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// Jianyong Qi 1, 2, * , Wenjun Pan 1, 2, * , Yafang Tan 1, 2, * , Jiaru Luo 1, 2, * , Dancai Fan 1, 2 , Juan Yu 3 , Jiashin Wu 4 and Minzhou Zhang 1, 2 1 AMI Key Laboratory of Chinese Medicine in Guangzhou, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangdong Province Hospital of Chinese Medicine, Guangzhou 510006, China 2 Intensive Care Research Team of Traditional Chinese Medicine, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangdong Province Hospital of Chinese Medicine, Guangzhou 510006, China 3 Animal Center, The Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangdong Province Hospital of Chinese Medicine, Guangzhou 510006, China 4 Department of Pharmaceutical Sciences, College of Pharmacy, Northeast Ohio Medical University, Rootstown, Ohio 44272, USA * These authors have contributed equally to this work and share the first authorship Correspondence to: Minzhou Zhang, email: minzhouzhang8@163.com Keywords: myocardial ischemia; isoproterenol; signaling pathway; rat; Shexiang Tongxin dropping pill Received: July 27, 2017 Accepted: October 28, 2017 Published: November 14, 2017 ABSTRACT Shexiang Tongxin dropping pill (STDP) is a formulae of Chinese Medicine commonly used to treating angina pectoris in China. However, its mechanism of action is still yet unclear. This study investigated the roles of STDP on myocardial ischemia injury. We constructed a rat model of myocardial injury (isoproterenol subcutaneous injection, i.h, 85 mg/kg/day for 2 days), and compared among 4 groups: CON (control), ISO (ischemic injury model), MET (metoprolol), and STDP. Serum contents of Troponin I (cTnI), creatine kinase (CK), CK-MB, lactate dehydrogenase (LDH), alpha-hydroxybutyric dehydrogenase (α-HBD), and Aspartate Aminotransferase were detected and five STDP doses (1, 10, 100, 1000 and 10000 mg/kg/day) were chosen to obtain a dose-response curve. Western-blot was used to detect phosphorylations of extracellular signal-regulated kinase 1/2 (ERK1/2), protein kinase B (AKT), and camodulin kinase II (CamkII). Furthermore, an ERK1/2 inhibitor PD98059, a phosphatidylinositol-3-kinase inhibitor, LY294002, and a CamKII inhibitor, KN-93 were administered i.h. Results: cTnI, CK, CK-MB, α-HBD, and LDH were significantly lower in STDP than ISO ( P

To analyze the effects of salvianolate on myocardial infarction in a murine in vivo model of ischemia and reperfusion (I/R) injury.Myocardial I/R injury model was constructed in mice by 30 min of coronary occlusion followed by 24 h of reperfusion and pretreated with salvianolate 30 min before I/R (SAL group). The SAL group was compared with SHAM (no I/R and no salvianolate), I/R (no salvianolate), and ischemia preconditioning (IPC) groups. Furthermore, an ERK1/2 inhibitor PD98059 (1 mg/kg), and a phosphatidylinositol-3-kinase (PI3-K) inhibitor, LY294002 (7.5 mg/kg), were administered intraperitoneal injection (i.p) for 30 min prior to salvianolate, followed by I/R surgery in LY and PD groups. By using a double staining method, the ratio of the infarct size (IS) to left ventricle (LV) and of risk region (RR) to LV were compared among the groups. Correlations between IS and RR were analyzed. Western-blot was used to detect the extracellular signal-regulated kinase 1/2 (ERK1/2) and protein kinase B (AKT) phosphorylation changes.There were no significant differences between RR to LV ratio among the SHAM, I/R, IPC and SAL groups (P0.05). The SAL and IPC groups had IS of 26.1%±1.4% and 22.3%±2.9% of RR, respectively, both of which were significantly smaller than the I/R group (38.5%±2.9% of RR, P0.05, P0.01, respectively). Moreover, the phosphorylation of ERK1/2 was increased in SAL group (P0.05), while AKT had no significant change. LY294002 further reduced IS, whereas the protective role of salvianolate could be attenuated by PD98059, which increased the IS. Additionally, the IS was not linearly related to the RR (r=0.23, 0.45, 0.62, 0.17, and 0.52 in the SHAM, I/R, SAL, LY and PD groups, respectively).Salvianolate could reduce myocardial I/R injury in mice in vivo, which involves an ERK1/2 pathway, but not a PI3-K signaling pathway.

Ethnopharmacological relevance Cardiac hypertrophy is a key pathologic process in heart failure. Songling Xuemaikang Capsule (SXC), is a formulae of Chinese Medicine commonly used in China to treat hypertension and heart failure. However, its mechanism of effects on cardiac hypertrophy is still unclear. Aim of the study The aims of the present study were to investigate the cardio-protection roles and detailed mechanisms of SXC on cardiac hypertrophy in vivo and in vitro. Materials and methods A rat model of cardiac hypertrophy was constructed by isoproterenol (ISO) intraperitoneal injection (i.p), 10 mg/kg/day for 3 days, and 4 groups were compared: CON (n = 8), ISO (n = 8), MET (metoprolol, positive drug treatment, n = 7), and SXC (SXC treatment, n = 6). Cardiac structure and function were evaluated with echocardiography in vivo. Dose-dependent curve was obtained with SXC different concentrations. In addition, H9C2 rat cardiomyocytes were cultured in vitro and the phosphorylation of ERK1/2, p38, JNK, AKT, and protein expression of CaN, CaMKIIδ, GATA4 were detected with Western blot test. Results The results showed that SXC reduced diastolic thickness of left ventricular posterior wall, while did not change ejection fraction and fraction shortening significantly (P > 0.05). SXC inhibit ISO-induced cardiac hypertrophy dose-dependently with 50% inhibiting concentration (IC50) is 0.504 g/kg/day. Moreover, SXC inhibited the protein expression of CaMKIIδ, and the phosphorylation of ERK1/2, so inhibiting protein expression of GATA4 in nucleus, and brain natriuretic peptide in serum (P Conclusion The mechanism of SXC in the treatment of heart diseases involves SXC dose-dependently inhibited the ISO-induced cardiac hypertrophy via inhibiting CaMKIIδ and ERK1/2/GATA4 signaling pathway.

