Reexamining the origin of the directionality of myosin V.

The nature of the conversion of chemical energy to directional motion in myosin V is examined by careful simulations that include two complementary methods: direct Langevin Dynamics (LD) simulations with a scaled-down potential that provided a detailed time-resolved mechanism, and kinetic equations solution for the ensemble long-time propagation (based on information collected for segments of the landscape using LD simulations and experimental information). It is found that the directionality is due to the rate-limiting ADP release step rather than the potential energy of the lever arm angle. We show that the energy of the power stroke and the barriers involved in it are of minor consequence to the selectivity of forward over backward steps and instead suggest that the selective release of ADP from a postrigor myosin motor head promotes highly selective and processive myosin V. Our model is supported by different computational methods-LD simulations, Monte Carlo simulations, and kinetic equations solution-as well as by structure-based binding energy calculations.

Tensiomyographic Assessment of Muscle Contractile Properties in 9- to 14-Year Old Children.

While there are numerous data on the skeletal muscle fiber type composition in adults, little is known about the changes in fiber type composition and contractile properties during maturational growth in children. Using noninvasive tensiomyography, we measured contraction time (Tc), an indirect estimate of the myosin heavy chain I (MHC-I) proportion, to assess the longitudinal changes of the biceps brachii (BB), biceps femois (BF), vastus lateralis (VL), and erector spinae (ES) muscles in 53 boys and 54 girls. The children were 9 years at the start of the study and returned for 5 follow-up measurements until the age of 14 years. The ES has the shortest and the BF has the longest Tc. The VL and ES of boys have shorter Tc than those of girls. When applying the relationship between proportion of MHC-I and Tc established in adults to children's TMG data, we found a slow-to-fast transition in the VL between, at least, the ages of 6 to 10 years, when it stabilized to adult proportions. Regular participation in sports was associated with a faster BF, but not in the VL. Our data represents a first non-invasive indication of the developmental changes in muscle fiber type composition in children.

Gokyo Khumbu/Ama Dablam Trek 2012: effects of physical training and high-altitude exposure on oxidative metabolism, muscle composition, and metabolic cost of walking in women.

PURPOSE: We investigated the effects of moderate-intensity training at low and high altitude on VO2 and QaO2 kinetics and on myosin heavy-chain expression (MyHC) in seven women (36.3 yy ± 7.1; 65.8 kg ± 11.7; 165 cm ± 8) who participated in two 12- to 14-day trekking expeditions at low (598 m) and high altitude (4132 m) separated by 4 months of recovery. METHODS: Breath-by-breath VO2 and beat-by-beat QaO2 at the onset of moderate-intensity cycling exercise and energy cost of walking (Cw) were assessed before and after trekking. MyHC expression of vastus lateralis was evaluated before and after low-altitude and after high-altitude trekking; muscle fiber high-resolution respirography was performed at the beginning of the study and after high-altitude trekking. RESULTS: Mean response time of VO2 kinetics was faster (P = 0.002 and P = 0.001) and oxygen deficit was smaller (P = 0.001 and P = 0.0004) after low- and high-altitude trekking, whereas ˙ QaO2 kinetics and Cw did not change. Percentages of slow and fast isoforms of MyHC and mitochondrial mass were not affected by low- and high-altitude training. After training altitude, muscle fiber ADP-stimulated mitochondrial respiration was decreased as compared with the control condition (P = 0.016), whereas leak respiration was increased (P = 0.031), leading to a significant increase in the respiratory control ratio (P = 0.016). CONCLUSIONS: Although training did not significantly modify muscle phenotype, it induced beneficial adaptations of the oxygen transport-utilization systems witnessed by faster VO2 kinetics at exercise onset.

Effect of Plyometrics on the Energy Cost of Running and MHC and Titin Isoforms.

