Myocyte Shortening Research Articles

Measurement of Strain in Cardiac Myocytes at Micrometer Scale Based on Rapid Scanning Confocal Microscopy and Non-Rigid Image Registration.

Measurement of cell shortening is an important technique for assessment of physiology and pathophysiology of cardiac myocytes. Many types of heart disease are associated with decreased myocyte shortening, which is commonly caused by structural and functional remodeling. Here, we present a new approach for local measurement of 2-dimensional strain within cells at high spatial resolution. The approach applies non-rigid image registration to quantify local displacements and Cauchy strain in images of cells undergoing contraction. We extensively evaluated the approach using synthetic cell images and image sequences from rapid scanning confocal microscopy of fluorescently labeled isolated myocytes from the left ventricle of normal and diseased canine heart. Application of the approach yielded a comprehensive description of cellular strain including novel measurements of transverse strain and spatial heterogeneity of strain. Quantitative comparison with manual measurements of strain in image sequences indicated reliability of the developed approach. We suggest that the developed approach provides researchers with a novel tool to investigate contractility of cardiac myocytes at subcellular scale. In contrast to previously introduced methods for measuring cell shorting, the developed approach provides comprehensive information on the spatio-temporal distribution of 2-dimensional strain at micrometer scale.

New Findings on the Effects of Tannic Acid: Inhibition of L-Type Calcium Channels, Calcium Transient and Contractility in Rat Ventricular Myocytes.

Tannic acid (TA) is a group of water-soluble polyphenolic compounds that occur mainly in plant-derived feeds, food grains and fruits. Many studies have explored its biomedical properties, such as anticancer, antibacterial, antimutagenic, antioxidant, antidiabetic, antiinflammatory and antihypertensive activities. However, the effects of TA on the L-type Ca(2+) current (ICa-L) of cardiomyocytes remain undefined. The present study examined the effects of TA on ICa-L using the whole-cell patch-clamp technique and on intracellular Ca(2+) handling and cell contractility in rat ventricular myocytes with the aid of a video-based edge detection system. Exposure to TA resulted in a concentration- and voltage-dependent blockade of ICa-L, with the half maximal inhibitory concentration of 1.69 μM and the maximal inhibitory effect of 46.15%. Moreover, TA significantly inhibited the amplitude of myocyte shortening and peak value of Ca(2+) transient and increased the time to 10% of the peak. These findings provide new experimental evidence for the cellular mechanism of action of TA and may help to expand clinical treatments for cardiovascular disease.

Persistent increases in Ca(2+) influx through Cav1.2 shortens action potential and causes Ca(2+) overload-induced afterdepolarizations and arrhythmias.

Persistent elevation of Ca(2+) influx due to prolongation of the action potential (AP), chronic activation of the β-adrenergic system and molecular remodeling occurs in stressed and diseased hearts. Increases in Ca(2+) influx are usually linked to prolonged myocyte action potentials and arrhythmias. However, the contribution of chronic enhancement of Cav1.2 activity on cardiac electrical remodeling and arrhythmogenicity has not been completely defined and is the subject of this study. Chronically increased Cav1.2 activity was produced with a cardiac specific, inducible double transgenic (DTG) mouse system overexpressing the β2a subunit of Cav (Cavβ2a). DTG myocytes had increased L-type Ca(2+) current (ICa-L), myocyte shortening, and Ca(2+) transients. DTG mice had enhanced cardiac performance, but died suddenly and prematurely. Telemetric electrocardiograms revealed shortened QT intervals in DTG mice. The action potential duration (APD) was shortened in DTG myocytes due to significant increases of potassium currents and channel abundance. However, shortened AP in DTG myocytes did not fully limit excess Ca(2+) influx and increased the peak and tail ICa-L. Enhanced ICa promoted sarcoplasmic reticulum (SR) Ca(2+) overload, diastolic Ca(2+) sparks and waves, and increased NCX activity, causing increased occurrence of early and delayed afterdepolarizations (EADs and DADs) that may contribute to premature ventricular beats and ventricular tachycardia. AV blocks that could be related to fibrosis of the AV node were also observed. Our study suggests that increasing ICa-L does not necessarily result in AP prolongation but causes SR Ca(2+) overload and fibrosis of AV node and myocardium to induce cellular arrhythmogenicity, arrhythmias, and conduction abnormalities.