Background Ischemia suppresses action potentials (APs) by elevating interstitial K + and activating K ATP channels and alters cytosolic Ca 2+ transients (CaTs) via metabolic inhibition. Objective To test the hypothesis that AP and CaT respond to ischemia with different spatiotemporal courses and patterns. Methods Thirty-four transmural wedges were isolated from canine left ventricular free walls, perfused arterially, and stained with voltage- and Ca 2+ -sensitive dyes. Twenty-eight wedges underwent 15 minutes of arterial occlusion during pacing at a cycle length (PCL) of 300 ms (n = 19) or 600 ms (n = 9). Six other wedges had a sequential reduction of perfusion flow from full to 50%, 25%, and 10% at 300 ms PCL. AP and CaT were recorded on the cut-exposed transmural surfaces with an optical mapping system. Results Although ischemia suppressed APs, it enhanced CaT to 150% ± 10% (more in the endocardium than in the epicardium) and induced CaT alternans during the first 2 minutes of arterial occlusion and then suppressed CaT (PCL = 300 ms). Enhancement of CaT (to 159% ± 23%) also occurred during low flow (25%) perfusion (PCL = 300 ms). Faster suppression of AP occurred with subepicardial preference as compared to that of CaT. After 15 minutes of arterial occlusion, AP and CaT remained in only small regions during 300 ms PCL but were preserved in most regions during 600 ms PCL. Conclusions Early ischemia induced a surge and alternans in CaT and caused its dissociation from AP both in time course of suppression and in spatial distribution. These results suggested that there were different cellular regulatory mechanisms of AP and of CaT in responding to ischemia from arterial occlusion.

Patients with heart failure (HF) have high mortality and mobility. Xinmailong (XML) injection, a Chinese Medicine, is clinically effective in treating HF. However, the mechanism of XML’s effectiveness on HF was unclear, and thus, was the target of the present study. We created a mouse model of pressure-overload-induced HF with transverse aortic constriction (TAC) surgery and compared among 4 study groups: SHAM (n = 10), TAC (n = 12), MET (metoprolol, positive drug treatment, n = 7) and XML (XML treatment, n = 14). Dynamic changes in cardiac structure and function were evaluated with echocardiography in vivo. In addition, H9C2 rat cardiomyocytes were cultured in vitro and the phosphorylation of ERK1/2, AKT, GSK3β and protein expression of GATA4 in nucleus were detected with Western blot experiment. The results showed that XML reduced diastolic thickness of left ventricular posterior wall, increased ejection fraction and fraction shortening, so as to inhibit HF at 2 weeks after TAC. Moreover, XML inhibited the phosphorylation of ERK1/2, AKT and GSK3β, subsequently inhibiting protein expression of GATA4 in nucleus (P

Myocardial ischemia/reperfusion (I/R) can induce lethal ventricular arrhythmia and myocardial infarction. One of the clinical strategies for managing patients with high risk of myocardial I/R is to prevent the occurrence of arrhythmias and limit the size of infarction following a coronary episode. Tongguan Capsule (TGC) is one of the popular herbal remedies in treating coronary artery disease in the clinics of Chinese medicine. However, the potential roles and mechanisms of TGC in reducing I/R injury are still unclear. The present study statistically assessed the effectiveness of TGC in reducing I/R injury by comparing the infarct size (IS), risk region (RR), and arrhythmia (in electrocardiogram) among four groups of surgically created mice models of myocardial I/R: SHAM, I/R, VER (I/R with verapamil 20 mg/kg pretreatment), and TGC (I/R with TGC 5 g/kg/d pretreatment). We found that IS was significantly smaller in the TGC and VER groups than I/R group, and the incidence of arrhythmias was reduced in the TGC group compared with I/R group, although there were no differences in RR among the four groups. We conclude that TGC is effective in reducing I/R injury in mice. These results provided an experimental basis for clinical application of TGC in reducing I/R injury.

Background Heart failure(HF) is a dangerous disease that affects millions of patients. Radix Salvia is widely used in Chinese clinics to treat heart diseases. Salvianolic acid B(SalB) is the major active component of Radix Salvia. This study investigated the mechanisms of action and effects of SalB on HF in an experimental mouse model of HF. Methods We created a mouse model of HF by inducing pressure overload with transverse aortic constriction(TAC) surgery for 2 weeks and compared among 4 study groups: SHAM group (n = 10), TAC group (n = 9), TAC+MET group (metprolol, positive drug treatment, n = 9) and TAC+SalB group (SalB, 240 mg•kg-1•day-1, n = 9). Echocardiography was used to evaluate the dynamic changes in cardiac structure and function in vivo. Plasma brain natriuretic peptide (BNP) concentration was detected by Elisa method. In addition, H9C2 rat cardiomyocytes were cultured and Western blot were implemented to evaluate the phosphorylation of ERK1/2, AKT, and protein expression of GATA4. Results SalB significantly inhibited the phosphorylation of Thr202/Tyr204 sites of ERK1/2, but not Ser473 site of AKT, subsequently inhibited protein expression of GATA4 and plasma BNP(P < 0.001), and then inhibited HF at 2 weeks after TAC surgery. Conclusions Our data provide a mechanism of inactivating the ERK1/2/GATA4 signaling pathway for SalB inhibition of the TAC-induced HF.

Patients with Brugada syndrome (BS) often have spontaneous changes in their electrocardiogram (ECG).To evaluate the significance of ECG alterations, we investigated the relationships between the ECG and the occurrence of ventricular fibrillation (VF) in both patients and an experimental model of BS.In study 1, we evaluated ECG alterations in BS patients with (VF+, n = 33) and without (VF-, n = 41) spontaneous VF. We defined type 0 ECG as coved-type ST elevation without a negative T wave, which represents the existence of loss-of-dome (LOD) type action potentials (APs). In study 2, we optically mapped epicardial APs and recorded transmural ECGs in 34 canine right ventricular tissues with a drug-induced BS model by a combination of pinacidil and pilsicainide.In study 1, changes in ST level ≥0.2 mV were more frequent in the VF+ group than in the VF- group (P.01). Spontaneous ECG alterations and appearances of types 1 and 0 ECGs were more frequent in the VF+ group than in the VF- group (P.01). In study 2, BS model with spike-and-dome (SAD) epicardial APs exhibited type 1 ECG. Deepening of the phase 1 notch of the APs induced heterogeneous conversion of the APs (SAD→LOD) and resulted in ECG conversion from type 1 to type 0. Significant AP heterogeneity often appeared during AP alterations and initiated phase 2 reentry. Tissues having ventricular tachycardia (VT; n = 20) had more frequent alterations in APs and ECG than in tissues without VT (n = 14; P.01).ECG alterations, especially conversion between types 0 and 1, are associated with significant AP heterogeneity that can initiate VF in BS.

