Depressed Frank-Starling mechanism in the left ventricular muscle of the knock-in mouse model of dilated cardiomyopathy with troponin T deletion mutation ΔK210.

It has been reported that the Frank-Starling mechanism is coordinately regulated in cardiac muscle via thin filament "on-off" equilibrium and titin-based lattice spacing changes. In the present study, we tested the hypothesis that the deletion mutation ΔK210 in the cardiac troponin T gene shifts the equilibrium toward the "off" state and accordingly attenuate the sarcomere length (SL) dependence of active force production, via reduced cross-bridge formation. Confocal imaging in isolated hearts revealed that the cardiomyocytes were enlarged, especially in the longitudinal direction, in ΔK210 hearts, with striation patterns similar to those in wild type (WT) hearts, suggesting that the number of sarcomeres is increased in cardiomyocytes but the sarcomere length remains unaltered.

Structure of the Ca(2+)-saturated C-terminal domain of scallop troponin C in complex with a troponin I fragment.

Troponin C (TnC) is the Ca(2+)-sensing subunit of troponin that triggers the contraction of striated muscles. In scallops, the striated muscles consume little ATP energy in sustaining strong contractile forces. The N-terminal domain of TnC works as the Ca(2+) sensor in vertebrates, whereas scallop TnC uses the C-terminal domain as the Ca(2+) sensor, suggesting that there are differences in the mechanism of the Ca(2+)-dependent regulation of muscles between invertebrates and vertebrates.

Sarcomere length-dependent Ca2+ activation in skinned rabbit psoas muscle fibers: coordinated regulation of thin filament cooperative activation and passive force.

In skeletal muscle, active force production varies as a function of sarcomere length (SL). It has been considered that this SL dependence results simply from a change in the overlap length between the thick and thin filaments. The purpose of this study was to provide a systematic understanding of the SL-dependent increase in Ca(2+) sensitivity in skeletal muscle, by investigating how thin filament "on-off" switching and passive force are involved in the regulation.

Depressed contractile performance and reduced fatigue resistance in single skinned fibers of soleus muscle after long-term disuse in rats.

Long-term disuse results in atrophy in skeletal muscle, which is characterized by reduced functional capability, impaired locomotor condition, and reduced resistance to fatigue. Here we show how long-term disuse affects contractility and fatigue resistance in single fibers of soleus muscle taken from the hindlimb immobilization model of the rat. We found that long-term disuse results in depression of caffeine-induced transient contractions in saponin-treated single fibers.

Biological actions of green tea catechins on cardiac troponin C.

Background And Purpose: Catechins, biologically active polyphenols in green tea, are known to have a protective effect against cardiovascular diseases. In this study, we investigated direct actions of green tea catechins on cardiac muscle function to explore their uses as potential drugs for cardiac muscle disease.

Experimental Approach: The effects of catechins were systematically investigated on the force-pCa relationship in skinned cardiac muscle fibres to determine their direct effects on cardiac myofilament contractility.

Regulatory mechanism of length-dependent activation in skinned porcine ventricular muscle: role of thin filament cooperative activation in the Frank-Starling relation.

Cardiac sarcomeres produce greater active force in response to stretch, forming the basis of the Frank-Starling mechanism of the heart. The purpose of this study was to provide the systematic understanding of length-dependent activation by investigating experimentally and mathematically how the thin filament "on-off" switching mechanism is involved in its regulation. Porcine left ventricular muscles were skinned, and force measurements were performed at short (1.

Titin and troponin: central players in the frank-starling mechanism of the heart.

The basis of the Frank-Starling mechanism of the heart is the intrinsic ability of cardiac muscle to produce greater active force in response to stretch, a phenomenon known as length-dependent activation. A feedback mechanism transmitted from cross-bridge formation to troponin C to enhance Ca(2+) binding has long been proposed to account for length-dependent activation. However, recent advances in muscle physiology research technologies have enabled the identification of other factors involved in length-dependent activation.

Protein kinase A-dependent modulation of Ca2+ sensitivity in cardiac and fast skeletal muscles after reconstitution with cardiac troponin.