Ketamine attenuates the Na+-dependent Ca2+ overload in rabbit ventricular myocytes in vitro by inhibiting late Na+ and L-type Ca2+ currents.

Intracellular Ca(2+) ([Ca(2+)]i) overload occurs in myocardial ischemia. An increase in the late sodium current (INaL) causes intracellular Na(+) overload and subsequently [Ca(2+)]i overload via the reverse-mode sodium-calcium exchanger (NCX). Thus, inhibition of INaL is a potential therapeutic target for cardiac diseases associated with [Ca(2+)]i overload. The aim of this study was to investigate the effects of ketamine on Na(+)-dependent Ca(2+) overload in ventricular myocytes in vitro. Ventricular myocytes were enzymatically isolated from hearts of rabbits. INaL, NCX current (INCX) and L-type Ca(2+) current (ICaL) were recorded using whole-cell patch-clamp technique. Myocyte shortening and [Ca(2+)]i transients were measured simultaneously using a video-based edge detection and dual excitation fluorescence photomultiplier system. Ketamine (20, 40, 80 μmol/L) inhibited INaL in a concentration-dependent manner. In the presence of sea anemone toxin II (ATX, 30 nmol/L), INaL was augmented by more than 3-fold, while ketamine concentration-dependently suppressed the ATX-augmented INaL. Ketamine (40 μmol/L) also significantly suppressed hypoxia or H2O2-induced enhancement of INaL. Furthermore, ketamine concentration-dependently attenuated ATX-induced enhancement of reverse-mode INCX. In addition, ketamine (40 μmol/L) inhibited ICaL by 33.4%. In the presence of ATX (3 nmol/L), the rate and amplitude of cell shortening and relaxation, the diastolic [Ca(2+)]i, and the rate and amplitude of [Ca(2+)]i rise and decay were significantly increased, which were reverted to control levels by tetrodotoxin (TTX, 2 μmol/L) or by ketamine (40 μmol/L). Ketamine protects isolated rabbit ventricular myocytes against [Ca(2+)]i overload by inhibiting INaL and ICaL.

Crocin, a carotenoid component of Crocus cativus, exerts inhibitory effects on L-type Ca(2+) current, Ca(2+) transient, and contractility in rat ventricular myocytes.

Crocin, a carotenoid component of Crocus sativus L. belonging to the Iridaceae family, has demonstrated cardioprotective effects. To investigate the cellular mechanisms of these cardioprotective effects, here we studied the influence of crocin on L-type Ca(2+)current (I(Ca-L)), intracellular Ca(2+) ([Ca(2+)]i), and contraction of isolated rat cardiomyocytes by using the whole-cell patch-clamp technique and video-based edge detection and dual excitation fluorescence photomultiplier systems. Crocin inhibited I(Ca-L) in a concentration-dependent manner with the half-maximal inhibitory concentration (IC50) of 45 μmol/L and the maximal inhibitory effect of 72.195% ± 1.54%. Neither current-voltage relationship of I(Ca-L), reversal potential of I(Ca-L), nor the activation/inactivation of I(Ca-L) was significantly changed. Crocin at 1 μmol/L reduced cell shortening by 44.64% ± 2.12% and the peak value of the Ca(2+) transient by 23.66% ± 4.52%. Crocin significantly reduced amplitudes of myocyte shortening and [Ca(2+)]i with an increase in the time to reach 10% of the peak (Tp) and a decrease in the time to 10% of the baseline (Tr). Thus, the cardioprotective effects of crocin may be attributed to the attenuation of [Ca(2+)]i through the inhibition of I(Ca-L) in rat cardiomyocytes and negative inotropic effects on myocardial contractility.

