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Abnormal respiratory muscle function is thought to contribute to breathlessness and exercise intolerance in heart failure but little is known about possible alterations in the function of such muscle. We have measured tetanic force and intracellular Ca(2+) concentration ([Ca(2+)](i)) in isolated, arterially perfused hemidiaphragm preparations from a rabbit coronary artery ligation model of heart failure. Increasing stimulation frequency (10-100 Hz) caused a progressive increase of force and [Ca(2+)](i) in control preparations, whereas force and [Ca(2+)](i) only increased between 10 and 25 Hz stimulation (decreasing at higher frequencies) in preparations from ligated animals. Cyclopiazonic acid produced a dose-dependent shift in the relationship between stimulation frequency and [Ca(2+)](i) in control preparations that was similar to the shift observed in the diaphragm of coronary-ligated animals. These data indicate that the in vitro contractile characteristics of the diaphragm are significantly altered in our model and that altered [Ca(2+)](i) regulation contributes to the reduced diaphragm strength observed in heart failure.

This study aimed to determine structural alterations occurring in cardiac myofilaments after exogenous application of oxidants and the effects of oxidants on contractile protein function in a rabbit coronary artery ligation model of heart failure. Myocardial "stiffness" was higher in the ligated animals (Lig) than sham-operated controls (Sh, 4.9+/-1.5 versus 1.6+/-0.8 mN.mm-1). Superoxide anion (O2-) exposure decreased active stiffness in both groups, whereas hypochlorous acid (HOCl) had no effect in Lig but increased stiffness in Sh. Resting stiffness was higher in Lig than Sh (0.6+/-0.2 versus 0.2+/-0.1 mN.mm-1), remaining unchanged after O2- exposure but increasing after HOCl in both groups. The frequency at minimum stiffness was lower in Lig than Sh (0.9+/-0.2 versus 1. 7+/-0.6 Hz) and was reduced in both groups after oxidant exposure. Myofilament calcium sensitivity (pCa50) was not altered by O2- in Sh but increased in Lig (pCa50 increased from 5.41+/-0.05 to 5.56+/-0. 06). Protease contamination in the xanthine oxidase used to generate O2- did not affect myofilament ultrastructure at the concentrations used here. These data demonstrate that contractile proteins from "failed" myocardium have a similar response to exogenously applied oxidants as controls and that application of protease-contaminated xanthine oxidase system does not degrade the contractile protein structure.

Calcium uptake by cardiac sarcoplasmic reticulum (SR) is reported to be reduced in heart failure in the human and in a number of animal models. However, the majority of studies have examined end-stage heart failure in the human and few animal studies have taken account of the duration and severity of left ventricular dysfunction. In this study we have compared SR Ca2+ loading in a haemodynamically assessed, coronary artery ligation model of heart failure at 8 and 15 weeks after ligation. Trabeculae were isolated from the right ventricle and mounted for isometric tension measurement. They were treated with saponin to permeabilize the sarcolemma but retain SR function and bathed in a mock intracellular solution including adenosine triphosphate (ATP) and buffered Ca2+. Caffeine was used to release Ca2+ from the SR. The amplitude of the caffeine-induced contracture was used as a quantitative gauge of the Ca2+ content of the SR. Eight weeks after ligation, trabeculae demonstrated enhanced SR Ca2+ uptake as manifest by larger caffeine-induced contractures (e.g. 200 nM [Ca2+], 120 s loading - 38.2+/-9.2 versus 67.3+/-10.1% of maximum Ca2+-activated force, FCa, max, P=0.03). At 15 weeks, trabeculae from ligated hearts were not significantly different from controls with SR Ca2+ loading returning to control levels (e.g. 200 nM [Ca2+], 120 s loading - 47.3+/-9.6 versus 30.2+/-12.8% FCa, max, P=0.12). These data suggest that SR Ca2+ loading may increase in the early stages of heart failure and fall back to normal with an increasing duration of left ventricular dysfunction. Increased incidence of spontaneous Ca2+ release observed from the SR at 8 weeks and not at 15 weeks may represent an arrhythmogenic mechanism specific to the early phase of heart failure.

Cardiac sarcoplasmic reticulum (SR) Ca(2+)-loading ability was assessed in a coronary artery ligation model of heart failure. Heart failure was produced in New Zealand White rabbits by ligation of the left marginal coronary artery. Sham-operated animals were used as controls. After hemodynamic and echocardiographic assessment 8 wk after coronary ligation, a free-running trabecula was isolated from the left or right ventricle, mounted for isometric tension measurement, and permeabilized with the chemical skinning agent saponin, leaving the SR functionally intact. The SR was Ca2+ loaded by exposure of the preparation to a mock intracellular solution with a Ca2+ concentration ([Ca2+]) of 150-300 nM. The amplitude of the caffeine-induced contracture was used as a measure of Ca2+ loaded by the SR. The same preparation was then treated with Triton X-100 to disrupt all cell membranes, and Ca2+ sensitivity {expressed as [Ca2+] required to produce 50% of maximal activation (pCa50)} of isometric tension production and maximum Ca2+ activated force (Cmax) were measured. Ligated animals demonstrated enhanced SR Ca(2+)-loading ability that correlated with the degree of left ventricular dysfunction. Enhanced SR Ca2+ loading was associated with evidence of SR Ca2+ overload revealed as spontaneous tension oscillations. Cmax and pCa50 were not significantly different from controls. Increased SR Ca(2+)-loading ability may predispose the SR to Ca2+ overload and could contribute to both contractile dysfunction and arrhythmogenesis in heart failure.