UNLABELLED: Several training strategies such as plyometrics have been shown to improve running economy; however, its physiological basis remains elusive. PURPOSE: To examine the effect of plyometric training on the energy cost of running (ECR, J · kg(-1) · min(-1)), titin, and myosin heavy chain (MHC) isoforms. METHODS: Subjects were randomly assigned to a 6-wk plyometric treatment (P; n = 11) or control group (C; n = 11). Preintervention and postintervention outcomes included body composition, vertical jump, sit-and-reach, maximal oxygen consumption (VO2max), speed at onset of blood lactate, 3-km time trial performance, ECR, and a vastus lateralis muscle biopsy for protein analysis. RESULTS: Plyometric intervention resulted in improved time trial (P, 2.6% faster, P = 0.04; C, 1.6%, P = 0.17). VO2max improved in the P group (5.2%, P = 0.03), whereas the C group increased by 3.1% (P = 0.20). The ECR decreased in the P group as the result of 6 wk of plyometric training (P = 0.02 for stage 3), whereas it increased in the C group (P = 0.02 for stage 3). The ECR correlated strongly with performance at stages 2, 3, and 4 (r > 0.8, P < 0.001) independent of group. There was no significant main effect of group, time, or interaction on any of the protein isoforms analyzed. A negative correlation was found between the ECR at stage 7 and MHC IIa (r = -0.96, P < 0.001), and the ECR at stage 6 with titin isoform 1 (T1)/T2 ratio (r = -0.69, P = 0.007) independent of group. CONCLUSION: Six weeks of plyometric training improved running performance and the ECR despite no measurable changes in MHC and titin isoforms. However, higher MHC IIa and lower T1/T2 isoform ratios correlated to lower ECR.

Simultaneous assessment of glomerular filtration and barrier function in live zebrafish.

The zebrafish pronephros is a well-established model to study glomerular development, structure, and function. A few methods have been described to evaluate glomerular barrier function in zebrafish larvae so far. However, there is a need to assess glomerular filtration as well. In the present study, we extended the available methods by simultaneously measuring the intravascular clearances of Alexa fluor 647-conjugated 10-kDa dextran and FITC-conjugated 500-kDa dextran as indicators of glomerular filtration and barrier function, respectively. After intravascular injection of the dextrans, mean fluorescence intensities of both dextrans were measured in the cardinal vein of living zebrafish (4 days postfertilization) by confocal microscopy over time. We demonstrated that injected 10-kDa dextran was rapidly cleared from the circulation, became visible in the lumen of the pronephric tubule, quickly accumulated in tubular cells, and was detectably excreted at the cloaca. In contrast, 500-kDa dextran could not be visualized in the tubule at any time point. To check whether alterations in glomerular function can be quantified by our method, we injected morpholino oligonucleotides (MOs) against zebrafish nonmuscle myosin heavy chain IIA (zMyh9) or apolipoprotein L1 (zApol1). While glomerular filtration was reduced in zebrafish nonmuscle myosin heavy chain IIA MO-injected larvae, glomerular barrier function remained intact. In contrast, in zebrafish apolipoprotein L1 MO-injected larvae, glomerular barrier function was compromised as 500-kDa dextran disappeared from the circulation and became visible in tubular cells. In summary, we present a novel method that allows to simultaneously assess glomerular filtration and barrier function in live zebrafish.

Faster cross-bridge detachment and increased tension cost in human hypertrophic cardiomyopathy with the R403Q MYH7 mutation.

The first mutation associated with hypertrophic cardiomyopathy (HCM) is the R403Q mutation in the gene encoding ß-myosin heavy chain (ß-MyHC). R403Q locates in the globular head of myosin (S1), responsible for interaction with actin, and thus motor function of myosin. Increased cross-bridge relaxation kinetics caused by the R403Q mutation might underlie increased energetic cost of tension generation; however, direct evidence is absent. Here we studied to what extent cross-bridge kinetics and energetics are related in single cardiac myofibrils and multicellular cardiac muscle strips of three HCM patients with the R403Q mutation and nine sarcomere mutation-negative HCM patients (HCMsmn). Expression of R403Q was on average 41 ± 4% of total MYH7 mRNA. Cross-bridge slow relaxation kinetics in single R403Q myofibrils was significantly higher (P < 0.0001) than in HCMsmn myofibrils (0.47 ± 0.02 and 0.30 ± 0.02 s(-1), respectively). Moreover, compared to HCMsmn, tension cost was significantly higher in the muscle strips of the three R403Q patients (2.93 ± 0.25 and 1.78 ± 0.10 µmol l(-1) s(-1) kN(-1) m(-2), respectively) which showed a positive linear correlation with relaxation kinetics in the corresponding myofibril preparations. This correlation suggests that faster cross-bridge relaxation kinetics results in an increase in energetic cost of tension generation in human HCM with the R403Q mutation compared to HCMsmn. Therefore, increased tension cost might contribute to HCM disease in patients carrying the R403Q mutation.

