Myofilament Calcium Sensitivity Research Articles

Abstract 1238: Myofilament Calcium Sensitization Creates Arrhythmia Susceptibility in Familial Hypertrophic Cardiomyopathy

Although sudden cardiac death is a common feature in hypertrophic cardiomyopathy (HCM), the underlying mechanisms are unclear. We recently reported that some but not all mice expressing troponin mutations associated with HCM display increased myofilament calcium sensitivity. Here, we used these mice as well as acute challenge with the Ca 2+ sensitizing agent EMD57033 (EMD) in isolated mouse and rabbit hearts to test the hypothesis that myofilament sensitization leads directly to arrhythmia susceptibility. HCM mice with Ca 2+ sensitizing troponin mutations displayed significantly higher rate of ventricular ectopy (TnT-I79N 10±3, TnT-F110I 11±5, ssTnI 15±4 PVC/h) than mice expressing non-sensitizing troponins (TnT-R278C 1±1, TnT-WT 1±1, non-Tg 2±1 PVC/h, n=5–14 per group, p<0.01). Myofilament sensitization by troponin mutations or EMD shortened the effective refractory period and the ventricular action potential of isolated perfused mouse and rabbit hearts, and caused early afterdepolarizations and triggered beats after fast pacing trains. The action potential shortening was attributable to increased cytosolic Ca 2+ buffering by the Ca 2+ sensitized myofilaments, which resulted in decreased systolic and increased end-diastolic Ca 2+ during fast pacing. Optical mapping demonstrated that Ca 2+ sensitization slowed the ventricular conduction velocity and increased the size of the vulnerable window for ventricular tachycardia both in mouse and rabbit hearts, resulting in a significantly higher incidence of sustained ventricular tachycardia in Ca 2+ sensitized compared with control hearts (5/6 vs 0/6, p<0.05). Conclusion: Myofilament Ca 2+ sensitization alters Ca 2+ buffering to render hearts susceptible to ventricular arrhythmias by creating both an arrhythmogenic substrate and trigger. These results identify a novel mechanism linking sarcomeric mutations to susceptibility to ventricular arrhythmias and raise the prospect of a molecular approach to stratifying arrhythmia risk in HCM.

Dilated Cardiomyopathy Mutant Tropomyosin Mice Develop Cardiac Dysfunction With Significantly Decreased Fractional Shortening and Myofilament Calcium Sensitivity

Mutations in striated muscle alpha-tropomyosin (alpha-TM), an essential thin filament protein, cause both dilated cardiomyopathy (DCM) and familial hypertrophic cardiomyopathy. Two distinct point mutations within alpha-tropomyosin are associated with the development of DCM in humans: Glu40Lys and Glu54Lys. To investigate the functional consequences of alpha-TM mutations associated with DCM, we generated transgenic mice that express mutant alpha-TM (Glu54Lys) in the adult heart. Results showed that an increase in transgenic protein expression led to a reciprocal decrease in endogenous alpha-TM levels, with total myofilament TM protein levels remaining unaltered. Histological and morphological analyses revealed development of DCM with progression to heart failure and frequently death by 6 months. Echocardiographic analyses confirmed the dilated phenotype of the heart with a significant decrease in the left ventricular fractional shortening. Work-performing heart analyses showed significantly impaired systolic, and diastolic functions and the force measurements of cardiac myofibers revealed that the myofilaments had significantly decreased Ca(2+) sensitivity and tension generation. Real-time RT-PCR quantification demonstrated an increased expression of beta-myosin heavy chain, brain natriuretic peptide, and skeletal actin and a decreased expression of the Ca(2+) handling proteins sarcoplasmic reticulum Ca(2+)-ATPase and ryanodine receptor. Furthermore, our study also indicates that the alpha-TM54 mutation decreases tropomyosin flexibility, which may influence actin binding and myofilament Ca(2+) sensitivity. The pathological and physiological phenotypes exhibited by these mice are consistent with those seen in human DCM and heart failure. As such, this is the first mouse model in which a mutation in a sarcomeric thin filament protein, specifically TM, leads to DCM.

