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‣ Regulation of cardiac excitation and contraction by p21 activated kinase-1

Ke, Yunbo; Lei, Ming; Solaro, R. John
Fonte: PubMed Publicador: PubMed
Tipo: Artigo de Revista Científica
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Cardiac excitation and contraction are regulated by a variety of signaling molecules. Central to the regulatory scheme are protein kinases and phosphatases that carry out reversible phosphorylation of different effectors. The process of β-adrenergic stimulation mediated by cAMP dependent protein kinase (PKA) forms a well-known pathway considered as the most significant control mechanism in excitation and contraction as well as many other regulatory mechanisms in cardiac function. However, although dephosphorylation pathways are critical to these regulatory processes, signaling to phosphatases is relatively poorly understood. Emerging evidence indicates that regulation of phosphatases, which dampen the effect of β-adrenergic stimulation, is also important. We review here functional studies of p21 activated kinase-1 (Pak1) and its potential role as an upstream signal for protein phosphatase PP2A in the heart. Pak1 is a serine/threonine protein kinase directly activated by the small GTPases Cdc42 and Rac1. Pak1 is highly expressed in different regions of the heart and modulates the activities of ion channels, sarcomeric proteins, and other phosphoproteins through up-regulation of PP2A activity. Coordination of Pak1 and PP2A activities is not only potentially involved in regulation of normal cardiac function...

‣ Ionic mechanisms of cellular electrical and mechanical abnormalities in Brugada syndrome

Dong, Min; Niklewski, Paul J.; Wang, Hong-Sheng
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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The Brugada syndrome (BrS) is a right ventricular (RV) arrhythmia that is responsible for up to 12% of sudden cardiac deaths. The aims of our study were to determine the cellular mechanisms of the electrical abnormality in BrS and the potential basis of the RV contractile abnormality observed in the syndrome. Tetrodotoxin was used to reduce cardiac Na+ current (INa) to mimic a BrS-like setting in canine ventricular myocytes. Moderate reduction (<50%) of INa with tetrodotoxin resulted in all-or-none repolarization in a fraction of RV epicardial myocytes. Dynamic clamp and modeling show that reduction of INa shifts the action potential (AP) duration-transient outward current (Ito) density curve to the left and has a biphasic effect on AP duration. In the presence of a large Ito, INa reduction either prolongs or collapses the AP, depending on the exact density of Ito. These repolarization changes reduce Ca2+ influx and sarcoplasmic reticulum load, resulting in marked attenuation of myocyte contraction and Ca2+ transient in RV epicardial myocytes. We conclude that INa reduction alters repolarization by reducing the threshold for Ito-induced all-or-none repolarization. These cellular electrical changes suppress myocyte excitation-contraction coupling and contraction and may be a contributing factor to the contractile abnormality of the RV wall in BrS.

‣ Quantification of calsequestrin 2 (CSQ2) in sheep cardiac muscle and Ca2+-binding protein changes in CSQ2 knockout mice

Murphy, Robyn M.; Mollica, Janelle P.; Beard, Nicole A.; Knollmann, Bjorn C.; Lamb, Graham D.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Calsequestrin 2 (CSQ2) is generally regarded as the primary Ca2+-buffering molecule present inside the sarcoplasmic reticulum (SR) in cardiac cells, but findings from CSQ2 knockout experiments raise major questions about its role and necessity. This study determined the absolute amount of CSQ2 present in cardiac ventricular muscle to gauge its likely influence on SR free Ca2+ concentration ([Ca2+]) and maximal Ca2+ capacity. Ventricular tissue from hearts of freshly killed sheep was examined by SDS-PAGE without any fractionation, and CSQ2 was detected by Western blotting; this method avoided the >90% loss of CSQ2 occurring with usual fractionation procedures. Band intensities were compared against those for purified CSQ2 run on the same blots. Fidelity of quantification was verified by demonstrating that CSQ2 added to homogenates was detected with equal efficacy as purified CSQ2 alone. Ventricular tissue from sheep (n = 8) contained 24 ± 2 μmol CSQ2/kg wet wt. Total Ca2+ content of the ventricular tissue, measured by atomic absorption spectroscopy, was 430 ± 20 μmol/kg (with SR Ca2+ likely <250 μmol/kg) and displayed a linear correlation with CSQ2 content, with gradient of ∼10 Ca2+ per CSQ2. The large amount of CSQ2 bestows the SR with a high theoretical maximal Ca2+-binding capacity (∼1 mmol Ca2+/kg ventricular tissue...