Palmitic Acid Regulation of Cardiac Inotropy and Calcium Handling in Healthy and Angiotensin II‐Induced Hypertensive Rat

Background and aimPalmitic acid (PA) is a predominant metabolic substrate. However, the effect of PA on cardiac contraction and underlying mechanisms in healthy and diseased heart remain unclear. Here, we analyzed PA‐regulation of myocyte contraction in left ventricular (LV) myocytes from sham and angiotensin II (Ang II)‐induced hypertensive rats.ResultsOur results showed that PA (100 μM) increased the amplitude of sarcomere shortening in LV myocytes (field stimulation, 2Hz,IonOptix Corp) from shams but not in hypertension. PA increased intracellular ATP level in shams.In contrast, etomoxir, a selective carnitine palmitoyl transferase I inhibitor, blunted the positive inotropic effect of PA, suggesting that increased beta‐oxidation may be fundamentally importantin PA‐regulation of cardiac inotropy. Mechanistically, PA did not increase the amplitude of Ca2+ transients or the density of peak L‐type Ca2+ current (ICa) or Na+‐Ca2+ exchanger activity, INCX, in either group. Paradoxically, PA did not affect the amplitude of Ca2+ transient or myofilament Ca2+ sensitivity (Myo‐Ca‐Sen) in shams. In hypertension, PA did not affect myocyte contraction or Ca2+ transient amplitude. Reactive oxygen species (ROS) scavenger did not affect PA‐induced increase in LV myocyte contraction in sham or hypertension.PA reduced neuronal nitric oxide synthase (nNOS) production of NO in shams but increased nNOS production of NO in hypertension. Importantly, nNOS inhibition with S‐methyl‐l‐thiocitrulline (SMTC) increased the amplitude of Ca2+ transients and restored PA‐induced cardiac inotropy in hypertension.ConclusionPA increases myocardial ATP production and potentiates LV myocyte contraction in healthy heart. nNOS restricts PA‐dependent cardiac inotropy by modulating Ca2+ handling in hypertension.

Effects of cannabidiol on contractions and calcium signaling in rat ventricular myocytes

Cannabidiol (CBD), a major nonpsychotropic cannabinoid found in Cannabis plant, has been shown to influence cardiovascular functions under various physiological and pathological conditions. In the present study, the effects of CBD on contractility and electrophysiological properties of rat ventricular myocytes were investigated. Video edge detection was used to measure myocyte shortening. Intracellular Ca2+ was measured in cells loaded with the Ca2+ sensitive fluorescent indicator fura-2 AM. Whole-cell patch clamp was used to measure action potential and Ca2+ currents. Radioligand binding was employed to study pharmacological characteristics of CBD binding. CBD (1μM) caused a significant decrease in the amplitudes of electrically evoked myocyte shortening and Ca2+ transients. However, the amplitudes of caffeine-evoked Ca2+ transients and the rate of recovery of electrically evoked Ca2+ transients following caffeine application were not altered. CBD (1μM) significantly decreased the duration of APs. Further studies on L-type Ca2+ channels indicated that CBD inhibits these channels with IC50 of 0.1μM in a voltage-independent manner. Radioligand studies indicated that the specific binding of [3H]Isradipine, was not altered significantly by CBD. The results suggest that CBD depresses myocyte contractility by suppressing L-type Ca2+ channels at a site different than dihydropyridine binding site and inhibits excitation–contraction coupling in cardiomyocytes.

Effects of total flavones from Acanthopanax senticosus on L-type calcium channels, calcium transient and contractility in rat ventricular myocytes.

Acanthopanax senticosus (Rupr. et Maxim.) Harms (AS), a traditional herbal medicine, has been widely used to treat ischemic heart disease. However, the underlying cellular mechanisms of its benefits to cardiac function remain unclear. The present study examined the effects of total flavones from AS (TFAS) on L-type Ca(2+) channel currents (ICa-L ) using the whole cell patch-clamp technique and on intracellular calcium ([Ca(2+) ]i ) handling and cell contractility in rat ventricular myocytes with the aid of a video-based edge-detection system. Exposure to TFAS resulted in a concentration- and voltage-dependent blockade of ICa-L , with the half-maximal inhibitory concentration (IC50 ) of 283.12 µg/mL and the maximal inhibitory effect of 36.49 ± 1.95%. Moreover, TFAS not only increased the maximum current in the current-voltage relationship but also shifted the activation and inactivation curves of ICa-L toward the hyperpolarizing direction. Meanwhile, TFAS significantly reduced amplitudes of myocyte shortening and [Ca(2+) ]i with an increase in the time to 10% of the peak (Tp) and a decrease in the time to 10% of the baseline (Tr). Thus, the cardioprotective effects of TFAS may be attributed mainly to the attenuation of [Ca(2+) ]i through the direct inhibition of ICa-L in rat ventricular myocytes and consequent negative effect on myocardial contractility.