Interspecific and intragenic differences in codon usage bias among vertebrate myosin heavy-chain genes.

Synonymous codon usage bias is a broadly observed phenomenon in bacteria, plants, and invertebrates and may result from selection. However, the role of selective pressures in shaping codon bias is still controversial in vertebrates, particularly for mammals. The myosin heavy-chain (MyHC) gene family comprises multiple isoforms of the major force-producing contractile protein in cardiac and skeletal muscles. Slow and fast genes are tandemly arrayed on separate chromosomes, and have distinct patterns of functionality and expression in muscle. We analyze both full-length MyHC genes (~5400 bp) and a larger collection of partial sequences at the 3' end (~500 bp). The MyHC isoforms are an interesting system in which to study codon usage bias because of their length, expression, and critical importance to organismal mobility. Codon bias and GC content differs among MyHC genes with regards to functional type, isoform, and position within the gene. Codon bias even varies by isoform within a species. We find evidence in favor of both chromosomal influences on nucleotide composition and selection against nonsense errors (SANE) acting on codon usage in MyHC genes. Intragenic variation in codon bias and elongation rate is significant, with a strong trend for increasing codon bias and elongation rate towards the 3' end of the gene, although the trend is dependent upon the degeneracy class of the codons. Therefore, patterns of codon usage in MyHC genes are consistent with models supporting SANE as a major force shaping codon usage.

Random walk of processive, quantum dot-labeled myosin Va molecules within the actin cortex of COS-7 cells.

Myosin Va (myoVa) is an actin-based intracellular cargo transporter. In vitro experiments have established that a single myoVa moves processively along actin tracks, but less is known about how this motor operates within cells. Here we track the movement of a quantum dot (Qdot)-labeled myoVa HMM in COS-7 cells using total internal reflectance fluorescence microscopy. This labeling approach is unique in that it allows myoVa, instead of its cargo, to be tracked. Single-particle analysis showed short periods (

Training induced decrease in oxygen cost of cycling is accompanied by down-regulation of SERCA expression in human vastus lateralis muscle.

We have examined the effect of 5 week cycling endurance training program on the sarco(endo)plasmic reticulum Ca2+ ATPase isoforms (SERCA1 and 2) and myosin heavy chain (MyHC) in the vastus lateralis muscle as well as on the oxygen uptake to power output ratio (VO2/PO) during incremental cycling. Fifteen untrained men performed an incremental cycling exercise until exhaustion before and after moderate intensity training. Muscle biopsies were taken from vastus lateralis before and after training program. Training resulted in higher (P = 0.048) maximal oxygen uptake (VO(2max)) as well as in higher power output reached at VO(2max) (P = 0.0001). Moreover, lower (P = 0.02) VO2/PO ratio determined during incremental moderate intensity cycling (i.e. 30-120 W) as well as lower (P = 0.003) VO2/PO ratio reached at VO(2max) were observed after the training. A significant down regulation of SERCA2 protein (P = 0.03) and tendency (P = 0.055) to lower SERCA1 content accompanied by lower (P<10(-4)) plasma thyroid hormone concentration, with no changes (P = 0.67) in MyHC composition in vastus lateralis muscle were found after training. We have concluded that the increase in mechanical efficiency of cycling occurring during first weeks of endurance training is not related to changes in MyHC composition but it may be due to down-regulation of SERCA pumps.

A DIGE approach for the assessment of rat soleus muscle changes during unloading: effect of acetyl-L-carnitine supplementation.

After hind limb suspension, a remodeling of postural muscle phenotype is observed. This remodeling results in a shift of muscle profile from slow-oxidative to fast-glycolytic. These metabolic changes and fiber type shift increase muscle fatigability. Acetyl-L-carnitine (ALCAR) influences the skeletal muscle phenotype of soleus muscle suggesting a positive role of dietary supplementation of ALCAR during unloading. In the present study, we applied a 2-D DIGE, mass spectrometry and biochemical assays, to assess qualitative and quantitative differences in the proteome of rat slow-twitch soleus muscle subjected to disuse. Meanwhile, the effects of ALCAR administration on muscle proteomic profile in both unloading and normal-loading conditions were evaluated. The results indicate a modulation of troponin I and tropomyosin complex to regulate fiber type transition. Associated, or induced, metabolic changes with an increment of glycolytic enzymes and a decreased capacity of fat oxidation are observed. These metabolic changes appear to be counteracted by ALCAR treatment, which restores the mitochondrial mass and decreases the glycolytic enzyme expression, suggesting a normalization of the metabolic shift observed in unloaded animals. This normalization is accompanied by a maintenance of body weight and seems to prevent a switch of fiber type.