Postnatal maturation attenuates pressure-evoked myogenic tone and stretch-induced increases in Ca2+in rat cerebral arteries

Although postnatal maturation potently modulates agonist-induced cerebrovascular contractility, its effects on the mechanisms mediating cerebrovascular myogenic tone remain poorly understood. Because the regulation of calcium influx and myofilament calcium sensitivity change markedly during early postnatal life, the present study tested the general hypothesis that early postnatal maturation increases the pressure sensitivity of cerebrovascular myogenic tone via age-dependent enhancement of pressure-induced calcium mobilization and myofilament calcium sensitivity. Pressure-induced myogenic tone and changes in artery wall intracellular calcium concentrations ([Ca(2+)](i)) were measured simultaneously in endothelium-denuded, fura-2-loaded middle cerebral arteries (MCA) from pup [postnatal day 14 (P14)] and adult (6-mo-old) Sprague-Dawley rats. Increases in pressure from 20 to 80 mmHg enhanced myogenic tone in MCA from both pups and adults although the normalized magnitudes of these increases were significantly greater in pup than adult MCA. At each pressure step, vascular wall [Ca(2+)](i) was also significantly greater in pup than in adult MCA. Nifedipine significantly attenuated pressure-evoked constrictions in pup MCA and essentially eliminated all responses to pressure in the adult MCA. Both pup and adult MCA exhibited pressure-dependent increases in calcium sensitivity, as estimated by changes in the ratio of pressure-induced myogenic tone to wall [Ca(2+)](i). However, there were no differences in the magnitudes of these increases between pup and adult MCA. The results support the view that regardless of postnatal age, changes in both calcium influx and myofilament calcium sensitivity contribute to the regulation of cerebral artery myogenic tone. The greater cerebral myogenic response in P14 compared with adult MCA appears to be due to greater pressure-induced increases in [Ca(2+)](i), rather than enhanced augmentation of myofilament calcium sensitivity.

Rescue of tropomyosin-induced familial hypertrophic cardiomyopathy mice by transgenesis

Familial hypertrophic cardiomyopathy (FHC) is a disease caused by mutations in contractile proteins of the sarcomere. Our laboratory developed a mouse model of FHC with a mutation in the thin filament protein alpha-tropomyosin (TM) at amino acid 180 (Glu180Gly). The hearts of these mice exhibit dramatic systolic and diastolic dysfunction, and their myofilaments demonstrate increased calcium sensitivity. The mice also develop severe cardiac hypertrophy, with death ensuing by 6 mo. In an attempt to normalize calcium sensitivity in the cardiomyofilaments of the hypertrophic mice, we generated a chimeric alpha-/beta-TM protein that decreases calcium sensitivity in transgenic mouse cardiac myofilaments. By mating mice from these two models together, we tested the hypothesis that an attenuation of myofilament calcium sensitivity would modulate the severe physiological and pathological consequences of the FHC mutation. These double-transgenic mice "rescue" the hypertrophic phenotype by exhibiting a normal morphology with no pathological abnormalities. Physiological analyses of these rescued mice show improved cardiac function and normal myofilament calcium sensitivity. These results demonstrate that alterations in calcium response by modification of contractile proteins can prevent the pathological and physiological effects of this disease.

Myogenic contractility is more dependent on myofilament calcium sensitization in term fetal than adult ovine cerebral arteries

Regulation of cytosolic calcium and myofilament calcium sensitivity varies considerably with postnatal age in cerebral arteries. Because these mechanisms also govern myogenic tone, the present study used graded stretch to examine the hypothesis that myogenic tone is less dependent on calcium influx and more dependent on myofilament calcium sensitization in term fetal compared with adult cerebral arteries. Term fetal and adult posterior communicating cerebral arteries exhibited similar myogenic responses, with peak tensions averaging 24 and 26% of maximum contractile force produced in any given tissue in response to an isotonic Krebs buffer containing 122 mM K(+) (K(max)) at optimum stretch ratios (working diameter/unstressed diameter) of 2.19 and 2.23, respectively. Graded stretch increased cytosolic Ca(2+) concentration at stretch ratios >2.0 in adult arteries, but increased Ca(2+) concentration only at stretch ratios >2.3 in fetal arteries. In permeabilized arteries, myogenic tone peaked at a stretch ratio of 2.1 in both fetal and adult arteries. The fetal %K(max) values at peak myogenic tone were not significantly different at either pCa 7.0 (23%) or pCa 5.5 (25%) but were significantly less at pCa 8.0 (8.4 +/- 2.3%). Conversely, adult %K(max) values at peak myogenic tone were significantly less at both pCa 8.0 (10.4 +/- 1.8%) and pCa 7.0 (16%) than at pCa 5.5 (27%). The maximal extents of stretch-induced increases in myosin light chain phosphorylation in intact fetal (20%) and adult (17%) arteries were similar. The data demonstrate that the cerebrovascular myogenic response is highly conserved during postnatal maturation but is mediated differently in fetal and adult cerebral arteries.