Although radiofrequency catheter ablation (RFCA) has been used to treat patients with Brugada syndrome (BS), it is difficult to eliminate polymorphic ventricular tachycardias (VTs) completely.The purpose of this study was to determine the efficacy of RFCA in eliminating recurrent VTs in an experimental model of BS.We optically mapped electrical activity on the epicardial (n = 9) or transmural (n = 8) surface in 17 arterially perfused canine right ventricle preparations. Using pinacidil (5 microM) and pilsicainide (5 microM), we induced a model of BS that showed spontaneous VT. We then applied RFCA to the earliest activation site of premature ventricular complexes (PVCs) in the epicardium (EPI) or endocardium (ENDO) of the RV.After induction of BS, the transmural electrocardiogram (ECG) showed BS-type ECG in association with prominent heterogeneity of action potential duration (APDs) within the EPI (APD: maximum 272 +/- 39 ms, minimum 200 +/- 39 ms, P.01), but not within the ENDO. PVCs originated in the EPI region having short APDs and triggered functional reentry causing VT. Multiple epicardial foci of PVCs existed in each tissue (3.7 +/- 1.9 foci/tissue). RFCA at the earliest activation site of PVCs in the EPI disconnected the short and long APD regions and eliminated all PVCs and VTs, although APD heterogeneity still existed. All successful RFCA lesions were confined to the EPI. RFCA in the ENDO failed to eliminate VT or PVCs.These experimental observations suggest that RFCA applied to the EPI may be more effective than applied to the ENDO in eliminating VT in patients with BS.

Background Acute ischemia depresses tissue excitability more rapidly in the epicardium than in the endocardium of the canine left ventricular (LV) free wall. However, the effects of acute ischemia on conduction in the interventricular septum (IVS), which is composed of right ventricular (RV) and LV endocardium and midmyocardium without epicardium, are less known. Objective The purpose of this study was to evaluate the hypothesis that the IVS exhibits transseptal differences in local tissue response to acute ischemia. Methods Isolated canine IVS preparations were perfused through the septal branch of the anterior descending coronary artery, and conduction on the cut-exposed transseptal surfaces was optically mapped before and after two sequential episodes of 8 minutes of global ischemia (separated by >60 minutes of reperfusion). The preparations were paced alternately between the RV endocardium and LV endocardium at cycle lengths of 250, 300, and 1,500 ms. Results Prior to ischemia, transseptal conduction was radial and symmetric during either RV endocardial or LV endocardial pacing at all cycle lengths. Eight minutes of ischemia depressed conduction velocity more in the RV half than in the LV half of the IVS and caused local conduction block in the sub-RV endocardium, especially during rapid pacing. The K ATP channel blocker glibenclamide (10 μmol/L) prevented development of this transseptal asymmetry and conduction block during ischemia. Conclusion Acute global ischemia increased transseptal heterogeneity and induced sub-RV endocardial block of conduction via activation of the ATP-sensitive potassium current. Such changes could contribute to initiation of arrhythmia in patients with septal infarction.

Background Brugada syndrome (BrS) is characterized by repolarization abnormality with ST-segment elevation in the right ventricular outflow tract (RVOT). Objective Although action potential (AP) heterogeneity is associated with induction of ventricular arrhythmias (VA) in BrS, clinical evidence and its experimental correlations are still absent and are the focus of this study. Methods We evaluated repolarization heterogeneity in 15 patients with BrS using body surface mapping and in 8 pairs of isolated canine RVOT and right ventricular anteroinferior (RVAI) preparations having drug-induced BrS using optical mapping. Results Patients had large J-ST-segment elevation and long QT interval in the RVOT at baseline. Administration of pilsicainide (1 mg/kg) exaggerated J-ST-segment elevation, caused simultaneous long and short QT intervals in the RVOT, and induced polymorphic ventricular tachycardia (VT) and T wave alternans (TWA). Dispersion of QT within the RVOT after pilsicainide was greater in patients that had syncope or ventricular fibrillation than those that did not. Ventricular arrhythmias originated from the RVOT along with local electrocardiogram changes and TWA. Repolarization heterogeneity was much less in areas outside the RVOT. Inducing BrS increased AP heterogeneity (with and without spike-and-dome) within the RVOT epicardium. Phase 2 reentry and TWA originated from the epicardium in 88% and 50% of RVOT preparations, respectively. In contrast, the RVOT endocardium and RVAI had little AP heterogeneity, with neither reentry nor TWA. Conclusion The instability and heterogeneity of repolarization within the epicardium of the RVOT seem to be associated with arrhythmogenesis in both patients and in the in vitro tissue models of BrS.