Expression profiling reveals heightened apoptosis and supports fiber size economy in the murine muscles of mastication.

Distinctions between craniofacial and axial muscles exist from the onset of development and throughout adulthood. The masticatory muscles are a specialized group of craniofacial muscles that retain embryonic fiber properties in the adult, suggesting that the developmental origin of these muscles may govern a pattern of expression that differs from limb muscles. To determine the extent of these differences, expression profiling of total RNA isolated from the masseter and tibialis anterior (TA) muscles of adult female mice was performed, which identified transcriptional changes in unanticipated functional classes of genes in addition to those attributable to fiber type. In particular, the masseters displayed a reduction of transcripts associated with contractile and cytoskeletal load-sensing and anabolic processes, and heightened expression of genes associated with stress. Associated with these observations was a significantly smaller fiber cross-sectional area in masseters, significantly elevated load-sensing signaling (phosphorylated focal adhesion kinase), and increased apoptotic index in masseters compared with TA muscles. Based on these results, we hypothesize that masticatory muscles may have a fundamentally different strategy for muscle design, compared with axial muscles. Specifically there are small diameter fibers that have an attenuated ability to hypertrophy, but an increased propensity to undergo apoptosis. These results may provide insight into the molecular basis for specific muscle-related pathologies associated with masticatory muscles.

Altered tension cost in (TG(mREN-2)27) rats overexpressing the mouse renin gene.

The present study aimed to characterize cardiac hypertrophy induced by activation of the renin-angiotensin system in terms of functional alterations on the level of the contractile proteins, employing transgenic rats harboring the mouse renin gene (TGR(mREN2)27). Ca2+-dependent tension and myosin ATPase activity were measured in skinned fiber preparations obtained from TGR(mREN2)27 and from age-matched Sprague-Dawley rats (SPDR). Western blots for troponin I (TnI) and troponin T (TnT) were performed and the phosphorylation status of TnI were evaluated in myocardial preparations. TnT and myosin heavy chain (MHC) isoforms were analyzed by RT-PCR. The pCa/tension relationship was shifted to the right in TGR(mREN2)27 compared to SPDR as indicated by increased Ca2+-concentrations required for half maximal activation of tension (SPDR 5.80, 95% confidence limits 5.77-5.82 vs. TGR(mREN2)27 5.69, 95% confidence limits 5.67-5.72, pCa units), while maximal developed tension was unaltered. Even more pronounced was the shift in the relationship between pCa and myosin-ATPase (SPDR 6.01, 95% confidence limits 5.99-6.03 vs. TGR(mREN2)27 5.77, 95% confidence limits 5.73-5.79, pCa units). The maximal myosin-ATPase activity was reduced in TGR(mREN2)27 compared to SPDR, respectively (211.0 +/- 28.77 micromol ADP/s vs. 271.6 +/- 43.66 micromol ADP/s, P < 0.05). Tension cost (ATPase activity/tension) was significantly reduced in TGR(mREN2)27. The beta-MHC expression was significantly increased in TGR(mREN2)27. There was no isoform shift for TnT (protein and mRNA), as well as TnI, and no alteration of the phosphorylation of TnI in TGR(mREN2)27 compared to SPRD. The present study demonstrates that cardiac hypertrophy, induced by an activation of the renin-angiotensin system, leads to adapting alterations on the level of the contractile filaments, which reduce tension cost.

Assessment of contractility of purified smooth muscle cells derived from embryonic stem cells.

