The mechanism of action of adrenaline on cardiac contractility in rat papillary muscles containing V1 and V3 isomyosins was analyzed during barium-activated contractures at 25 degrees C by frequency domain analysis using pseudo-random binary noise-modulated perturbations. The analysis characterizes a frequency (fmin) at which dynamic stiffness of a muscle is a minimum, a parameter that reflects the rate of cycling of crossbridges. We have previously shown that fmin for V1- and V3-containing papillary muscles were 2.1 +/- 0.2 Hz (mean +/- SD) (n = 10) and 1.1 +/- 0.2 Hz (n = 8), respectively, and that these values were independent of the level of activation. The present study's goal was to determine whether the inotropic action of adrenaline was associated with an increased rate of crossbridge cycling. The results show that a saturating dose of adrenaline increased fmin in V1 hearts by 49 +/- 2% (n = 11). The action on V3 hearts was significantly less; the increase in fmin was 26 +/- 2% (n = 6). The increase in fmin for V1 hearts was shown to be sensitive to the beta-blocking agent propranolol. These results suggest that adrenaline significantly increases the rate of crossbridge cycling by a beta-receptor-mediated mechanism. We conclude that the increased contractility of the heart in the presence of adrenaline arises not only from more complete activation of the contractile proteins but also from the increased rate at which each crossbridge can transduce energy.

Experiments were done on four-week-old rats, containing biochemically verified V1 only, and thyroidectomized adult rats, treated with propylthiouracil, verified to contain V3 only. Contracture tension was induced in isolated papillary muscles either by high potassium solution or 0.5 mmol l-1 Ba2+. Small amplitude length perturbations with peak-to-peak value not exceeding 0.15% L0 were applied to the activated muscle. Both the applied length perturbations and the corresponding resulting force changes were analysed by computer for dynamic stiffness and phase values. In order to reduce data acquisition time, pseudo-random binary noise length changes, rather than the conventional sinusoidal length changes, were used. The plot of the dynamic stiffness against frequency displays a minimum, akin to a resonance phenomenon. The frequency, fmin, at which this resonance occurs, reflects crossbridge kinetics. It was found that the fmin values for the two types of papillary muscles differed by a factor of two. Experiments were also done on chemically skinned muscles containing V1 or V3 isomyosin activated by different concentrations of either barium or calcium ions. It was found that fmin values of skinned fibres were higher than those obtained from intact fibres. However, for each type of muscle the fmin was independent of the activator used as well as the level of activation. The ratio of fmin for V3 to that for V1 remained the same as for intact preparations. We conclude that the difference in mechanical parameters did not arise a possible difference in excitation-contraction coupling mechanism, but rather is a difference in the dynamic properties of the two types of crossbridges.

Inotropic agents that increase the intracellular levels of cAMP have been shown to increase crossbridge turnover kinetics in intact rat ventricular muscle, as measured by the parameter f min (the frequency at which dynamic stiffness is minimum). These agents are also known to increase the level of phosphorylation of two candidate myofibrillar proteins: myosin binding protein C (MyBPC) and Troponin I (TnI), but have no effect on myosin light chain 2 phosphorylation (MyLC2). The aim of this study was to investigate whether the phosphorylation of TnI and/or MyBPC was responsible for the increase in crossbridge cycling kinetics (as captured by f min ) seen with the elevation of cAMP within cardiac tissue. Using barium-activated intact rat papillary muscle, we investigated the actions of isobutylmethylxanthine (IBMX), an inhibitor of cAMP-dependent phosphatase, which simulates the action of b-adrenergic agents, and the chemical phosphatase 2,3-butanedione monoxime (BDM), which has been shown to dephosphorylate a number of contractile proteins. The presence of 0.6 mM IBMX approximately doubled the f min value of intact rat papillary muscle. This action was unaffected by the addition of BDM. In the presence of IBMX and BDM, the level of phosphorylation of MyBPC was unchanged, that of MyLC2 was reduced to 60 % of control, yet that of TnI was markedly increased (to 30 % above control levels). We conclude that TnI phosphorylation, mediated by cAMP-dependent protein kinase A, is the molecular basis for the enhanced crossbridge cycling seen during b-adrenergic stimulation of the heart.

The molecular mechanism of inotropic action of endothelin was investigated in rat ventricular muscle by studying its effects on characteristics of isometric twitch, barium‐induced steady contracture and the level of incorporation of 32Pi into myosin light chain 2. Exposure of rat papillary muscle to endothelin caused an increase in isometric twitch force but did not alter the twitch‐time parameters. Endothelin did not significantly change the maximum contracture tension but did cause an increase in contracture tension at submaximal levels of activation, without changes in the tension‐to‐stiffness ratio and kinetics of attached cross‐bridges. Kinetics of attached cross‐bridges were deduced during steady contracture from complex‐stiffness values, and in particular from the frequency at which muscle stiffness assumes a minimum value, fmin. Endothelin did not alter fmin. Endothelin caused an increase in the level of incorporation of 32Pi into myosin light chain 2 without a concurrent change in the level of incorporation of 32Pi into troponin I. We conclude that the inotropic action of endothelin is not due to an increase in the kinetics of attached cross‐bridges, nor due to a change in the force per unit cross‐bridge, but may result from an increased divalent cation sensitivity caused by elevated myosin light chain 2 phosphorylation, resembling post‐tetanic potentiation in fast skeletal muscle fibres.

A high resolution, time efficient method is described for determining the complex modulus of activated striated muscle. In this method the applied oscillation of muscle length takes the form of pseudo-random binary noise, PRBN. As a time-domain signal, PRBN is an extension of the double step approach to include n steps; as a frequency-domain signal, PRBN has the properties of quasi-white noise. Fourier analysis of the PRBN length oscillations and the resulting interrupted tension transients gives rise to the complex modulus values. PRBN provides a practical demonstration of the conceptual link between time and frequency domain descriptions of strain sensitive dynamics. The method is demonstrated with intact rat papillary muscle, and glycerol extracted rabbit psoas muscle.