Protein kinase A (PKA)-dependent phosphorylation of troponin (Tn)I represents a major physiological mechanism during beta-adrenergic stimulation in myocardium for the reduction of myofibrillar Ca2+ sensitivity via weakening of the interaction with TnC. By taking advantage of thin filament reconstitution, we directly investigated whether or not PKA-dependent phosphorylation of cardiac TnI (cTnI) decreases Ca2+ sensitivity in different types of muscle: cardiac (porcine ventricular) and fast skeletal (rabbit psoas) muscles. PKA enhanced phosphorylation of cTnI at Ser23/24 in skinned cardiac muscle and decreased Ca2+ sensitivity, of which the effects were confirmed after reconstitution with the cardiac Tn complex (cTn) or the hybrid Tn complex (designated as PCRF; fast skeletal TnT with cTnI and cTnC).

Propyl gallate, a strong antioxidant, increases the Ca2+ sensitivity of cardiac myofilament.

Ca(2+) sensitizers are cardiotonic agents that directly increase the Ca(2+) sensitivity of cardiac myofilament. To find a novel Ca(2+) sensitizer, we have screened a group of phenolic compounds by examining their effects on the Ca(2+)-dependent force generation in cardiac muscle fibers. We found that propyl gallate, a strong antioxidant, increased the Ca(2+) sensitivity of cardiac myofilament in a dose-dependent and reversible manner.

Structure-function relationships of molluscan troponin T revealed by limited proteolysis.

Molluscan troponin regulates muscle contraction through a novel Ca(2+)-dependent activating mechanism associated with Ca(2+)-binding to the C-terminal domain of troponin C. To elucidate the further details of this regulation, we performed limited chymotryptic digestion of the troponin complex from akazara scallop striated muscle. The results indicated that troponin T is very susceptible to the protease, compared to troponin C or troponin I.

Troponin and titin coordinately regulate length-dependent activation in skinned porcine ventricular muscle.

We investigated the molecular mechanism by which troponin (Tn) regulates the Frank-Starling mechanism of the heart. Quasi-complete reconstitution of thin filaments with rabbit fast skeletal Tn (sTn) attenuated length-dependent activation in skinned porcine left ventricular muscle, to a magnitude similar to that observed in rabbit fast skeletal muscle. The rate of force redevelopment increased upon sTn reconstitution at submaximal levels, coupled with an increase in Ca2+ sensitivity of force, suggesting the acceleration of cross-bridge formation and, accordingly, a reduction in the fraction of resting cross-bridges that can potentially produce additional active force.

Disuse-induced preferential loss of the giant protein titin depresses muscle performance via abnormal sarcomeric organization.

Persistent muscle weakness due to disuse-associated skeletal muscle atrophy limits the quality of life for patients with various diseases and individuals who are confined to bed. Fibers from disused muscle exhibit a marked reduction in active force production, which can exacerbate motor function, coupled with the well-known loss of muscle quantity. Despite recent understanding of the signaling pathways leading to the quantity loss, the molecular mechanisms of the depressed qualitative performance still remain elusive.

Troponin: regulatory function and disorders.

Study of the molecular biology of the calcium regulation of muscle contraction was initiated by Professor Ebashi's discovery of a protein factor that sensitized actomyosin to calcium ions. This protein factor was separated into two proteins: tropomyosin and a novel protein named troponin. Troponin is a Ca(2+)-receptive protein for the Ca(2+)-regulation of muscle contraction and, in association with tropomyosin, sensitizes actomyosin to Ca(2+).

Knock-in mouse model of dilated cardiomyopathy caused by troponin mutation.

We created knock-in mice in which a deletion of 3 base pairs coding for K210 in cardiac troponin (cTn)T found in familial dilated cardiomyopathy patients was introduced into endogenous genes. Membrane-permeabilized cardiac muscle fibers from mutant mice showed significantly lower Ca(2+) sensitivity in force generation than those from wild-type mice. Peak amplitude of Ca(2+) transient in cardiomyocytes was increased in mutant mice, and maximum isometric force produced by intact cardiac muscle fibers of mutant mice was not significantly different from that of wild-type mice, suggesting that Ca(2+) transient was augmented to compensate for decreased myofilament Ca(2+) sensitivity.

Crystallization and preliminary X-ray analysis of the Ca2+-bound C-terminal lobe of troponin C in complex with a troponin I-derived peptide fragment from Akazara scallop.