Properties of Atrial Myocyte Calcium Handling in Canine Model of Chronic Heart Failure

Weakened atrial contractility is a common comorbidity in heart failure (HF) that increases severity of HF symptoms. The role of intracellular calcium (Ca) handling remodeling in pathogenesis of ventricular contractile failure has been well established. However, the relationship between Ca signaling and atrial dysfunction in HF is not well recognized. We studied intracellular Ca handling in atrial myocytes isolated from control dogs and dogs with tachypacing-induced chronic HF. Ca transients and cellular shortening were reduced, while sarcoplasmic reticulum (SR) Ca content was increased in field-stimulated myocytes from HF compared to controls. Both control and HF myocytes have scarce t-tubule density, and Ca transients originated predominantly at the periphery of myocytes in both groups. Diminished Ca transients in HF myocytes were associated with decreased peripheral Ca release and impaired propagation of Ca signal from sarcolemma to the central areas of the myocyte. Peak density of the L-type Ca currents and gain of excitation-contraction coupling were reduced in voltage-clamped HF myocytes compared to control myocytes. Analysis of decay of caffeine- and electrically-induced Ca transients suggests reduced Na/Ca exchanger activity and increased SR Ca ATPase (SERCA) activity in HF myocytes. SERCA inhibition with cyclopiazonic acid did not improve centripetal propagation of Ca signal in HF myocytes. In contrast, L-type Ca channel agonist BayK 8644 increased peripheral Ca release, improved centripetal Ca wave propagation and restored shortening of HF myocytes to control levels.We conclude that defective activation and impaired propagation of the SR Ca release in atrial myocytes contribute to compromised atrial contractility in chronic HF. Our results suggest L-type Ca channels as a therapeutic target for treatment of atrial contractile dysfunction in HF.

Abstract 15295: Molecular Mechanism of Chronic Sildenafil-Caused Regression of Heart Failure: Effects on the Express of Cardiac SR Ca2+-ATPase, Subtype of β-Adrenergic Receptors and Nitric Oxide Synthase Isoforms

Background: Sildenafil (SIL), a selective inhibitor of PDE5 has been shown to exert profound beneficial effects in heart failure (HF). Recently we further found that SIL caused regression of cardiac dysfunction in a rat model with isoproterenol (ISO)-induced progressive HF. However, the molecular basis is unclear. We hypothesized that reversal of HF-induced detrimental alterations on the expressions of cardiac SR Ca 2+ -ATPase (SERCA2a), β-adrenergic receptors (AR) and nitric oxide synthase (NOS) isoforms by SIL may play a key role for its salutary role in HF. Methods: Left ventricular (LV) and myocyte function and the protein levels of myocyte β 1 - and β 3 - AR, SERCA2a, phospholamban (PLB) and three NOS were simultaneously evaluated in 3 groups of male rats (6/group): HF , 3 months (M) after receiving ISO (170 mg/kg sq for 2 days); HF/SIL , 2 M after receiving ISO, SIL (70 μg/kg/day sq via mini pump) was initiated and given for 1 M; and Controls (C). Results: Compared with controls, ISO-treated rats progressed to severe HF at 3 M after ISO followed by significantly decreased LV contractility (E ES , HF: 0.7 vs C: 1.2 mmHg/μl) and slowed LV relaxation, reductions in the peak velocity of myocyte shortening (77 vs 136 μm/sec), relengthening (62 vs 104 μm/sec) and [Ca 2+ ] iT (0.15 vs 0.24) accompanied by a diminished myocyte inotropic response to β-AR agonist, ISO (10 -8 M). These abnormalities were associated with concomitant significant decreases in myocyte protein levels of β 1 -AR (0.23 vs 0.64), SERCA2a (0.46 vs 0.80), PLB Ser16 /PLB ratio (0.24 vs 0.40) and eNOS (0.28 vs 0.46), but significantly increases in protein levels of β 3 -AR (0.29 vs 0.10) and iNOS (0.18 vs 0.08) with relatively unchanged nNOS. Chronic SIL prevented the HF-induced decreases in LV and myocyte contraction, relaxation, peak [Ca 2+ ] iT , and restored normal myocyte contractile response to ISO stimulation. With SIL, protein levels of myocyte β 1 - and β 3 -AR, SERCA2a were restored close to control values, but eNOS was significantly elevated than controls (0.77). Conclusions: Chronic SIL prevents HF-caused downregulation of cardiac β 1 -AR and reverse contrast changes between iNOS and β 3 -AR with SERCA 2a and eNOS expression, leading to the preservation of LV and myocyte function, [Ca 2+ ] iT , and β-adrenergic reserve.