PKC catalytic subunit and phenylephrine induce a comparable decrease in myofilament calcium sensitivity

The in vivo regulation of cardiomyocyte function by protein kinase C (PKC) and by phenylephrine (PE), which is also believed to occur partly via PKC activation, has not been clarified yet. We determined the effect of the PKC catalytic subunit and PE on force development in rat cardiomyocytes. Isometric force and rate of force redevelopment (Ktr) were studied in adult rat cardiomyocytes at various [Ca2+] before and after treatment with PKC (0.1 and 0.2 U/ml) and after stimulation with PE. In order to reveal PKC-dependent phosphorylation of contractile proteins two-dimensional gel electrophoresis was performed.

Protein kinase D expression and activity are unaltered during myofibrillogenesis in C2C12 cells

Protein kinase D (PKD) has been implicated in the signaling pathways leading to cardiac hypertrophy and heart failure, and in the regulation of myofilament calcium sensitivity. We have found that PKD associates with a number of myofibril proteins, including several (e.g. NRAP, telethonin and obscurin) which are important in the formation of myofibrils. This has led us to hypothesise that PKD may have a role in myofibrillogenesis, and to determine PKD expression and activity in the C2C12 skeletal myoblast cell line, which can be induced to differentiate into myotubes by the withdrawal of serum, with the de novo formation of myofibrils.

Frequency-dependent acceleration of relaxation involves decreased myofilament calcium sensitivity

The force-frequency relationship is an intrinsic modulator of cardiac contractility and relaxation. Force of contraction increases with frequency, while simultaneously a frequency-dependent acceleration of relaxation occurs. While frequency dependency of calcium handling and sarcoplasmic reticulum calcium load have been well described, it remains unknown whether frequency-dependent changes in myofilament calcium sensitivity occur. We hypothesized that an increase in heart rate that results in acceleration of relaxation is accompanied by a proportional decrease in myofilament calcium sensitivity. To test our hypothesis, ultrathin right ventricular trabeculae were isolated from New Zealand White rabbit hearts and iontophorically loaded with the calcium indicator bis-fura 2. Twitch and intracellular calcium handling parameters were measured and showed a robust increase in twitch force, acceleration of relaxation, and rise in both diastolic and systolic intracellular calcium concentration with increased frequency. Steady-state force-intracellular calcium concentration relationships were measured at frequencies 1, 2, 3, and 4 Hz at 37 degrees C using potassium-induced contractures. EC(50) significantly and gradually increased with frequency, from 475 +/- 64 nM at 1 Hz to 1,004 +/- 142 nM at 4 Hz (P < 0.05) and correlated with the corresponding changes in half relaxation time. No significant changes in maximal active force development or in the myofilament cooperativity coefficient were found. Myofilament protein phosphorylation was assessed using Pro-Q Diamond staining on protein gels of trabeculae frozen at either 1 or 4 Hz, revealing troponin I and myosin light chain-2 phosphorylation associated with the myofilament desensitization. We conclude that myofilament calcium sensitivity is substantially and significantly decreased at higher frequencies, playing a prominent role in frequency-dependent acceleration of relaxation.