Objective Andersen–Tawil syndrome (ATS) is a channelopathy affecting inward rectifier potassium I K1 with QT prolongation, large U waves, and frequent ventricular tachycardia (VT). Although ATS is clinically defined and genetically identified, its electrophysiological mechanism is still unclear, and thus, was the subject of the current study. Methods and results We replicated the major electrophysiological features of ATS with cesium chloride (CsCl, at I K1 blockade concentration of 5–10 mmol/l) in 23 isolated canine left ventricular tissues perfused arterially with Tyrode's solution having normal or low potassium concentrations, [K+]o. We mapped action potentials (APs) on the cut-exposed transmural surface of the wedges in control, after CsCl, and CsCl with 0.15 μmol/l isoproterenol (CsCl+ISP). CsCl delayed late phase 3 repolarization and prolonged the duration of the AP, more so during low [K+]o perfusion. Rapid pacing induced delayed afterdepolarizations (DADs) in all low [K+]o and in 71% of normal [K+]o preparations after CsCl treatment. Addition of ISP induced DADs in all preparations. DADs originated in mid-to-endocardium, and initiated VT after CsCl+ISP. Migration of DAD–VT foci resulted in multifocal VT. Alternating DADs at 2 foci resulted in bidirectional VT. There were more foci and longer durations of VT at low [K+]o than at normal [K+]o. Delayed late phase 3 repolarization of APs and DADs generated U waves. Verapamil abolished all DADs and VT. Conclusions CsCl blockade of I K1 produced a ventricular wedge model of ATS. Suppressing I K1 generated U waves by delaying late repolarization of APs and creating DADs, and promoted polymorphic VT by triggering DADs at multiple shifting sites.

Background Fever promotes ventricular arrhythmias in Brugada syndrome (BrS). Hypothermia can induce BrS electrocardiogram (ECG) and arrhythmia. However, the mechanisms are unclear. Objective We evaluated the hypothesis that pathological temperatures promoted arrhythmogenesis by modulating the spatial heterogeneity and functional dynamics of right ventricular electrophysiological activity. Methods We mapped action potentials (APs) on the epicardial or cut-exposed transmural surfaces and recorded transmural ECGs in 27 arterially perfused canine right ventricular preparations before and after inducing BrS at 32°C, 36.5°C, and 40°C. Results We observed major intraepicardial dispersion of AP duration (APD) and reversal of transmural gradient of APD in association with manifestation of BrS at 36.5°C. Reducing the temperature to 32°C prolonged APDs and enhanced the phase 1 notch of epicardial APs, while 40°C caused opposite changes. Prominent phase 2 domes of APs frequently led to spontaneous premature ventricular activations (PVAs), which conducted to surrounding regions having shorter APDs. Longer APDs at 32°C and 36.5°C frequently blocked reentry, although they promoted PVA, while shortened APDs at 40°C facilitated reentrant ventricular tachycardia. During bradycardia (2,000 ms), the J-ST elevation in the ECG was enhanced at 32°C and attenuated at 40°C. Rapid pacing (500 ms) eliminated the dome of epicardial APs and enhanced J-ST elevation at each temperature. Blocking the transient outward current, I to , with 4-aminopyridine reduced J-ST elevation and eliminated the PVA and reentry. Conclusions In this BrS model, prolongation and increased dispersion of APDs promoted spontaneous activation during hypothermia, while APD abbreviation facilitated reentry during hyperthermia. I to mediated the arrhythmogenicity.

Background The Brugada syndrome is characterized by ST-segment elevation on the ECG, especially in the right precordial leads sensitive to the right ventricular outflow tract (RVOT). Objectives The purpose of this study was to evaluate the hypothesis that right ventricular electrophysiologic heterogeneity caused arrhythmogenicity in the Brugada syndrome. Methods Action potentials (APs) were mapped on the epicardium of 14 RVOT preparations and on the transmural surfaces of 15 pairs of RVOT and right ventricular anteroinferior (RVAI) preparations isolated from canine hearts. Brugada ECG and arrhythmias were induced with pilsicainide (2.5–12.5 μmol/L), pinacidil (1.25–12.5 μmol/L), and terfenadine (2.0 μmol/L). Results Low doses of drugs elevated the J-ST segment and induced APs with both short and long action potential durations (APDs) in contiguous RVOT epicardial regions. In addition, APs in the RVOT had a larger phase 1 notch and longer APD than in RVAI. The longest APDs were in the epicardium in RVOT but in the endocardium in RVAI regions. High doses of drugs eliminated the phase 2 dome of the AP and abbreviated APDs in the epicardium but not in endocardium and reduced the epicardial heterogeneity of APs but increased the transmural gradient of APD in 14 (93%) of the RVOT preparations. In contrast, abbreviations of epicardial APDs occurred in only 4 (27%) of the RVAI preparations. Ventricular tachycardia occurred more frequently in the RVOT (47%) than in paired RVAI preparations (7%). Blocking the transient outward current reduced the heterogeneity of APs and eliminated arrhythmogenicity in all preparations. Conclusion Compared with the RVAI region, the RVOT has greater electrophysiologic heterogeneity that contributes to arrhythmogenicity in this model of Brugada syndrome.

Macroscopic T wave alternans (TWA) associated with increased occurrence of ventricular arrhythmias has been reported in patients with Brugada syndrome. However, the mechanisms in this syndrome are still unclear. We evaluated the hypothesis that TWA in Brugada syndrome was caused by the dynamic instability and heterogeneity of action potentials (APs) in the right ventricle. Using an optical mapping system, we mapped APs on the epicardium or transmural surfaces of 28 isolated and arterially perfused canine right ventricular preparations having drug-induced Brugada syndrome (in μmol/l: 2.5–15 pinacidil, 5.0 terfenadine, and 5.0–13 pilsicainide). Bradycardia at cycle length (CL) of 2,632 ± 496 ms ( n = 19) induced alternating deep and shallow T waves in the transmural electrocardiogram. Compared with the shallow T waves, deep T waves were associated with epicardial APs having longer durations and larger domes. Adjacent regions having APs with alternating domes, with constant domes, and without domes coexisted simultaneously in the epicardium and caused TWA. In contrast to the alternating epicardial APs, midmyocardial and endocardial APs did not change during TWA. Alternans could be terminated by rapid (CL: 529 ± 168 ms, n = 7) or very slow (CL: 3,000 ms, n = 7) pacing. The heterogeneic APs during TWA augmented the dispersion of repolarization both within the epicardium and from the epicardium to the endocardium and caused phase 2 reentry. In this drug-induced model of Brugada syndrome, heterogeneic AP contours and dynamic alternans in the dome of right ventricular epicardial, but not midmyocardial or endocardial, APs caused TWA and heightened arrhythmogenicity in part by increasing the dispersion of repolarization.