Troponin C (TnC) is the Ca(2+)-binding component of troponin and triggers muscle contraction. TnC of the invertebrate Akazara scallop can bind only one Ca(2+) at the C-terminal EF-hand motif. Recombinant TnC was expressed in Escherichia coli, purified, complexed with a 24-residue synthetic peptide derived from scallop troponin I (TnI) and crystallized.

Drastic Ca2+ sensitization of myofilament associated with a small structural change in troponin I in inherited restrictive cardiomyopathy.

Six missense mutations in human cardiac troponin I (cTnI) were recently found to cause restrictive cardiomyopathy (RCM). We have bacterially expressed and purified these human cTnI mutants and examined their functional and structural consequences. Inserting the human cTnI into skinned cardiac muscle fibers showed that these mutations had much greater Ca2+-sensitizing effects on force generation than the cTnI mutations in hypertrophic cardiomyopathy (HCM).

Comparative studies on the functional roles of N- and C-terminal regions of molluskan and vertebrate troponin-I.

Vertebrate troponin regulates muscle contraction through alternative binding of the C-terminal region of the inhibitory subunit, troponin-I (TnI), to actin or troponin-C (TnC) in a Ca(2+)-dependent manner. To elucidate the molecular mechanisms of this regulation by molluskan troponin, we compared the functional properties of the recombinant fragments of Akazara scallop TnI and rabbit fast skeletal TnI. The C-terminal fragment of Akazara scallop TnI (ATnI(232-292)), which contains the inhibitory region (residues 104-115 of rabbit TnI) and the regulatory TnC-binding site (residues 116-131), bound actin-tropomyosin and inhibited actomyosin-tropomyosin Mg-ATPase.

Functional importance of Ca2+-deficient N-terminal lobe of molluscan troponin C in troponin regulation.

Ca(2+)-binding sites I and II in the N-terminal lobe of molluscan troponin C (TnC) have lost the ability to bind Ca(2+) due to substitutions of the amino acid residues responsible for Ca(2+) liganding. To evaluate the functional importance of the Ca(2+)-deficient N-terminal lobe in the Ca(2+)-regulatory function of molluscan troponin, we constructed chimeric TnCs comprising the N-terminal lobes from rabbit fast muscle and squid mantle muscle TnCs and the C-terminal lobe from akazara scallop TnC, TnC(RA), and TnC(SA), respectively. We characterized their biochemical properties as compared with those of akazara scallop wild-type TnC (TnC(AA)).

SCH00013, a novel Ca(2+) sensitizer with positive inotropic and no chronotropic action in heart failure.

We investigated the effects of the agent SCH00013 on Ca(2+)-induced force generation in rabbit skinned cardiac muscle fibers and in vivo cardiac function in high-pacing-induced heart failure dogs. The Ca(2+)-induced force generation in skinned cardiac muscle fibers was determined at pH 6.2 - 7.

Cardiac troponin T mutation R141W found in dilated cardiomyopathy stabilizes the troponin T-tropomyosin interaction and causes a Ca2+ desensitization.

A missense mutation R141W in the strong tropomyosin-binding region of cardiac troponin T (cTnT) has recently been reported to cause dilated cardiomyopathy (DCM), following the first report of a DCM-causing deletion mutation DeltaK210. To clarify the molecular mechanism for the pathogenesis of DCM caused by this novel mutation in cTnT gene, functional analyses were made on the recombinant human cTnT mutant proteins. Exchanging human wild-type and mutant cTnTs into rabbit skinned cardiac muscle fibers revealed that R141W mutation resulted in a decrease in the Ca(2+) sensitivity of force generation, as in the case of DeltaK210 mutation lying outside the strong tropomyosin-binding region.

[Calcium regulation by troponin and its genetic disorder in striated muscle contraction].

Contraction of skeletal and cardiac muscles is regulated by Ca2+ through specific Ca(2+)-regulatory proteins, troponin and tropomyosin, located in the thin filament. Troponin is a Ca2+ receptive protein consisting of three differet components, troponins C, I, and T. The essential mechanisms of Ca(2+)-regulation are the inhibition of contractile interaction between myosin and actin by the inhibitory action of troponin I and the reversal of the inhibition by troponin I through the Ca(2+)-binding to troponin C.