Computational simulation of hypertrophic cardiomyopathy mutations in Troponin I: Influence of increased myofilament calcium sensitivity on isometric force, ATPase and [Ca 2+] i

Familial hypertrophic cardiomyopathy (FHC) is an inherited disease that is characterized by ventricular hypertrophy, cardiac arrhythmias and increased risk of premature sudden death. FHC is caused by autosomal-dominant mutations in genes for a number of sarcomeric proteins; many mutations in Ca 2+-regulatory proteins of the cardiac thin filament are associated with increased Ca 2+ sensitivity of myofilament function. Computational simulations were used to investigate the possibility that these mutations could affect the Ca 2+ transient and mechanical response of a myocyte during a single cardiac cycle. We used existing experimental data for specific mutations of cardiac troponin I that exhibit increased Ca 2+ sensitivity in physiological and biophysical assays. The simulated Ca 2+ transients were used as input for a three-dimensional half-sarcomere biomechanical model with filament compliance to predict the resulting force. Mutations with the highest Ca 2+ affinity (lowest K m) values, exhibit the largest decrease in peak Ca 2+ assuming a constant influx of Ca 2+ into the cytoplasm; they also prolong Ca 2+ removal but have little effect on diastolic Ca 2+. Biomechanical model results suggest that these cTnI mutants would increase peak force despite the decrease in peak [Ca 2+] i . There is a corresponding increase in net ATP hydrolysis, with no change in tension cost (ATP hydrolyzed per unit of time-integrated tension). These simulations suggest that myofilament-initiated hypertrophic signaling could be associated with decreased [Ca 2+] i , increased stress/strain, and/or increased ATP flux.

Myofilament calcium sensitivity does not affect cross-bridge activation-relaxation kinetics

We employed single myofibril techniques to test whether the presence of slow skeletal troponin-I (ssTnI) is sufficient to induce increased myofilament calcium sensitivity (EC(50)) and whether modulation of EC(50) affects the dynamics of force development. Studies were performed using rabbit psoas myofibrils activated by rapid solution switch and in which Tn was partially replaced for either recombinant cardiac Tn(cTn) or Tn composed of recombinant cTn-T (cTnT) and cTn-C (cTnC), and recombinant ssTnI (ssTnI-chimera Tn). Tn exchange was performed in rigor solution (0.5 mg/ml Tn; 20 degrees C; 2 h) and confirmed by SDS-PAGE. cTnI exchange induced a decrease in EC(50); ssTnI-chimera Tn exchange induced a further decrease in EC(50) (in microM: endogenous Tn, 1.35 +/- 0.08; cTnI, 1.04 +/- 0.13; ssTnI-chimera Tn, 0.47 +/- 0.03). EC(50) was also decreased by application of 100 microM bepridil (control: 2.04 +/- 0.03 microM; bepridil 1.35 +/- 0.03 microM). Maximum tension was not different between any groups. Despite marked alterations in EC(50), none of the dynamic activation-relaxation parameters were affected under any condition. Our results show that 1) incorporation of ssTnI into the fast skeletal sarcomere is sufficient to induce increased myofilament Ca(2+) sensitivity, and 2) the dynamics of actin-myosin interaction do not correlate with EC(50). This result suggests that intrinsic cross-bridge cycling rate is not altered by the dynamics of thin-filament activation.

Development and Cardiac Contractility: Cardiac Troponin T Isoforms and Cytosolic Calcium in Rabbit

Cardiac contractility depends on calcium sensitivity of the myofilaments and cytosolic free calcium concentration ([Ca(2+)](i)) during activation. During development, the cardiac troponin T isoform cTnT(1) is replaced by shorter cTnT isoforms, including cTnT(4), and changes occur in other myofibrillar proteins and in calcium regulation. We expressed rabbit recombinant (r)cTnT(1) and rcTnT(4) in Spodoptera frugiperda cells and determined their effect on calcium binding to TnC in solution and on the calcium sensitivity of myofilaments in skinned rabbit ventricular fibers in vitro. We measured [Ca(2+)](i) and L-type calcium current (I(Ca)) in ventricular myocytes from 3-wk-old and adult rabbits. The dissociation constant (K(d)) of Ca-Tn(cTnT1) in solution was smaller than that of Ca-Tn(cTnT4) (mean +/- SE: 0.52 +/- 0.08 mumol/L versus 0.83 +/- 0.09 mumol/L). The Ca(2+) sensitivity of force development was greater in fibers reconstituted with rcTnT(1) (pCa(50) 6.07 +/- 0.04) than those reconstituted with rcTnT(4) (pCa(50) 5.75 +/- 0.07). Systolic [Ca](i) was lower in 3-wk-old than adult cells (443 +/- 35 nmol/L versus 882 +/- 88 nmol/L) as was I(Ca) (5.8 +/- 0.9 pA/pF versus 14.2 +/- 1.6 pA/pF). The higher calcium sensitivity of Tn-Ca binding and of force development conferred by rcTnT(1) suggest that higher neonatal cTnT(1) expression may partially compensate for the lower systolic [Ca(2+)](i).