Objectives To explore the mechanism, we tested the hypothesis that premature epicardial stimulation transiently increased the dispersion of repolarization leading to VT. Background Premature stimulation initiated ventricular tachycardia (VT) when applied to the epicardium but not to the endocardium in a canine model of long QT syndrome (LOTS). Methods We optically mapped action potentials (APs) on the cut-exposed transmural surfaces of isolated wedges of canine ventricular walls perfused with anemone toxin II (ATX-II), which produced type 3 LQTS with an asymmetrical transmural profile of repolarization that was earliest in the epicardium and latest in deep subendocardium. Results Earliest excitable epicardial stimulation triggered VT in 5 of 18 wedges receiving ≥5 nmol/L ATX-II by direct activation of epicardium, which delayed repolarization in the still refractory midmyocardium and further enhanced the dispersion of repolarization. These VTs were initiated 197 ± 72 ms (n = 10) after the premature stimulation, from focal regions of earliest repolarization downstream to the steepest local spatial gradients of repolarization, and maintained by new focal activation and reentry. Transmural differences in the cycle lengths of activations altered conduction pathways and resulted in torsades de pointes-like polymorphic VT. In contrast, VTs were not initiated by endocardial stimulation at the same premature intervals or when ATX-II was ≤2.5 nmol/L. Failed VT initiation was associated with significantly lower maximum local gradient of repolarization. Conclusions Heterogeneic repolarization in LQTS provides a transmural asymmetrical substrate for the earliest excitable epicardial, but not endocardial, stimulation to further delay midmyocardial repolarization and produce a steep spatial gradient of repolarization potential initiating torsades de pointes-like polymorphic VT.

Objective: Ventricular tachyarrhythmias (VTs) occur frequently in patients having long QT syndrome (LQTS) or after acute myocardial ischemia. However, the synergistic effects of ischemia and LQTS on arrhythmia development are unclear. We evaluated the contribution of a prior episode of ischemia on the arrhythmogenicity of the LQTS. Methods: Using a 256-channel optical mapping system, we mapped action potentials on the cut-exposed transmural surfaces of perfused and preconditioned muscle wedges isolated from canine left ventricular walls and recorded their transmural electrocardiogram (ECG). Results: We observed that 40 min of global ischemia followed by 60 min of reperfusion, at which time action potential duration (APD) and conduction velocity had recovered, significantly enhanced the APD prolongation produced by 20 nmol/l anemone toxin II (ATX-II). Wedges after the above ischemia, reperfusion, and ATX-II procedures had 100% (8/8) occurrences of early afterdepolarizations (EADs) and 87.5% (7/8) occurrences of spontaneous VTs and reentry. We observed epicardial, midmyocardial, and endocardial occurrences of EADs in one, seven, and four wedges, respectively. Focal EADs and reentry were responsible for 73% and 18% of the repetitive activations in the VTs. In contrast, neither EADs nor VTs occurred in eight control wedges following identical procedures except without ischemia, and VT occurred in 20% wedges (2/10) after ischemia and reperfusion but before ATX-II. Conclusion: A prior episode of acute ischemia, even after apparent electrophysiologic recovery, enhances the arrhythmogenicity of LQTS induced by ATX-II through the development of EADs and reentry.

Ischemia depresses tissue excitability more rapidly in the ventricular epicardium than in the endocardium. We hypothesized that this would provide the substrate for transmural reentry originating in the epicardium. We mapped transmural conduction in isolated and perfused wedges taken from canine left ventricles during global ischemia while pacing alternately between the epicardium and endocardium. Ischemia reduced conduction velocity more in the epicardium than in the endocardium. We observed that the epicardial-initiated activation penetrated the ventricular wall transmurally while failing to conduct laterally along the epicardium, then conducted laterally along the endocardium and midmyocardium, and reentered the epicardium in 9 of 16 wedges during epicardial stimulation after 600 ± 182 s of ischemia. Endocardial stimulation applied immediately before or after the epicardial stimulation initiated activation that spread quickly along the endocardium and then transmurally to the epicardium without reentry in six of the nine wedges. The transmural asymmetric conduction was not observed in four separate wedges after the endocardium was removed. Therefore, ischemia-induced transmural gradient of excitability provided the substrate for reentry during epicardial stimulation.

Background— The junction between the coronary sinus (CS) musculature and both atria contributes to initiation of atrial tachyarrhythmias. The current study investigated the effects of CS isolation from the atria by radiofrequency catheter ablation on the induction and maintenance of atrial fibrillation (AF). Methods and Results— Using an optical mapping system, we mapped action potentials at 256 surface sites in 17 isolated and arterially perfused canine atrial tissues containing the entire musculature of the CS, right atrial septum, posterior left atrium, left inferior pulmonary vein, and vein of Marshal. Rapid pacing from each site before and after addition of acetylcholine (0.5 μmol/L) was applied to induce AF. Epicardial radiofrequency catheter ablation at CS-atrial junctions isolated the CS from the atria. Rapid pacing induced sustained AF in all tissues after acetylcholine. Microreentry within the CS drove AF in 88% of preparations. Reentries associated with the vein of Marshall (29%), CS-atrial junctions (53%), right atrium (65%), and pulmonary vein (76%) (frequently with 2–4 simultaneous circuits) were additional drivers of AF. Radiofrequency catheter ablation eliminated AF in 13 tissues before acetylcholine ( P P Conclusions— CS and its musculature developed unstable reentry and AF, which were prevented by isolation of CS musculature from atrial tissue. The results suggest that CS can be a substrate of recurrent AF in patients after pulmonary vein isolation and that CS isolation might help prevent recurrent AF.

Coronary occlusion and reperfusion produce tachyarrhythmias. We tested the hypothesis that variations in transmural activation after global ischemia and reperfusion were responsible for arrhythmias. We arterially perfused 36 isolated transmural wedges from canine left ventricular free walls. After ≥100 min of stabilization, the artery was occluded for 25 min, followed by reperfusion at various flow rates. We recorded 256 channels of fluorescent action potentials on transmural surfaces from preocclusion to >15 min after reperfusion. During endocardial pacing at 300 ms, ischemia of ≥570 ± 165 s ( n = 34) produced 1:1 endocardial conduction and then 2:1 and 4:1 block as the wave fronts conducted toward epicardium. Transmural reentry appeared after 535 ± 146 s of ischemia ( n = 31). Further ischemia caused epicardial inactivation and eliminated reentry ( n = 24). During reperfusion, tissues progressed through sequences of epicardial inactivation and reappearance of activation with 1:1, 2:1, and 4:1 conduction; both sustained and nonsustained reentry occurred. We conclude that heterogeneous activation responses to endocardial pacing during acute ischemia provide the substrate for initiating reentry, suppressed reentry during further ischemia, and caused reentry during reperfusion.

