cause regional variations in action potential duration (APD) at various intramural depths and along the apico-basal and circumferential directions in the left ventricle. This work extends our previous study of cartesian slabs to ventricular walls shaped as an ellipsoidal volume and including both transmural and apex-to-base APD heterogeneities. Our 3D simulation study investigates the combined effect on repolarization sequences and APD distributions of: (a) the intrinsic APD heterogeneity across the wall and along the apex-to-base direction, and (b) the electrotonic currents that modulate the APDs when myocytes are embedded in a ventricular wall with fiber rotation and orthotropic anisotropy. Our findings show that: (i) the transmural and apex-to-base heterogeneities have only a weak influence on the repolarization patterns on myocardial layers parallel to the epicardium; (ii) the patterns of APD distribution on the epicardial surface are mostly affected by the apex-to-base heterogeneities and do not reveal the APD transmural heterogeneity; (iii) the transmural heterogeneity is clearly discernible in both repolarization and APD patterns only on transmural sections; (iv) the apex-to-base heterogeneity is clearly discernible only in APD patterns on layers parallel to the epicardium. Thus, in our orthotropic ellipsoidal wall, the complex 3D electrotonic modulation of APDs does not fully mix the effects of the transmural and apex-to-base heterogeneity. The intrinsic spatial heterogeneity of the APDs is unmasked in the modulated APD patterns only in the appropriate transmural or intramural sections. These findings are independent of the stimulus location (epicardial, endocardial) and of Purkinje involvement.