Differences in myofilament calcium sensitivity in rat psoas fibers reconstituted with troponin T isoforms containing the α- and β-exons

The carboxy terminus of fast skeletal muscle troponin T (fsTnT) is highly conserved. However, mutually exclusive splicing of exons 16 and 17 in the fsTnT gene results in the expression of either the α- or β-fsTnT isoform. The α-isoform is expressed only in adult fast skeletal muscle, whereas the β-isoform is expressed in varying quantities throughout muscle development. Reconstitution of detergent-skinned adult rat psoas muscle fibers with rat fast skeletal troponin complexes containing either fsTnT isoform demonstrated that reconstitution with α-fsTnT resulted in greater myofilament Ca 2+ sensitivity than reconstitution with β-fsTnT, without changes to Ca 2+-activated maximal tension, ATPase activity or tension cost. The observed isoform-specific differences in myofilament Ca 2+ sensitivity may be due to changes in the transition of the thin-filament regulatory unit from the off to the on state, possibly due to altered interactions of the C-terminus of fsTnT with troponins I and/or C.

Impact of osmotic compression on sarcomere structure and myofilament calcium sensitivity of isolated rat myocardium

Changes in interfilament lattice spacing have been proposed as the mechanism underlying myofilament length-dependent activation. Much of the evidence to support this theory has come from experiments in which high-molecular-weight compounds, such as dextran, were used to osmotically shrink the myofilament lattice. However, whether interfilament spacing directly affects myofilament calcium sensitivity (EC(50)) has not been established. In this study, skinned isolated rat myocardium was osmotically compressed over a wide range (Dextran T500; 0-6%), and EC(50) was correlated to both interfilament spacing and I(1,1)/I(1,0) intensity ratio. The latter two parameters were determined by X-ray diffraction in a separate group of skinned muscles. Osmotic compression induced a marked reduction in myofilament lattice spacing, concomitant with increases in both EC(50) and I(1,1)/I(1,0) intensity ratio. However, interfilament spacing was not well correlated with EC(50) (r(2) = 0.78). A much better and deterministic relationship was observed between EC(50) and the I(1,1)/I(1,0) intensity ratio (r(2) = 0.99), albeit with a marked discontinuity at low levels of dextran compression; that is, a small amount of external osmotic compression (0.38 kPa, corresponding to 1% Dextran T500) produced a stepwise increase in the I(1,1)/I(1,0) ratio concomitant with a stepwise decrease in EC(50). These parameters then remained stable over a wide range of further applied osmotic compression (up to 6% dextran). These findings provide support for a "switch-like" activation mechanism within the cardiac sarcomere that is highly sensitive to changes in external osmotic pressure.

Myocardial Dysfunction and Potential Cardiac Hypoxia in Rats Induced by Carbon Monoxide Inhalation

Results from both animal and human being studies provide evidence that inhalation of concentrations of carbon monoxide (CO) at around 100 ppm has antiinflammatory effects. These low levels of CO are incriminated in ischemic heart diseases experienced by cigarette smokers and, in some cases, from air pollution. Although neurologic mechanisms have been investigated, the effects of CO on cardiovascular function are still poorly understood. The effects of CO (250 ppm; 90 min) inhalation on myocardial function were investigated in isolated heart of rats killed immediately, and 3, 24, 48, and 96 h after CO exposure. CO exposure at 250 ppm resulted in an arterial carboxyhemoglobin (HbCO) level of approximately 11%, which was not associated with changes in mean arterial pressure and heart rate. CO exposure induced coronary perfusion pressure increases, which were associated with endothelium-dependent and -independent vascular relaxation abnormalities. CO-induced coronary vascular relaxation perturbations were observed in the presence of increased heart contractility. Spontaneous peak to maximal Ca(2+)-activated left ventricular pressure ratio was markedly increased in CO-exposed rats, indicating increases in myofilament calcium sensitivity. Heart cyclic guanosine monophosphate/cAMP ratio and myocardial permeabilized fiber respiration (complex intravenous activity) were reduced in CO-exposed rats, which lasted after 48 h of reoxygenation in air. These findings suggest that CO deteriorates heart oxygen supply to utilization and potentially may induce myocardial hypoxia through mechanisms that include increased oxygen demand due to increased contractility, reduced coronary blood flow reserve, and cardiomyocyte respiration inhibition.