Cytochalasin D Induced Cardiac Contraction Failure. Introduction: 2,3-Butanedione monoxime (BDM) has been widely used to inhibit contraction during optical recordings of cardiac membrane voltage changes, even though it markedly abbreviates cardiac action potentials. Methods and Results: We compared the effects of BDM and of the F-actin disrupter cytochalasin D (cyto D) on isometric twitch force and transmembrane action potentials in isolated canine right ventricular trabeculae superfused with Tyrode's solution (2 mmol/L CaCl2, 37°C) and stimulated at 0.5 Hz. BDM at 10 mmol/L and cyto D at 80 μmol/L were equally effective in reducing peak isometric force to 10%± 3% (n = 6; mean ± SEM) and 8%± 1% (n = 8), respectively. Neither agent significantly altered resting tension. While 10 mmol/L BDM markedly shortened the action potential duration at 90% repolarization (APD90) from 198 ± 7 msec to 146 ± 9 msec (P < 0.001), 80 μmol/L cyto D had no significant effects on APD90 or on any other action potential parameter. The effects of BDM on peak isometric force and APD were completely reversible after 15 minutes of washout, whereas in the cyto D group contractile force continued to be reduced (13%± 3%) and action potential characteristics did not show significant changes from control values after a 60-minute period of superfusion with cyto D-free Tyrode's solution. Conclusion: We conclude that cyto D should be considered an alternative excitation-contraction uncoupler for optical mapping studies of cardiac repolarization.

Background. Cardiac hypertrophy occurs in many cardiovascular diseases. Apocynum tablet (AT), a traditional Chinese medicine, has been widely used in China to treat patients with hypertension. However, the underlying molecular mechanisms of AT on the hypertension-induced cardiac hypertrophy remain elusive. The current study evaluated the effect and mechanisms of AT on cardiac hypertrophy.Methods. We created a mouse model of cardiac hypertrophy by inducing pressure overload with surgery of transverse aortic constriction (TAC) and then explored the effect of AT on the development of cardiac hypertrophy using 46 mice in 4 study groups (combinations of AT and TAC). In addition, we evaluated the signaling pathway of phosphorylation of ERK1/2, AKT, and protein expression of GATA4 in the cardioprotective effects of AT using Western blot.Results. AT inhibited the phosphorylation of Thr202/Tyr204 sites of ERK1/2, Ser473 site of AKT, and protein expression of GATA4 and significantly inhibited cardiac hypertrophy and cardiac fibrosis at 2 weeks after TAC surgery (P<0.05).Conclusions. We experimentally demonstrated that AT inhibits cardiac hypertrophy via suppressing phosphorylation of ERK1/2 and AKT.

Objective. To determine the impact of an integrative medicine clinical pathways (CPs) on the length of in-hospital stay and on outcomes for patients with acute myocardial infarction (AMI).Methods. A multicenter nonrandomized controlled trial enrolling 197 consecutive patients with AMI at eight urban TCM hospitals was conducted between 1 January 2010 and 31 October 2010. These patients were enrolled in the interventional group after the CPs had been implemented. The control group included 405 patients with AMI from eight hospitals; these patients were treated between 1 January 2008 and 31 December 2009, before the CPs were implemented. Outcome measures were the length of hospital stay costs of medical care, and major cardiovascular events (MACEs) during hospitalization.Results. Compared with the control group, the patients in intervention group had a shorter length of hospital stay (9.2±4.2days versus12.7±8.6days,P<0.05), and reduced healthcare costs in hospital (46365.7±18266.9versus52866.0±35404.4,P<0.05). There were statistically significant differences in MACE between the two groups during the hospitalization period (2.5% versus 6.9%,P=0.03).Conclusion. These data suggest that the development and implementation of the clinical pathways based in Integrative Medicine could further improve quality of care and outcome for patients with AMI.

Coronary sinus (CS) musculature connects the right atria (RA) and the left atria (LA). However, the functional significance of the electrical junctions between the atria and the CS musculature is still unclear.We investigated electrophysiological properties of the CS-atrial connections and their role in atrial fibrillation.By using an optical mapping system, we mapped action potentials at 256 sites on the epicardial surface of 16 isolated and arterial-perfused canine atrial tissues containing the entire musculature of the CS, lower RA, posterior LA, left inferior pulmonary vein, and vein of Marshal. We paced from each of the above regions to measure electrophysiological properties and inducibility of atrial tachyarrhythmias.The CS musculature connected to the RA at the ostium of the CS and to the LA at proximal and distal CS sites. Electrical conduction across each of these CS-atrial junctions was slow (P.01), but not decremental. Rapid pacing often induced entrance block at the CS-atrial junctions and resulted in sequential changes of activation sequence in the CS. Macroreentrant circuit involving the CS musculature and the CS-atrial junctions occurred in association with conduction block at these junctions. The reentrant circuit was usually unstable and resulted in atrial fibrillation-like electrocardiographic activity.The anatomical and electrical connections between the CS musculature and the RA and the LA caused conduction slowing and block in the CS musculature and its atrial junctions, which frequently initiated unstable macroreentry and atrial fibrillation.