‣ Phorbol ester and endothelin-1 alter functional expression of Na+/Ca2+ exchange, K+, and Ca2+ currents in cultured neonatal rat myocytes

Puglisi, José L.; Yuan, Weilong; Timofeyev, Valeriy; Myers, Richard E.; Chiamvimonvat, Nipavan; Samarel, Allen M.; Bers, Donald M.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Endothelin-1 (ET-1) and activation of protein kinase C (PKC) have been implicated in alterations of myocyte function in cardiac hypertrophy and heart failure. Changes in cellular Ca2+ handling and electrophysiological properties also occur in these states and may contribute to mechanical dysfunction and arrhythmias. While ET-1 or PKC stimulation induces cellular hypertrophy in cultured neonatal rat ventricular myocytes (NRVMs), a system widely used in studies of hypertrophic signaling, there is little data about electrophysiological changes. Here we studied the effects of ET-1 (100 nM) or the PKC activator phorbol 12-myristate 13-acetate (PMA, 1 μM) on ionic currents in NRVMs. The acute effects of PMA or ET-1 (≤30 min) were small or insignificant. However, PMA or ET-1 exposure for 48–72 h increased cell capacitance by 100 or 25%, respectively, indicating cellular hypertrophy. ET-1 also slightly increased Ca2+ current density (T and L type). Na+/Ca2+ exchange current was increased by chronic pretreatment with either PMA or ET-1. In contrast, transient outward and delayed rectifier K+ currents were strongly downregulated by PMA or ET-1 pretreatment. Inward rectifier K+ current tended toward a decrease at larger negative potential...

‣ Regional increase of extracellular potassium leads to electrical instability and reentry occurrence through the spatial heterogeneity of APD restitution

Sidorov, Veniamin Y.; Uzelac, Ilija; Wikswo, John P.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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The heterogeneities of electrophysiological properties of cardiac tissue are the main factors that control both arrhythmia induction and maintenance. Although the local increase of extracellular potassium ([K+]o) due to coronary occlusion is a well-established metabolic response to acute ischemia, the role of local [K+]o heterogeneity in phase 1a arrhythmias has yet to be determined. In this work, we created local [K+]o heterogeneity and investigated its role in fast pacing response and arrhythmia induction. The left marginal vein of a Langendorff-perfused rabbit heart was cannulated and perfused separately with solutions containing 4, 6, 8, 10, and 12 mM of K+. The fluorescence dye was utilized to map the voltage distribution. We tested stimulation rates, starting from 400 ms down to 120 ms, with steps of 5–50 ms. We found that local [K+]o heterogeneity causes action potential (AP) alternans, 2:1 conduction block, and wave breaks. The effect of [K+]o heterogeneity on electrical stability and vulnerability to arrhythmia induction was largest during regional perfusion with 10 mM of K+. We detected three concurrent dynamics: normally propagating activation when excitation waves spread over tissue perfused with normal K+, alternating 2:2 rhythm near the border of [K+]o heterogeneity...

‣ Ca2+ sensitization of cardiac myofilament proteins contributes to exercise training-enhanced myocardial function in a porcine model of chronic occlusion

Sarin, Vandana; Muthuchamy, Mariappan; Heaps, Cristine L.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Exercise training has been shown to improve cardiac dysfunction in both patients and animal models of coronary artery disease; however, the underlying cellular and molecular mechanisms have not been completely understood. We hypothesized that exercise training would improve force generation in the myocardium distal to chronic coronary artery occlusion via altered intracellular Ca2+ concentration ([Ca2+]i) cycling and/or Ca2+ sensitization of myofilaments. Ameroid occluders were surgically placed around the proximal left circumflex coronary artery of adult female Yucatan pigs. Twenty-two weeks postoperatively, the myocardium was isolated from nonoccluded (left anterior descending artery dependent) and collateral-dependent (formerly left circumflex coronary artery dependent) regions of sedentary (pen confined) and exercise-trained (treadmill run, 5 days/wk for 14 wk) pigs. Force measurements in myocardial strips showed that the percent change in force at stimulation frequencies of 3 and 4 Hz relative to 1 Hz was significantly higher in exercise-trained pigs compared with sedentary pigs. β-Adrenergic stimulation with dobutamine significantly improved force kinetics in myocardial strips of sedentary but not exercise-trained pigs at 1 Hz. Additionally...

