Mechanical and energetic consequences of HCM-causing mutations.

Hypertrophic cardiomyopathy (HCM) was the first inherited heart disease to be characterized at the molecular genetic level with the demonstration that it is caused by mutations in genes that encode different components of the cardiac sarcomere. Early functional in vitro studies have concluded that HCM mutations cause a loss of sarcomere mechanical function. Hypertrophy would then follow as a compensatory mechanism to raise the work and power output of the affected heart.

Effects of chronic atrial fibrillation on active and passive force generation in human atrial myofibrils.

Rationale: Chronic atrial fibrillation (cAF) is associated with atrial contractile dysfunction. Sarcomere remodeling may contribute to this contractile disorder.

Objective: Here, we use single atrial myofibrils and fast solution switching techniques to directly investigate the impact of cAF on myofilament mechanical function eliminating changes induced by the arrhythmia in atrial myocytes membranes and extracellular components.

The familial hypertrophic cardiomyopathy-associated myosin mutation R403Q accelerates tension generation and relaxation of human cardiac myofibrils.

The R403Q mutation in beta-myosin heavy chain was the first mutation to be identified as responsible for familial hypertrophic cardiomyopathy (FHC). In spite of extensive work on the functional sequelae of this mutation, the mechanism by which the mutant protein causes the disease has not been definitely identified. Here we directly compare contraction and relaxation mechanics of single myofibrils from left ventricular samples of one patient carrying the R403Q mutation to those from a healthy control heart.

Tension generation and relaxation in single myofibrils from human atrial and ventricular myocardium.

Fast solution switching techniques in single myofibrils offer the opportunity to dissect and directly examine the sarcomeric mechanisms responsible for force generation and relaxation. The feasibility of this approach is tested here in human cardiac myofibrils isolated from small samples of atrial and ventricular tissue. At sarcomere lengths between 2.

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.

Impaired diastolic function after exchange of endogenous troponin I with C-terminal truncated troponin I in human cardiac muscle.

The specific and selective proteolysis of cardiac troponin I (cTnI) has been proposed to play a key role in human ischemic myocardial disease, including stunning and acute pressure overload. In this study, the functional implications of cTnI proteolysis were investigated in human cardiac tissue for the first time. The predominant human cTnI degradation product (cTnI(1-192)) and full-length cTnI were expressed in Escherichia coli, purified, reconstituted with the other cardiac troponin subunits, troponin T and C, and subsequently exchanged into human cardiac myofibrils and permeabilized cardiomyocytes isolated from healthy donor hearts.

Streptomycin and intracellular calcium modulate the response of single guinea-pig ventricular myocytes to axial stretch.

We tested the hypothesis that both stretch-activated channels (SACs) and intracellular calcium ([Ca(2+)](i)) are important in the electrical response of single guinea-pig ventricular myocytes to axial stretch. Myocytes were attached to carbon fibre transducers and stretched, sarcomere length increased by approximately 9 %, and there was a prolongation of the action potential duration. Streptomycin, a blocker of SACs, had no effect upon the shortening, [Ca(2+)](i) transients or action potentials of electrically stimulated, unstretched myocytes, at a concentration of 50 microM, but at 40 microM, prevented any stretch-induced increase in action potential duration.

Evidence that phosphate release is the rate-limiting step on the overall ATPase of psoas myofibrils prevented from shortening by chemical cross-linking.

It has been suggested that the mechanical condition determines the rate-limiting step of the ATPase of the myosin heads in fibers: when fibers are isometrically contracting, the ADP release kinetics are rate-limiting, but as the strain is reduced and the fibers are allowed to shorten, the ADP release kinetics accelerate and P(i) release becomes rate-limiting. We have put this idea to the test with myofibrils as a model because with these both mechanical and chemical kinetic measurements are possible. With relaxed or rapidly shortening myofibrils, P(i) release is rate-limiting and (A)M.

Effects of streptomycin sulphate on I(CaL), I(Kr) and I(Ks) in guinea-pig ventricular myocytes.

In single guinea pig ventricular myocytes, streptomycin sulphate (streptomycin) reduced intracellular Ca(2+) transients (IC(50) 1.9 mM) and contractility (IC(50) 1.0 mM), 2 mM streptomycin prolonged the action potential.