Background Genetic defects in the sodium channel or in the calcium channel have been identified in patients with Brugada syndrome (BS). However, the differences in their genotype-phenotype correlations are still unclear. Objective We evaluated the phenotypic differences and therapeutic effects between the sodium channel and calcium channel abnormalities in in vitro models of BS. Methods We created two models of BS in 18 isolated and arterially perfused canine right ventricular preparations: (1) sodium channel dysfunction model (Na model, n=11) by pilsicainide and pinacidil and (2) calcium channel dysfunction model (Ca model, n=7) by verapamil; optically mapped action potentials (APs) on their transmural surface; and evaluated APs and electrocardiograms (ECGs) at pacing cycle lengths (CLs) of 2,000 and 1,000 ms. Results CL = 1,000 ms: Both models had coved-type ST elevation in the ECG, longer AP duration (APD) in the epicardium than in the endocardium, and a similar incidence of spontaneous ventricular arrhythmias. However, the Ca model had a higher incidence of T wave alternans (TWA) than the Na-model. CL=2,000 ms: ECGs of the Ca model converted to saddleback-type ST elevation with shorter APDs in the epicardium than in the endocardium, whereas the Na model still had coved-type ST elevation and longer APDs in the epicardium. None of the Ca model preparations had ventricular arrhythmias or TWA, although the Na model had frequent ventricular arrhythmias and TWA. Conclusion Although both sodium channel and calcium channel dysfunction produced similar BS ECGs and arrhythmogenesis at 60 bpm, calcium channel dysfunction was associated with a higher incidence of TWA at 60 bpm, less ST elevation, and fewer arrhythmias at 30 bpm compared with sodium channel dysfunction.

Brugada syndrome (BrS), caused by ion channel abnormalities, is characterized by ST segment elevation and negative T waves in the right precordial electrocardiographic (ECG) leads recorded over the right ventricular outflow tract (RVOT). BrS is sensitive to body temperature and can lead to T-wave alternans (TWA), ventricular tachycardia, and sudden death. Recent studies in an isolated canine RVOT model of BrS demonstrated that reversal of the transmural gradient of repolarization caused the ECG characteristics and that major intraepicardial and transmural dispersion of action potentials (APs) initiated phase 2 reentry, premature ventricular activations, and tachyarrhythmias. Hypothermia enhanced the heterogeneity of the AP and promoted the origination of phase 2 reentry in the epicardium of the RVOT, but the prolonged AP duration frequently blocked reentry. Hyperthermia abbreviated the AP and facilitated the maintenance of reentry and tachyarrhythmias. Bradycardia promoted alternans in the phase 2 dome of the AP within the epicardium of the RVOT, resulting in TWA. The above phenomena were localized in the epicardium of the RVOT. Blockade of the transient outward current, I to , reduced AP heterogeneity and prevented arrhythmias in the BrS model. In addition, epicardial activation delay led to fragmented QRS, a risk marker of prognosis in BrS. Body surface mapping in patients with BrS supported these experimental findings. In conclusion, the AP heterogeneity within the epicardium of the RVOT contributes to the ECG characteristics, temperature sensitivity, TWA, and arrhythmias in BrS, and body surface mapping and fragmented QRS can be effective predictors of risk in patients with BrS.

Background— Conduction abnormalities serve as a substrate for ventricular fibrillation (VF) in patients with Brugada syndrome (BS). Signal-averaged electrograms can detect late potentials, but the significance of conduction abnormalities within the QRS complex is still unknown. The latter can present as multiple spikes within the QRS complex (fragmented QRS [f-QRS]). We hypothesized that f-QRS could indicate a substrate for VF and might predict a high risk of VF for patients with BS. Methods and Results— In study 1, we analyzed the incidence of f-QRS in 115 patients with BS (13 resuscitated from VF, 28 with syncope, and 74 asymptomatic). f-QRS was observed in 43% of patients, more often in the VF group (incidence of f-QRS: VF 85%, syncope 50%, and asymptomatic 34%, P SCN5A mutations occurred more often in patients with f-QRS (33%) than in patients without f-QRS (5%). In patients with syncope or VF, only 6% without f-QRS experienced VF during follow-up (43±25 months), but 58% of patients with f-QRS had recurrent syncope due to VF ( P Conclusions— f-QRS appears to be a marker for the substrate for spontaneous VF in BS and predicts patients at high risk of syncope.

Summary This Seminar presents the most recent information about the congenital long and short QT syndromes, emphasising the varied genotype-phenotype association in the ten different long QT syndromes and the five different short QT syndromes. Although uncommon, these syndromes serve as a Rosetta stone for the understanding of inherited ion-channel disorders leading to life-threatening cardiac arrhythmias. Ionic abnormal changes mainly affecting K + , Na + , or Ca 2+ currents, which either prolong or shorten ventricular repolarisation, can create a substrate of electrophysiological heterogeneity that predisposes to the development of ventricular tachyarrhythmias and sudden death. The understanding of the genetic basis of the syndromes is hoped to lead to genetic therapy that can restore repolarisation. Presently, symptomatic individuals are generally best treated with an implantable cardioverter defibrillator. Clinicians should be aware of these syndromes and realise that drugs, ischaemia, exercise, and emotions can precipitate sudden death in susceptible individuals.

The mechanisms responsible for ventricular tachyarrhythmia (VT) initiation have been studied extensively since the beginning of the last century. Multiple approaches including clinical observations; experiments in vivo, in vitro, and at cellular and genetic levels; theoretical analysis; and numeric simulations have been used. Most VTs are initiated by spontaneous discharge(s) at one or several sites in the ventricle, which can contribute to, or precipitate, reentry that then maintains the VT. Although most VTs occur in abnormal hearts such as in patients with coronary artery diseases, cardiomyopathy, abnormal ionic currents, or other problems, VT can also be functional, occurring in seemingly normal hearts when the conditions for its initiation are met. Because of the possible outcome of sudden cardiac death and the increasingly aging population in the industrialized countries, managing VT is a significant and important clinical task. Understanding the mechanisms of its initiation is one of the key steps in developing clinical strategies to prevent and treat VT. However, the mechanisms of VT initiation are still not fully understood and are the topic for a brief overview in this chapter.