‣ Post-transcriptional silencing of SCN1B and SCN2B genes modulates late sodium current in cardiac myocytes from normal dogs and dogs with chronic heart failure

Mishra, Sudhish; Undrovinas, Nidas A.; Maltsev, Victor A.; Reznikov, Vitaliy; Sabbah, Hani N.; Undrovinas, Albertas
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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The emerging paradigm for Na+ current in heart failure (HF) is that its transient component (INaT) responsible for the action potential (AP) upstroke is decreased, whereas the late component (INaL) involved in AP plateau is augmented. Here we tested whether Navβ1- and Navβ2-subunits can modulate INaL parameters in normal and failing ventricular cardiomyocytes (VCMs). Chronic HF was produced in nine dogs by multiple sequential coronary artery microembolizations, and six dogs served as a control. INa and APs were measured by the whole cell and perforated patch-clamp in freshly isolated and cultured VCMs, respectively. INaL was augmented with slower decay in HF VCMs compared with normal heart VCMs, and these properties remained unchanged within 5 days of culture. Post-transcriptional silencing SCN1B and SCN2B were achieved by virally delivered short interfering RNA (siRNA) specific to Navβ1 and Navβ2. The delivery and efficiency of siRNA were evaluated by green fluorescent protein expression, by the real-time RT-PCR, and Western blots, respectively. Five days after infection, the levels of mRNA and protein for Navβ1 and Navβ2 were reduced by >80%, but mRNA and protein of Nav1.5, as well as INaT, remained unchanged in HF VCMs. Navβ1-siRNA reduced INaL density and accelerated INaL two-exponential decay...

‣ The role of dynamic instability and wavelength in arrhythmia maintenance as revealed by panoramic imaging with blebbistatin vs. 2,3-butanedione monoxime

Lou, Qing; Li, Wenwen; Efimov, Igor R.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Unlike other excitation-contraction uncouplers, blebbistatin has few electrophysiological side effects and has gained increasing acceptance as an excitation-contraction uncoupler in optical mapping experiments. However, the possible role of blebbistatin in ventricular arrhythmia has hitherto been unknown. Furthermore, experiments with blebbistatin and 2,3-butanedione monoxime (BDM) offer an opportunity to assess the contribution of dynamic instability and wavelength of impulse propagation to the induction and maintenance of ventricular arrhythmia. Recordings of monophasic action potentials were used to assess effects of blebbistatin in Langendorff-perfused rabbit hearts (n = 5). Additionally, panoramic optical mapping experiments were conducted in rabbit hearts (n = 7) that were sequentially perfused with BDM, then washed out, and subsequently perfused with blebbistatin. The susceptibility to arrhythmia was investigated using a shock-on-T protocol. We found that 1) application of blebbistatin did not change action potential duration (APD) restitution; 2) in contrast to blebbistatin, BDM flattened APD restitution curve and reduced the wavelength; and 3) incidence of sustained arrhythmia was much lower under blebbistatin than under BDM (2/123 vs. 23/99). While arrhythmias under BDM were able to stabilize...

‣ Constitutive overexpression of phosphomimetic phospholemman S68E mutant results in arrhythmias, early mortality, and heart failure: potential involvement of Na+/Ca2+ exchanger

Song, Jianliang; Gao, Erhe; Wang, JuFang; Zhang, Xue-Qian; Chan, Tung O.; Koch, Walter J.; Shang, Xiying; Joseph, Jeffrey I.; Peterson, Blaise Z.; Feldman, Arthur M.; Cheung, Joseph Y.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Expression and activity of cardiac Na+/Ca2+ exchanger (NCX1) are altered in many disease states. We engineered mice in which the phosphomimetic phospholemman S68E mutant (inhibits NCX1 but not Na+-K+-ATPase) was constitutively overexpressed in a cardiac-specific manner (conS68E). At 4–6 wk, conS68E mice exhibited severe bradycardia, ventricular arrhythmias, increased left ventricular (LV) mass, decreased cardiac output (CO), and ∼50% mortality compared with wild-type (WT) littermates. Protein levels of NCX1, calsequestrin, ryanodine receptor, and α1- and α2-subunits of Na+-K+-ATPase were similar, but sarco(endo)plasmic reticulum Ca2+-ATPase was lower, whereas L-type Ca2+ channels were higher in conS68E hearts. Resting membrane potential and action potential amplitude were similar, but action potential duration was dramatically prolonged in conS68E myocytes. Diastolic intracellular Ca2+ ([Ca2+]i) was higher, [Ca2+]i transient and maximal contraction amplitudes were lower, and half-time of [Ca2+]i transient decline was longer in conS68E myocytes. Intracellular Na+ reached maximum within 3 min after isoproterenol addition, followed by decline in WT but not in conS68E myocytes. Na+/Ca2+ exchange, L-type Ca2+, Na+-K+-ATPase, and depolarization-activated K+ currents were decreased in conS68E myocytes. At 22 wk...