Measurement of myofilament calcium sensitivity at physiological temperature in intact cardiac trabeculae

Cardiac contraction-relaxation coupling is determined by both the free intracellular calcium concentration ([Ca2+]i) and myofilament properties. We set out to develop a technique where we could assess these parameters (twitch and steady-state force [Ca2+]i) under near physiological conditions. Bis-fura-2 was iontophorically introduced into ultrathin rat trabeculae preparations to monitor the [Ca2+]i, and steady-state contractures were achieved by using a modified Krebs-Henseleit solution containing high K+. During K+ contractures, the very slow changes in [Ca2+]i and force development were in equilibrium and allowed for the construction of a steady-state, force-[Ca2+]i relationship. Twitch contractions before and after this myofilament calcium sensitivity assessment were unaltered, and this protocol could be repeated several times. For the first time, this novel protocol allows us to measure myofilament calcium sensitivity under physiological temperature. Not only do the data so obtained allow us to assess myofilament calcium sensitivity, the data also will allow us, in the same preparation under nearly identical conditions, to compare the dynamic to the steady-state, force-calcium relationship. To test whether the steady-state relationship between force and calcium in our novel protocol reproduces expected changes, we determined this relationship in the presence of isoproterenol and under acidosis and alkalosis. As expected, beta-adrenergic stimulation resulted in an increase of calcium amplitude and twitch force and a desensitization of the myofilaments as indicated by a rightward shift of the obtained steady-state, force-calcium relationship. An increase in pH shifted the curve leftward, whereas a decrease in pH resulted in the expected rightward shift.

Insight into the role of DL-α-lipoic acid against cyclophosphamide induced alterations in calcium sensitivity of cardiac myofilaments

Cyclophosphamide (CP), a potent antitumor drug, is known to cause severe cardiotoxicity. The present study is aimed at evaluating the role of DL-alpha-Lipoic acid (LA) on the calcium responsiveness of cardiac myofilaments isolated from CP treated rats. Adult male Wistar rats were divided into four treatment groups. Two groups received single intraperitoneal injection of CP (200 mg/kg b.wt.) to induce cardiotoxicity, one of these groups received LA treatment (25 mg/kg b.wt. for 10 days). A vehicle treated control group and a LA drug control were also included. Cardiotoxicity was evident from increased levels of cardiac Troponin I in serum of CP treated rats. The pCa-actomyosin ATPase relationship of myofilaments demonstrated a rightward shift indicating diminished responsiveness in CP treated rats. The hill coefficient was reduced and the myofibrillar myosin Ca(2+)-ATPase and K(+)-(EDTA) activities were also significantly (P < 0.05) reduced. Ultrastructural observations were also in agreement with the above abnormal changes, wherein loss of myofilaments occurred. LA effectively normalized these abnormalities and restored the cardiac function in CP administered rats.

Age‐dependent stretch‐induced myofilament calcium sensitization in ovine cerebral arteries

Myogenic tone is a well-established characteristic of cerebral arteries, but the mechanisms of this response are not fully understood, particularly in immature cerebral arteries. Given that myofilament calcium sensitivity is typically greater in fetal than adult cerebral arteries, we explored the hypothesis that the characteristics of myogenic tone should differ in fetal and adult arteries. To test this idea, we measured intracellular calcium via Fura-2 in intact ovine posterior communicating arteries and found that stretch induced a graded increase in calcium (max ?ratio = .292), but only in adult arteries even though the magnitudes of stretch-induced contraction were similar in adult (265% Kmax) and fetal (247% Kmax) arteries. Maximum levels of stretch-induced myosin light chain (MLC) phosphorylation were also similar (?41–44%) in fetal and adult arteries. However, when length-tension measurements were performed in permeabilized arteries held at pCa 7, graded stretch produced graded increases in force that were greater in adult (207% Kmax) than fetal (110% Kmax) arteries, indicating stretch-induced calcium sensitization. Similar results were obtained at pCa 5.5. Together, these results indicate that myogenic tone is of greater magnitude in adult than fetal posterior communicating arteries, and that this difference is due to a greater calcium transient and a greater calcium sensitization response in adult compared to fetal arteries. Supported by NIH HD31226 and HL54120.