Salvia miltiorrhiza (Danshen) inhibits L-type calcium current and attenuates calcium transient and contractility in rat ventricular myocytes

Ethnopharmacological relevanceSalvia miltiorrhiza (SM, Danshen), a traditional Chinese herbal drug, has been widely used for hundreds of years to treat coronary artery disease. Materials and methodsWe studied the effects of SM on the L-type Ca2+ current (ICa-L) with whole-cell patch-clamp technique in rat ventricular myocytes, and its influence on Ca2+ transient and contractility using video-based edge detection and dual excitation fluorescence photomultiplier systems as well. ResultsExposure to SM solution caused a concentration- and voltage-dependent blockade of ICa-L, and the dose of SM solution (10g/l) decreased the maximal inhibitory effect of 35.2±1.2%. However, SM solution did not significantly change the current–voltage relationship or reversal potential of ICa-L, nor did it altered the activation and inactivation gating properties of cardiac Ca2+ channels. Meanwhile, SM decreased the amplitude of myocyte shortening and the peak value of Ca2+ transient with a significant decrease in the time to 90% of the baseline (Tr), but the time to 10% of the peak (Tp) was not dramatically prolonged. ConclusionsThe results indicated that SM significantly inhibited L-type Ca2+ channels, decreased [Ca2+]i and contractility in adult rat cardiac myocytes. These findings may be relevant to the cardioprotective efficacy of SM.

Dapagliflozin reduces the amplitude of shortening and Ca(2+) transient in ventricular myocytes from streptozotocin-induced diabetic rats.

In the management of type 2 diabetes mellitus, Dapagliflozin (DAPA) is a newly introduced selective sodium-glucose co-transporter 2 inhibitor which promotes renal glucose excretion. Little is known about the effects of DAPA on the electromechanical function of the heart. This study investigated the effects of DAPA on ventricular myocyte shortening and intracellular Ca(2+) transport in streptozotocin (STZ)-induced diabetic rats. Shortening, Ca(2+) transients, myofilament sensitivity to Ca(2+) and sarcoplasmic reticulum Ca(2+), and intracellular Ca(2+) current were measured in isolated rats ventricular myocytes by video edge detection, fluorescence photometry, and whole-cell patch-clamp techniques. Diabetes was characterized in STZ-treated rats by a fourfold increase in blood glucose (440 ± 25 mg/dl, n = 21) compared to Controls (98 ± 2 mg/dl, n = 19). DAPA reduced the amplitude of shortening in Control (76.68 ± 2.28 %, n = 37) and STZ (76.58 ± 1.89 %, n = 42) ventricular myocytes, and reduced the amplitude of the Ca(2+) transients in Control and STZ ventricular myocytes with greater effects in STZ (71.45 ± 5.35 %, n = 16) myocytes compared to Controls (92.01 ± 2.72 %, n = 17). Myofilament sensitivity to Ca(2+) and sarcoplasmic reticulum Ca(2+) were not significantly altered by DAPA in either STZ or Control myocytes. L-type Ca(2+) current was reduced in STZ myocytes compared to Controls and was further reduced by DAPA. In conclusion, alterations in the mechanism(s) of Ca(2+) transport may partly underlie the negative inotropic effects of DAPA in ventricular myocytes from STZ-treated and Control rats.

Ceramide-mediated depression in cardiomyocyte contractility through PKC activation and modulation of myofilament protein phosphorylation.