‣ Regional increase in extracellular potassium can be arrhythmogenic due to nonuniform muscle contraction in rat ventricular muscle

Miura, Masahito; Hattori, Taiki; Murai, Naomi; Nagano, Tsuyoshi; Nishio, Taichi; Boyden, Penelope A.; Shindoh, Chiyohiko
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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In the ischemic myocardium, extracellular potassium ([K+]o) increases to ≥20 mmol/l. To determine how lethal arrhythmias occur during ischemia, we investigated whether the increased spatial pattern of [K+]o, i.e., a regional or a global increase, affects the incidence of arrhythmias. Force, sarcomere length, membrane potential, and nonuniform intracellular Ca2+ ([Ca2+]i) were measured in rat ventricular trabeculae. A “regional” or “global” increase in [K+]o was produced by exposing a restricted region of muscle to a jet of 30 mmol/l KCl or by superfusing trabeculae with a solution containing 30 mmol/l KCl, respectively. The increase in [Ca2+]i (CaCW) during Ca2+ waves was measured (24°C, 3.0 mmol/l [Ca2+]o). A regional increase in [K+]o caused nonuniform [Ca2+]i and contraction. In the presence of isoproterenol, the regional increase in [K+]o induced sustained arrhythmias in 10 of 14 trabeculae, whereas the global increase did not induce such arrhythmias. During sustained arrhythmias, Ca2+ surged within the jet-exposed region. In the absence of isoproterenol, the regional increase in [K+]o increased CaCW, whereas the global increase decreased it. This increase in CaCW with the regional increase in [K+]o was not suppressed by 100 μmol/l streptomycin...

‣ Refractoriness of sarcoplasmic reticulum Ca2+ release determines Ca2+ alternans in atrial myocytes

Shkryl, Vyacheslav M.; Maxwell, Joshua T.; Domeier, Timothy L.; Blatter, Lothar A.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Cardiac alternans is a recognized risk factor for cardiac arrhythmia and sudden cardiac death. At the cellular level, Ca2+ alternans appears as cytosolic Ca2+ transients of alternating amplitude at regular beating frequency. Cardiac alternans is a multifactorial process but has been linked to disturbances in intracellular Ca2+ regulation. In atrial myocytes, we tested the role of voltage-gated Ca2+ current, sarcoplasmic reticulum (SR) Ca2+ load, and restitution properties of SR Ca2+ release for the occurrence of pacing-induced Ca2+ alternans. Voltage-clamp experiments revealed that peak Ca2+ current was not affected during alternans, and alternans of end-diastolic SR Ca2+ load, evaluated by application of caffeine or measured directly with an intra-SR fluorescent Ca2+ indicator (fluo-5N), were not a requirement for cytosolic Ca2+ alternans. Restitution properties and kinetics of refractoriness of Ca2+ release after activation during alternans were evaluated by four different approaches: measurements of 1) the delay (latency) of occurrence of spontaneous global Ca2+ releases and 2) Ca2+ spark frequency, both during rest after a large and small alternans Ca2+ transient; 3) the magnitude of premature action potential-induced Ca2+ transients after a large and small beat; and 4) the efficacy of a photolytically induced Ca2+ signal (Ca2+ uncaging from DM-nitrophen) to trigger additional Ca2+ release during alternans. The results showed that the latency of global spontaneous Ca2+ release was prolonged and Ca2+ spark frequency was decreased after the large Ca2+ transient during alternans. Furthermore...