Spatial Segmentation in the Simulation of Propagation. Introduction: Spatial segmentation is essential for the numerical simulation of excitation propagation in cardiac muscle. Methods and Results: This study evaluated the effects of spatial segmentation on action potential and on the velocity of propagation in a continuous one-dimensional model of cardiac muscle [intracellular and extracellular resistivities along (L) and transverse (T) to the muscle fibers: 402 ωcm (Ri, L), 3,620 ωcm (Ri, T), 48 ωcm (Re, L), and 126 ωcm (Re, T), J of Physiol 255:335-346, 1976) and either Luo-Rudy (L-R, Circ Res 68:1501-1526, 1991) or Beeler-Reuter (B-R, J Physiol 268:177-210, 1977) ionic currents. Related cable equations for active membrane are derived. Spatial segmentations of < 31.2 μm (L, L-R), < 11.5 μm (T, L-R), < 44.7 μm (L, B-R), and < 16.5 μm (T, B-R) were required for < 1% errors in the characteristic parameters of action potential. Similarly, spatial segmentations of < 54.5 μm (L, L-R), < 20.1 μm (T, L-R), < 84.3 μm (L, B-R), and < 31.2 μm (T, B-R) were required for < 1% errors in the velocity of conduction. Conclusion: In general, spatial segmentations of < 26.9% and < 50.8% of the space constant of a fully activated membrane gave < 1.0% errors in the characteristic parameters of action potential and in the velocity of propagation, respectively, for both membranes. The action potential duration was relatively insensitive to the spatial segmentation. Our analysis suggests that λfull is a better criterion for the selection of spatial segmentation in numerical simulation than the space constant of the resting membrane.

Background —Heterogeneous electrophysiological properties, which may be due in part to autonomic innervation, are important in the maintenance of atrial fibrillation (AF). We hypothesized that heterogeneous sympathetic denervation with phenol would create a milieu for sustained AF. Methods and Results —After the determination of baseline inducibility, 15 dogs underwent atrial epicardial phenol application and 11 underwent a sham procedure. After 2 weeks of recovery, the animals had repeat attempts at inducing AF and effective refractory period (ERP) testing. Epicardial maps were obtained to determine local AF cycle lengths. ERPs were determined at baseline and during sympathetic, vagal, and simultaneous vagal/sympathetic stimulation. Dogs then underwent PET imaging with either a sympathetic ([ 11 C]hydroxyephedrine, HED) or parasympathetic (5-[ 11 C]methoxybenzovesamicol, MOBV) nerve label. None of the animals had sustained AF (>60 minutes) at baseline. None of the sham dogs and 14 of 15 phenol dogs had sustained AF at follow-up. Sites to which phenol was applied had a significantly shorter ERP (136±17.6 ms) than those same sites in the sham controls (156±19.1 ms) ( P =0.01). Although there was no difference in the ERP change with either vagal or sympathetic stimulation alone between phenol and nonphenol sites, the percent decrease in ERP with simultaneous vagal/sympathetic stimulation was greater in the phenol sites (17±8%) than in the nonphenol sites (9±9%) ( P =0.01). There was a significantly increased dispersion of refractoriness (21±6.4 ms in the sham versus 58±14 ms in the phenol dogs, P =0.01) as well as dispersion of AF cycle length (49±10 ms in the sham versus 105±12 ms in the phenol dogs, P =0.0001). PET images demonstrated defects of HED uptake in the areas of phenol application, with no defect of MOBV uptake. Conclusions —Heterogeneous sympathetic atrial denervation with phenol facilitates sustained AF.

Torsades de pointes (TdP) is defined as a polymorphic ventricular tachycardia (VT) with a twisting QRS morphology associated with a prolonged QT interval and/or increased U wave amplitude in the ECG.1,2⇓ Patients with acquired or congenital long-QT syndrome (LQTS) can develop TdP that results in sudden cardiac death. Fifty or more drugs, both typical antiarrhythmic agents such as quinidine as well as other classes of drugs such as some antibiotics, affect membrane ionic currents, prolong the duration of the QT interval in the ECG, and have been associated with the acquired LQTS.3 Similarly, inherited genetic defects in the cardiac membrane ion channels can cause congenital LQTS. Some patients with apparent acquired LQTS may actually have mild congenital forms and remain asymptomatic until exposed to one of the drugs noted above.4 Congenital LQTS can be divided into multiple subtypes depending on the affected membrane ionic currents (eg, LQT1, LQT2, and LQT3 for alterations in I Ks, I Kr, and the inactivation process of I Na+). Due to the possible fatal outcome and involvement of many drugs, it is important to understand the mechanism of TdP, predict its occurrence, and manage it clinically. See p 770 It has been suggested that an early afterdepolarization (EAD) exceeding the activation threshold, arising from the endocardium (including the Purkinje network)5 or from the mid-myocardial region,6 could initiate TdP.2,7⇓ EADs have been observed experimentally in monophasic action potential recordings at multiple epicardial and endocardial sites.8 Increased dispersion of repolarization preceding the TdP induction has been noted in an isolated rabbit heart model of LQT2.9 Long intervals between activations, ie, slow rates, prolong the duration of the action potential and increase the transmural dispersion of …

Myocardium Immobilization for Repolarization Mapping. Introduction: Cytochalasin D In tissue bath superfusate inhihits the contraction of isolated thin trabeculae from canine right ventricle without affecting the intracellular action potential recorded with glass microelectrode. The purpose of this study was to test whether cytochalasin D could also be used to immobilize perfused wedges of ventricular muscle without affecting the action potential duration or propagation, and also to determine the optimal concentration and time duration of drug in the perfusate. Methods and Results: Using a membrane potential sensitive dye, di-4-ANEPPS, and a high-resolution photodiode optical mapping system at a rate of 1,000 frames/sec, we recorded action potentials on the transmural surface of arterially perfused wedges of muscle from the canine left ventricular free wall. We also recorded arterial pulse pressure as a surrogate for tissue contraction. Cytochalasin D at ≥ 20 μmol/L in the perfusate for ≥ 6 minutes reduced the arterial pulse pressure to approximately one tenth of its initial value and significantly reduced or eliminated motion artifacts in the action potentials. A sustained concentration of 10μmol/L cytochalasin D in the perfusate prevented contraction from recurring after the tissue was immobilized with an initial concentration of 25 μmol/L. Cytochalasin D had little effect on the action potential duration and on its transmural gradient, and did not slow the transmural velocity of excitation propagation. Conclusion: Cytochalasin D can be used to uncouple excitation and contraction in perfused canine cardiac muscle for the fluorescent-optical mapping of action potentials without affecting action potential duration or slowing transmural propagation.