The aims of this study were to develop a method for deriving purified populations of contractile smooth muscle cells (SMCs) from embryonic stem cells (ESCs) and to characterize their function. Transgenic ESC lines were generated that stably expressed a puromycin-resistance gene under the control of either a smooth muscle alpha-actin (SMalphaAlpha) or smooth muscle-myosin heavy chain (SM-MHC) promoter. Negative selection, either overnight or for 3 days, was then used to purify SMCs from embryoid bodies. Purified SMCs expressed multiple SMC markers by immunofluorescence, immunoblotting, quantitative reverse transcription-polymerase chain reaction, and flow cytometry and were designated APSCs (SMalphaAlpha-puromycin-selected cells) or MPSCs (SM-MHC-puromycin-selected cells), respectively. Both SMC lines displayed agonist-induced Ca(2+) transients, expressed functional Ca(2+) channels, and generated contractile force when aggregated within collagen gels and stimulated with vasoactive agonists, such as endothelin-1, or in response to depolarization with KCl. Importantly, subcutaneous injection of APSCs or MPSCs subjected to 18 hours of puromycin selection led to the formation of teratomas, presumably due to residual contamination by pluripotent stem cells. In contrast, APSCs or MPSCs subjected to prolonged puromycin selection for 3 days did not form teratomas in vivo. These studies describe for the first time a method for generating relatively pure populations of SMCs from ESCs which display appropriate excitation and contractile responses to vasoactive agonists. However, studies also indicate the potential for teratoma development in ESC-derived cell lines, even after prolonged differentiation, highlighting the critical requirement for efficient methods of separating differentiated cells from residual pluripotent precursors in future studies that use ESC derivatives, whether SMC or other cell types, in tissue engineering applications.

Increased resistance to fatigue in creatine kinase deficient muscle is not due to improved contractile economy.

There has been speculation on the origin of the increased endurance of skeletal muscles in creatine kinase (CK)-deficient mice. Important factors that have been raised include the documented increased mitochondrial capacity and alterations in myosin heavy chain (MyHC) isoform composition in CK-deficient muscle. More recently, the absence of inorganic phosphate release from phosphocreatine hydrolysis in exercising CK-deficient muscle has been postulated to contribute to the lower fatigueability in skeletal muscle. In this study, we tested the hypothesis that the reported shift in MyHC composition to slower isoforms in CK-deficient muscle leads to a decrease in oxygen cost of twitch performance. To that aim, extensor digitorum longus (EDL) and soleus (SOL) muscles were isolated from wild-type (WT) and knock-out mice deficient in the cytoplasmic muscle-type and sarcomeric mitochondrial isoenzymes of CK, and oxygen consumption per twitch time-tension-integral (TTI) was measured. The results show that the adaptive response to loss of CK function does not involve any major change to contractile economy of skeletal muscle.

Myosin heavy chain composition and the economy of contraction in healthy and diseased human myocardium.

Changes in myosin heavy chain (MHC) isoform expression and protein composition occur during cardiac disease and it has been suggested that even a minor shift in MHC composition may exert a considerable effect on myocardial energetics and performance. Here an overview is provided of the cellular basis of the energy utilisation in cardiac tissue and novel data are presented concerning the economy of myocardial contraction in diseased atrial and ventricular human myocardium. ATP utilisation and force development were measured at various Ca(2+) concentrations during isometric contraction in chemically skinned atrial trabeculae from patients in sinus rhythm (SR) or with chronic atrial fibrillation (AF) and in ventricular muscle strips from non-failing donor or end-stage failing hearts. Contractile protein composition was analysed by one-dimensional gel electrophoresis. Atrial fibrillation was accompanied by a significant shift from the fast alpha-MHC isoform to the slow beta-MHC isoform, whereas both donor and failing ventricular tissue contained almost exclusively the beta-MHC isoform. Simultaneous measurements of force and ATP utilisation indicated that economy of contraction is preserved in atrial fibrillation and in end-stage human heart failure.

Myocardial contraction is 5-fold more economical in ventricular than in atrial human tissue.

OBJECTIVE: Cardiac energetics and performance depend on the expression level of the fast (alpha-) and slow (beta-) myosin heavy chain (MHC) isoform. In ventricular tissue, the beta-MHC isoform predominates, whereas in atrial tissue a variable mixture of alpha- and beta-MHC is found. In several cardiac diseases, the slow isoform is upregulated; however, the functional implications of this transition in human myocardium are largely unknown. The aim of this study was to determine the relation between contractile properties and MHC isoform composition in healthy human myocardium using the diversity in atrial tissue. METHODS: Isometric force production and ATP consumption were measured in chemically skinned atrial trabeculae and ventricular muscle strips, and rate of force redevelopment was studied using single cardiomyocytes. MHC isoform composition was determined by one-dimensional SDS-gel electrophoresis. RESULTS: Force development in ventricular tissue was about 5-fold more economical, but nine times slower, than in atrial tissue. Significant linear correlations were found between MHC isoform composition, ATP consumption and rate of force redevelopment. CONCLUSION: These results clearly indicate that even a minor shift in MHC isoform expression has considerable impact on cardiac performance in human tissue.