‣ Mapping cardiac surface mechanics with structured light imaging

Laughner, Jacob I.; Zhang, Song; Li, Hao; Shao, Connie C.; Efimov, Igor R.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Cardiovascular disease often manifests as a combination of pathological electrical and structural heart remodeling. The relationship between mechanics and electrophysiology is crucial to our understanding of mechanisms of cardiac arrhythmias and the treatment of cardiac disease. While several technologies exist for describing whole heart electrophysiology, studies of cardiac mechanics are often limited to rhythmic patterns or small sections of tissue. Here, we present a comprehensive system based on ultrafast three-dimensional (3-D) structured light imaging to map surface dynamics of whole heart cardiac motion. Additionally, we introduce a novel nonrigid motion-tracking algorithm based on an isometry-maximizing optimization framework that forms correspondences between consecutive 3-D frames without the use of any fiducial markers. By combining our 3-D imaging system with nonrigid surface registration, we are able to measure cardiac surface mechanics at unprecedented spatial and temporal resolution. In conclusion, we demonstrate accurate cardiac deformation at over 200,000 surface points of a rabbit heart recorded at 200 frames/s and validate our results on highly contrasting heart motions during normal sinus rhythm, ventricular pacing...

‣ Regulation of cardiac alternans by β-adrenergic signaling pathways

Florea, Stela M.; Blatter, Lothar A.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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In cat atrial myocytes, β-adrenergic receptor (β-AR) stimulation exerts profound effects on excitation-contraction coupling and cellular Ca2+ cycling that are mediated by β1- and β2-AR subtypes coupled to G proteins (Gs and Gi). In this study, we determined the effects of β-AR stimulation on pacing-induced Ca2+ alternans. Ca2+ alternans was recorded from single cat atrial myocytes with the fluorescent Ca2+ indicator indo-1. Stable Ca2+ alternans occurred at an average pacing frequency of 1.7 Hz at room temperature with a mean alternans ratio of 0.43. Nonselective β-AR stimulation as well as selective stimulation of β1/Gs, β2/Gs + Gi, and β2/Gs coupled pathways all abolished pacing-induced Ca2+ alternans. β1-AR stimulation abolished alternans through stimulation of PKA and Ca2+/calmodulin-dependent protein kinase II, whereas β2-AR stimulation exclusively involved PKA and was mediated via Gs, whereas a known second pathway in cat atrial myocytes acting through Gi and nitric oxide production was not involved in alternans regulation. Inhibition of various mitochondrial functions (dissipation of the mitochondrial membrane potential or inhibition of mitochondrial F1/F0-ATP synthase, mitochondrial Ca2+ uptake via the mitochondrial Ca2+ uniporter...

‣ Effects of mitochondrial uncoupling on Ca2+ signaling during excitation-contraction coupling in atrial myocytes

Zima, Aleksey V.; Pabbidi, Malikarjuna R.; Lipsius, Stephen L.; Blatter, Lothar A.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Mitochondria play an important role in intracellular Ca2+ concentration ([Ca2+]i) regulation in the heart. We studied sarcoplasmic reticulum (SR) Ca2+ release in cat atrial myocytes during depolarization of mitochondrial membrane potential (ΔΨm) induced by the protonophore FCCP. FCCP caused an initial decrease of action potential-induced Ca2+ transient amplitude and frequency of spontaneous Ca2+ waves followed by partial recovery despite partially depleted SR Ca2+ stores. In the presence of oligomycin, FCCP only exerted a stimulatory effect on Ca2+ transients and Ca2+ wave frequency, suggesting that the inhibitory effect of FCCP was mediated by ATP consumption through reverse-mode operation of mitochondrial F1F0-ATPase. ΔΨm depolarization was accompanied by cytosolic acidification, increases of diastolic [Ca2+]i, intracellular Na+ concentration ([Na+]i), and intracellular Mg2+ concentration ([Mg2+]i), and a decrease of intracellular ATP concentration ([ATP]i); however, glycolytic ATP production partially compensated for the exhaustion of mitochondrial ATP supplies. In conclusion, the initial inhibition of Ca2+ transients and waves resulted from suppression of ryanodine receptor SR Ca2+ release channel activity by a decrease in [ATP]...