Although ceramide accumulation in the heart is considered a major factor in promoting apoptosis and cardiac disorders, including heart failure, lipotoxicity and ischemia-reperfusion injury, little is known about ceramide's role in mediating changes in contractility. In the present study, we measured the functional consequences of acute exposure of isolated field-stimulated adult rat cardiomyocytes to C6-ceramide. Exogenous ceramide treatment depressed the peak amplitude and the maximal velocity of shortening without altering intracellular calcium levels or kinetics. The inactive ceramide analog C6-dihydroceramide had no effect on myocyte shortening or [Ca(2+)]i transients. Experiments testing a potential role for C6-ceramide-mediated effects on activation of protein kinase C (PKC) demonstrated evidence for signaling through the calcium-independent isoform, PKCε. We employed 2-dimensional electrophoresis and anti-phospho-peptide antibodies to test whether treatment of the cardiomyocytes with C6-ceramide altered myocyte shortening via PKC-dependent phosphorylation of myofilament proteins. Compared to controls, myocytes treated with ceramide exhibited increased phosphorylation of myosin binding protein-C (cMyBP-C), specifically at Ser273 and Ser302, and troponin I (cTnI) at sites apart from Ser23/24, which could be attenuated with PKC inhibition. We conclude that the altered myofilament response to calcium resulting from multiple sites of PKC-dependent phosphorylation contributes to contractile dysfunction that is associated with cardiac diseases in which elevations in ceramides are present.

Cardiac P2X purinergic receptors as a new pathway for increasing Na⁺ entry in cardiac myocytes.

P2X4 receptors (P2X4Rs) are ligand-gated ion channels capable of conducting cations such as Na(+). Endogenous cardiac P2X4R can mediate ATP-activated current in adult murine cardiomyocytes. In the present study, we tested the hypothesis that cardiac P2X receptors can induce Na(+) entry and modulate Na(+) handling. We further determined whether P2X receptor-induced stimulation of the Na(+)/Ca(2+) exchanger (NCX) has a role in modulating the cardiac contractile state. Changes in Na(+)-K(+)-ATPase current (Ip) and NCX current (INCX) after agonist stimulation were measured in ventricular myocytes of P2X4 transgenic mice using whole cell patch-clamp techniques. The agonist 2-methylthio-ATP (2-meSATP) increased peak Ip from a basal level of 0.52 ± 0.02 to 0.58 ± 0.03 pA/pF. 2-meSATP also increased the Ca(2+) entry mode of INCX (0.55 ± 0.09 pA/pF under control conditions vs. 0.82 ± 0.14 pA/pF with 2-meSATP) at a membrane potential of +50 mV. 2-meSATP shifted the reversal potential of INCX from -14 ± 2.3 to -25 ± 4.1 mV, causing an estimated intracellular Na(+) concentration increase of 1.28 ± 0.42 mM. These experimental results were closely mimicked by mathematical simulations based on previously established models. KB-R7943 or a structurally different agent preferentially opposing the Ca(2+) entry mode of NCX, YM-244769, could inhibit the 2-meSATP-induced increase in cell shortening in transgenic myocytes. Thus, the Ca(2+) entry mode of INCX participates in P2X agonist-stimulated contractions. In ventricular myocytes from wild-type mice, the P2X agonist could increase INCX, and KB-R7943 was able to inhibit the contractile effect of endogenous P2X4Rs, indicating a physiological role of these receptors in wild-type cells. The data demonstrate a novel Na(+) entry pathway through ligand-gated P2X4Rs in cardiomyocytes.

Abstract 93: A Gain-of-Function Mutation in Cardiac Myosin Binding Protein-C Increases Viscoelastic Load and Slows Shortening Velocity in Myocytes from Transgenic Mice

Cardiac myosin binding protein C (cMyBP-C) is a sarcomeric protein involved in the regulation of cardiac muscle contraction. Effects of cMyBP-C on contraction are thought to be mediated in part by limiting the interactions of actin and myosin to slow myocyte shortening velocity and power output. Although interactions with myosin S2 on the thick filament have been proposed as a way in which cMyBP-C could limit shortening velocity (e.g., by creating a drag force on myosin heads), interactions of cMyBP-C with actin could also account for slowed shortening velocity. For instance, cMyBP-C could create a drag that opposes filament sliding by transiently linking thick and thin filaments together. To explore this possibility we created transgenic mice that express a mutant cMyBP-C with a point mutation, L348P (human L352P), located in a conserved sequence within the regulatory M-domain that increases cMyBP-C binding to actin in vitro. We reasoned that if the mutation also enhanced binding to actin in sarcomeres then shortening velocity would be slowed in myocytes from L348P mice. Results show that transgenic mice expressing the L348P mutation are viable and that L348P cMyBP-C is expressed in sarcomeres. Permeabilized myocytes from transgenic mice showed altered force production including reduced maximal force and enhanced calcium sensitivity of tension. Shortening velocity and power output were significantly reduced whereas passive stiffness and myocyte visco-elasticity were significantly increased. Together these data are consistent with the idea that cMyBP-C creates an internal load in the sarcomere by binding to actin.