New Inotropic Agent-Levosimendan

Several clinical studies suggest substantial limitations of currently available positive inotropic substances, including beta1-adrenoceptor agonists and phosphodiesterase III inhibitors. Levosimendan, a myofilament calcium sensitizer with inotropic effects, increases myocardial performance without substantial changes in oxygen consumption and with neutral effects on heart rhythm. In addition, levosimendan has vasodilatory effects that are achieved by stimulation of adenosine triphosphate-dependent potassium channels. This review article briefly discusses the pharmacology of levosimendan and evaluates current available evidence to assess the safety and efficacy of levosimendan. (Korean J Anesthesiol 2006; 51: 519~27)

Cardiac and Renal Effects of Levosimendan, Arginine Vasopressin, and Norepinephrine in Lipopolysaccharide-treated Rabbits

Because sepsis-induced myocardial dysfunction related to sepsis is at least partially related to a decrease in cardiac myofilament response to calcium, the use of the new myofilament-calcium sensitizer, levosimendan, has been proposed. In addition, arginine vasopressin is increasingly proposed as a vasopressor in septic patients, although data on its effects on cardiac function are still scarce. The aim of the current study was to assess, invasively and noninvasively, whether levosimendan, arginine vasopressin, and norepinephrine, either alone or combined, may modify sepsis-induced myocardial dysfunction and renal hemodynamics. Thirty-six hours after lipopolysaccharide or saline administration, rabbits were studied either after slight sedation for echocardiography or after general anesthesia with sodium pentobarbital for the following measurements: aortic flow velocity and maximum acceleration of blood flow in the ascending aorta and renal macrocirculation and microcirculation. Levosimendan improved, within 30 min of administration, both maximum acceleration of blood flow by 20 +/- 12% (n = 8; P < 0.05) and left ventricular shortening fraction by a similar extent. Furthermore, low doses of arginine vasopressin markedly deteriorated cardiac function via an afterload-independent mechanism, even when animals were pretreated with levosimendan, whereas norepinephrine showed no detrimental effects on cardiac function. The study also showed that norepinephrine often improved renal medullary blood flow, whereas arginine vasopressin consistently decreased it. Levosimendan and norepinephrine both exert beneficial effects in endotoxemic animals and should be further explored in human sepsis trials.

Functional Consequences of Sarcomeric Protein Abnormalities in Failing Myocardium

Sarcomeric protein abnormalities have been recognized for many years in heart failure due to dilated cardiomyopathy (DCM). In contrast, virtually nothing is known about myofilament abnormalities in heart failure occurring in association with diastolic dysfunction. With the exception of sarcomeric protein mutations that cause DCM, the most important mechanism of myofilament dysfunction in DCM is probably altered post-translational modification, in particular the phosphorylation state of troponins I and T and possibly myosin light chain. Other modifications, including oxidation and glycation, may also play a role. Myosin heavy chain isoform switching occurs in human heart failure, but its functional significance is uncertain. Myofilament abnormalities contribute significantly to myocardial dysfunction in DCM, although their relative importance compared with abnormal calcium handling is debated. One consistent functional abnormality in DCM is increased myofilament calcium sensitivity of tension generation, which contributes to slowed or incomplete relaxation. More recently, decreases in the optimal frequency of myofilament work and power generation have been recognized. These changes may contribute to depression of the force-frequency relation in DCM. Altered mechanoenergetics constitute one of the most important manifestations of myofilament dysfunction in heart failure. DCM and hemodynamic overload are associated with more economical and efficient energy utilization by the contractile machinery, which may be protective of the myocardium. This change is strongly associated with depressed myofibrillar ATPase activity. We speculate that the effectiveness of mechanical therapies such as resynchronization may at least in part be related to improved mechanical function without loss of this mechanoenergetic advantage.