‣ Modulation of ventricular transient outward K+ current by acidosis and its effects on excitation-contraction coupling

Saegusa, Noriko; Garg, Vivek; Spitzer, Kenneth W.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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The contribution of transient outward current (Ito) to changes in ventricular action potential (AP) repolarization induced by acidosis is unresolved, as is the indirect effect of these changes on calcium handling. To address this issue we measured intracellular pH (pHi), Ito, L-type calcium current (ICa,L), and calcium transients (CaTs) in rabbit ventricular myocytes. Intracellular acidosis [pHi 6.75 with extracellular pH (pHo) 7.4] reduced Ito by ∼50% in myocytes with both high (epicardial) and low (papillary muscle) Ito densities, with little effect on steady-state inactivation and activation. Of the two candidate α-subunits underlying Ito, human (h)Kv4.3 and hKv1.4, only hKv4.3 current was reduced by intracellular acidosis. Extracellular acidosis (pHo 6.5) shifted Ito inactivation toward less negative potentials but had negligible effect on peak current at +60 mV when initiated from −80 mV. The effects of low pHi-induced inhibition of Ito on AP repolarization were much greater in epicardial than papillary muscle myocytes and included slowing of phase 1, attenuation of the notch, and elevation of the plateau. Low pHi increased AP duration in both cell types, with the greatest lengthening occurring in epicardial myocytes. The changes in epicardial AP repolarization induced by intracellular acidosis reduced peak ICa...

‣ Cardiac hypertrophy associated with impaired regulation of cardiac ryanodine receptor by calmodulin and S100A1

Yamaguchi, Naohiro; Chakraborty, Asima; Huang, Tai-Qin; Xu, Le; Gomez, Angela C.; Pasek, Daniel A.; Meissner, Gerhard
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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The cardiac ryanodine receptor (RyR2) is inhibited by calmodulin (CaM) and S100A1. Simultaneous substitution of three amino acid residues (W3587A, L3591D, F3603A; RyR2ADA) in the CaM binding domain of RyR2 results in loss of CaM inhibition at submicromolar (diastolic) and micromolar (systolic) Ca2+, cardiac hypertrophy, and heart failure in Ryr2ADA/ADA mice. To address whether cardiac hypertrophy results from the elimination of CaM and S100A1 inhibition at diastolic or systolic Ca2+, a mutant mouse was generated with a single RyR2 amino acid substitution (L3591D; RyR2D). Here we report that in single-channel measurements RyR2-L3591D isolated from Ryr2D/D hearts lost CaM inhibition at diastolic Ca2+ only, whereas S100A1 regulation was eliminated at both diastolic and systolic Ca2+. In contrast to the ∼2-wk life span of Ryr2ADA/ADA mice, Ryr2D/D mice lived longer than 1 yr. Six-month-old Ryr2D/D mice showed a 9% increase in heart weight-to-body weight ratio, modest changes in cardiac morphology, and a twofold increase in atrial natriuretic peptide mRNA levels compared with wild type. After 4-wk pressure overload with transverse aortic constriction, heart weight-to-body weight ratio and atrial natriuretic peptide mRNA levels increased and echocardiography showed changes in heart morphology of Ryr2D/D mice compared with sham-operated mice. Collectively...

‣ Novel role of transient receptor potential vanilloid 2 in the regulation of cardiac performance

Rubinstein, Jack; Lasko, Valerie M.; Koch, Sheryl E.; Singh, Vivek P.; Carreira, Vinicius; Robbins, Nathan; Patel, Amit R.; Jiang, Min; Bidwell, Philip; Kranias, Evangelia G.; Jones, W. Keith; Lorenz, John N.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Transient receptor potential cation channels have been implicated in the regulation of cardiovascular function, but only recently has our laboratory described the vanilloid-2 subtype (TRPV2) in the cardiomyocyte, though its exact mechanism of action has not yet been established. This study tests the hypothesis that TRPV2 plays an important role in regulating myocyte contractility under physiological conditions. Therefore, we measured cardiac and vascular function in wild-type and TRPV2−/− mice in vitro and in vivo and found that TRPV2 deletion resulted in a decrease in basal systolic and diastolic function without affecting loading conditions or vascular tone. TRPV2 stimulation with probenecid, a relatively selective TRPV2 agonist, caused an increase in both inotropy and lusitropy in wild-type mice that was blunted in TRPV2−/− mice. We examined the mechanism of TRPV2 inotropy/lusitropy in isolated myocytes and found that it modulates Ca2+ transients and sarcoplasmic reticulum Ca2+ loading. We show that the activity of this channel is necessary for normal cardiac function and that there is increased contractility in response to agonism of TRPV2 with probenecid.