Effects of a sucrose‐enriched diet on the pattern of gene expression, contraction and Ca2+ transport in Goto–Kakizaki type 2 diabetic rat heart

There has been a spectacular rise in the global prevalence of type 2 diabetes mellitus (T2DM), and cardiovascular disease is the major cause of morbidity and mortality in diabetic patients. A variety of diastolic and systolic dysfunctions have been demonstrated in type 2 diabetic heart. The consumption of sugar-sweetened beverages has been linked to rising rates of obesity, which in turn is a risk factor for development of T2DM. In this study, the effects of a sucrose-enriched diet on the pattern of gene expression, contraction and Ca(2+) transport in the Goto-Kakizaki T2DM rat heart were investigated. Genes encoding cardiac muscle proteins (Myh7, Mybpc3, Myl1, Myl3 and Mylpf), intercellular proteins (Gja4), cell membrane transport (Atp1b1), calcium channels (Cacna1c, Cacna1g and Cacnb1) and potassium channels (Kcnj11) were upregulated and genes encoding potassium channels (Kcnb1) were downregulated in GK compared with control rats. Genes encoding cardiac muscle proteins (Myh6, Mybpc3 and Tnn2), intercellular proteins (Gja1 and Gja4), intracellular Ca(2+) transport (Atp2a1 and Ryr2), cell membrane transport (Atp1a2 and Atp1b1) and potassium channel proteins (Kcnj2 and Kcnj8) were upregulated and genes encoding cardiac muscle proteins (Myh7) were downregulated in control rats fed sucrose compared with control rats. Genes encoding cardiac muscle proteins (Myh7) and potassium channel proteins (Kcnj11) were downregulated in control and GK rats fed sucrose compared with control and GK rats, respectively. The amplitude of shortening was reduced in myocytes from the control-sucrose group compared with control rats and in the GK-sucrose group compared with GK rats. The amplitude of the Ca(2+) transient was increased in myocytes from control-sucrose compared with control rats and decreased in GK-sucrose compared with GK rats. Subtle alterations in the pattern of expression of genes encoding a variety of cardiac muscle proteins are associated with changes in shortening and intracellular Ca(2+) transport in ventricular myocytes from GK T2DM and control rats fed a sucrose-enriched diet.

Mechanisms by which calcium receptor stimulation modifies electromechanical coupling in isolated ventricular cardiomyocytes

The calcium-sensing receptor (CaR) is widely expressed throughout the entire cardiovascular system and is capable of activating signaling pathways in different cells. Alongside calcium, the CaR also responds to physiological polycations such as putrescine underlining a participation in physiological and pathophysiological processes. Here, we aimed to determine mechanisms as to how CaR activation affects the contractile responsiveness of ventricular cardiomyocytes under basal and stimulated conditions. For that purpose, cardiac myocytes from 3-month-old male Wistar rats were isolated, and the acute effects of an antagonist (NPS2390), agonists (putrescine and gadolinium), or of downregulation of the CaR by siRNA on cell shortening were recorded in a cell-edge-detection system. In addition, experiments were performed on muscle stripes and Langendorff preparations. Mechanistic insights were taken from calcium transients of beating fura-2 AM-loaded cardiomyocytes and western blots. Isolated ventricular cardiomyocytes constitutively express CaR. The expression in the atria is less pronounced. Acute inhibition of CaR reduced basal cell shortening of ventricular myocytes at nearly physiological levels of extracellular calcium. Inhibition of CaR strongly reduced contractility of ventricular muscle stripes but not of atria. Activation of CaR by putrescine and gadolinium influences the contractile responsiveness of isolated cardiomyocytes. Increased calcium mobilization from the sarcoplasmic reticulum via an IP3-dependent mechanism was responsible for amplified systolic calcium transients and a subsequent improvement in cell shortening. Alongside with these effects, activation of CaR increased relaxation velocity of the cells. In conclusion, ventricular CaR expression affects contractile parameters of ventricular heart muscle cells and modifies electromechanical coupling of cardiomyocytes.