Skeletal muscle aging in F344BN F1-hybrid rats: II. Improved contractile economy in senescence helps compensate for reduced ATP-generating capacity.

We used a pump-perfused rat hind-limb preparation to compare young adult (YA: 8-9- month-old), late middle-aged (LMA: 28-29-month-old), and senescent (SEN: 36-month-old) rats at similar rates of convective O(2) delivery during a 4-minute contraction bout. We hypothesized that not only would VO(2) and lactate production be reduced, but also that contractile economy would be altered with aging. Peak tension was lower in LMA (42%) and SEN (71%) versus YA. VO(2) and lactate efflux was progressively lower with increasing age. Estimated adenosine triphosphate per N of force was increased in LMA (35%) and reduced in SEN (31%) versus YA. Myosin heavy chain (MHC) analysis by sodium dodecyl sulphate-polyacrylamide gel electrophoresis showed a lower MHC type IIb and higher MHC type IIa/IIx in SEN versus YA. Therefore, whereas contractile economy is impaired in LMA, it is improved in SEN, and this latter effect may be due in part to reduced type IIb MHC.

Depressed cardiac tension cost in experimental diabetes is due to altered myosin heavy chain isoform expression.

Cardiac disease in diabetes presents as impaired left ventricular contraction and relaxation; however, the mechanisms underlying contractile protein dysfunction during the progression of disease are unknown. Accordingly, we assessed Ca(2+)-dependent tension development and tension-dependent ATP consumption (tension cost) in a rat model early (6 wk) and late (12 wk) after the onset of diabetes (50 mg/kg iv streptozotocin) using mechanical force- and enzyme-coupled UV absorbance measurements. Myofilament Ca(2+) sensitivity and maximal tension were unchanged between groups at either time point. Cross-bridge cycling rate was significantly decreased in diabetes, as indexed by tension cost (early control 5.4 +/- 0.4 and early diabetes 4.2 +/- 0.3; and late control 6.0 +/- 0.2 and late diabetes 4.2 +/- 0.2; P < 0.05). Because rodent models of cardiac disease are confounded by altered myosin isoform distribution, myosin content was determined by SDS-PAGE and densitometry. The cardiac content of alpha-myosin in diabetes was decreased to 41% +/- 4.1 at 6 wk and 32.5% +/- 2.9 at 12 wk of diabetes (early control 77.8% +/- 3.3 and late control 73.6% +/- 2.5). Separate control experiments demonstrated a linear decrease in tension cost with decreased alpha-myosin content. Given this, the depression of tension cost in this rodent model of diabetes could be fully explained by the altered myosin isoform distribution.

Age-dependent changes in myosin composition correlate with enhanced economy of contraction in guinea-pig hearts.

1. The composition of myosin heavy chains (MHCs) was investigated in young (1- to 8-week-old) and mature (9- to 26-week-old) guinea-pigs using two monoclonal antibodies directed specifically against alpha-MHC and beta-MHC. In addition, maximum force and the rate of ATP consumption during isometric contraction were measured in chemically skinned trabeculae taken from the same hearts. 2. An age-dependent shift in the MHC composition was found. The alpha-MHC fraction decreased from 0.17 +/- 0.02 (mean +/- S.E.M.; n = 24) in young to 0.04 +/- 0.01 (n = 43) in mature hearts. This shift was correlated with a decrease in tension cost (i.e. ATP consumption per second per trabecula volume/force per cross-sectional area) from 4.1 +/- 0.2 mmol kN-1 m-1 s-1 (n = 23) in young to 2.5 +/- 0.1 mmol kN-1 m-1 s-1 (n = 57) in mature hearts. 3. From the results it follows that the slow beta-MHC isoform, which predominates in hearts of mature guinea-pigs, is about 5 times more economical than the fast alpha-MHC isoform. Calcium sensitivity of force and ATP consumption decreased with age, but stabilized within a few weeks after birth. The pronounced dependence of cardiac energetics on MHC composition should be taken into account in long-term studies of cardiac overload.