‣ The absence of insulin signaling in the heart induces changes in potassium channel expression and ventricular repolarization

Lopez-Izquierdo, Angelica; Pereira, Renata O.; Wende, Adam R.; Punske, Bonnie B.; Abel, E. Dale; Tristani-Firouzi, Martin
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Diabetes mellitus increases the risk for cardiac dysfunction, heart failure, and sudden death. The wide array of neurohumoral changes associated with diabetes pose a challenge to understanding the roles of specific pathways that alter cardiac function. Here, we use a mouse model with cardiomyocyte-restricted deletion of insulin receptors (CIRKO, cardiac-specific insulin receptor knockout) to study the specific effects of impaired cardiac insulin signaling on ventricular repolarization, independent of the generalized metabolic derangements associated with diabetes. Impaired insulin action caused a reduction in mRNA and protein expression of several key K+ channels that dominate ventricular repolarization. Specifically, components of transient outward K+ current fast component (Ito,fast; Kv4.2 and KChiP2) were reduced, consistent with a reduction in the amplitude of Ito,fast in isolated left ventricular CIRKO myocytes, compared with littermate controls. The reduction in Ito,fast resulted in ventricular action potential prolongation and prolongation of the QT interval on the surface ECG. These results support the notion that the lack of insulin signaling in the heart is sufficient to cause the repolarization abnormalities described in other animal models of diabetes.

‣ Adverse perinatal environment contributes to altered cardiac development and function

Velten, Markus; Gorr, Matthew W.; Youtz, Dane J.; Velten, Christina; Rogers, Lynette K.; Wold, Loren E.
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Epidemiological observations report an association between intrauterine growth restriction (IUGR) and cardiovascular diseases. Systemic maternal inflammation is the most common stress during pregnancy, leading to IUGR. We hypothesized that perinatal inflammation and hyperoxygenation induce discernible alterations in cardiomyocyte contractility and calcium signaling, causing early cardiac dysfunction. Pregnant C3H/HeN mice were injected with LPS or saline on embryonic day 16. Newborn mice were placed in 85% O2 or room air (RA) for 14 days. Pups born to LPS-injected dams had reduced birth weight. Echocardiographic measurements revealed that in vivo LV function was compromised in LPS/O2 mice as early as 3 days of life. Isolated cardiomyocytes from LPS/O2 mice at day 14 exhibited decreased sarcomere fractional shortening, along with decreased time-to-90% peak shortening. Calcium transient amplitude was greatest in LPS/O2 mice. SERCA2a mRNA and protein levels were increased and phospholamban mRNA levels were decreased in LPS/O2 mice. Phosphorylation of phospholamban was increased, along with Sorcin mRNA levels in LPS/O2 mice. Combined exposure to perinatal inflammation and hyperoxia resulted in growth restriction, in vivo and in vitro cardiac dysfunction...

‣ Cardiac tissue slices: preparation, handling, and successful optical mapping

Wang, Ken; Lee, Peter; Mirams, Gary R.; Sarathchandra, Padmini; Borg, Thomas K.; Gavaghan, David J.; Kohl, Peter; Bollensdorff, Christian
Fonte: American Physiological Society Publicador: American Physiological Society
Tipo: Artigo de Revista Científica
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Cardiac tissue slices are becoming increasingly popular as a model system for cardiac electrophysiology and pharmacology research and development. Here, we describe in detail the preparation, handling, and optical mapping of transmembrane potential and intracellular free calcium concentration transients (CaT) in ventricular tissue slices from guinea pigs and rabbits. Slices cut in the epicardium-tangential plane contained well-aligned in-slice myocardial cell strands (“fibers”) in subepicardial and midmyocardial sections. Cut with a high-precision slow-advancing microtome at a thickness of 350 to 400 μm, tissue slices preserved essential action potential (AP) properties of the precutting Langendorff-perfused heart. We identified the need for a postcutting recovery period of 36 min (guinea pig) and 63 min (rabbit) to reach 97.5% of final steady-state values for AP duration (APD) (identified by exponential fitting). There was no significant difference between the postcutting recovery dynamics in slices obtained using 2,3-butanedione 2-monoxime or blebistatin as electromechanical uncouplers during the cutting process. A rapid increase in APD, seen after cutting, was caused by exposure to ice-cold solution during the slicing procedure...