Select Articles Published on the Topic of Arrhythmia and Electrophysiogy in 2013

Select Articles Published on the Topic of Arrhythmia and Electrophysiogy in 2013

Effects of endogenous cannabinoid anandamide on excitation–contraction coupling in rat ventricular myocytes

A role for anandamide (N-arachidonoyl ethanolamide; AEA), a major endocannabinoid, in the cardiovascular system in various pathological conditions has been reported in earlier reports. In the present study, the effects of AEA on contractility, Ca2+ signaling, and action potential (AP) characteristics were investigated in rat ventricular myocytes. Video edge detection was used to measure myocyte shortening. Intracellular Ca2+ was measured in cells loaded with the fluorescent indicator fura-2 AM. AEA (1μM) caused a significant decrease in the amplitudes of electrically evoked myocyte shortening and Ca2+ transients. However, the amplitudes of caffeine-evoked Ca2+ transients and the rate of recovery of electrically evoked Ca2+ transients following caffeine application were not altered. Biochemical studies in sarcoplasmic reticulum (SR) vesicles from rat ventricles indicated that AEA affected Ca2+-uptake and Ca2+-ATPase activity in a biphasic manner. [3H]-ryanodine binding and passive Ca2+ release from SR vesicles were not altered by 10μM AEA. Whole-cell patch-clamp technique was employed to investigate the effect of AEA on the characteristics of APs. AEA (1μM) significantly decreased the duration of AP. The effect of AEA on myocyte shortening and AP characteristics was not altered in the presence of pertussis toxin (PTX, 2μg/ml for 4h), AM251 and SR141716 (cannabinoid type 1 receptor antagonists; 0.3μM) or AM630 and SR 144528 (cannabinoid type 2 receptor antagonists; 0.3μM). The results suggest that AEA depresses ventricular myocyte contractility by decreasing the action potential duration (APD) in a manner independent of CB1 and CB2 receptors.

Well-preserved ventricular myocyte shortening in Goto–Kakizaki type 2 diabetic rats

The global prevalence of type 2 diabetes mellitus (T2DM) is rising at a spectacular rate and the prevalence of T2DM in the United Arab Emirates is among the highest in the world. Risk factors for diabetes mellitus include obesity, sedentary lifestyle, unhealthy eating habits, family history of diabetes mellitus, genetics, increasing age, high blood pressure and high cholesterol. A variety of diastolic and systolic dysfunctions have been reported in the hearts of type 2 diabetic patients and the severity of abnormalities is partly dependent on the patient's age and the duration of diabetes mellitus. The Goto–Kakizaki (GK) rat is a genetic model of T2DM and its general characteristics include fasting hyperglycaemia, impaired insulin secretion and insulin resistance. In the GK rat heart, there are a variety of dysfunctions including decreased heart rate, decreased ejection fraction (mainly due to loss of left ventricular longitudinal contraction) and prolonged shortening and/or relaxation in ventricular myocytes. Widespread consumption of sugar-sweetened beverages has been linked to higher incidence of obesity and T2DM. Ventricular myocyte shortening and Ca2+ transport in GK type 2 diabetic rats that were fed a sucrose-enriched diet have been investigated. GK and control rats received either water or water enriched with increasing concentrations of sucrose (100–400 mmol/l) for a period of 6 months. Ventricular myocyte shortening and intracellular Ca2+ were measured by video edge detection and fluorescence photometry, respectively. Resting cell length, time to peak (TPK) shortening, time to half-relaxation of shortening and amplitude of shortening were not significantly altered in ventricular myocytes from GK/sucrose compared with GK rats and control/sucrose compared with control rats. TPK Ca2+ transient was prolonged in myocytes from GK/sucrose compared with GK and control/sucrose rats and amplitude of the Ca2+ transient was increased in myocytes from GK/sucrose compared with control rats. Myofilament sensitivity to Ca2+ was unaltered in myocytes from GK/sucrose and control/sucrose compared with GK and control rats. In conclusion, ventricular myocyte shortening was well preserved despite some alterations in Ca2+ transport in GK rats receiving a sucrose-enriched diet.