Twitch and tetanic tension during culture of mature Xenopus laevis single muscle fibres

Investigation of the mechanisms of muscle adaptation requires independent control of the regulating factors. The aim of the present study was to develop a serum-free medium to culture mature single muscle fibres of Xenopus laevis. As an example, we used the culture system to study adaptation of twitch and tetanic force characteristics, number of sarcomeres in series and fibre cross-section. Fibres dissected from m. iliofibularis (n = 10) were kept in culture at a fibre mean sarcomere length of 2.3 µm in a culture medium without serum. Twitch and tetanic tension were determined daily. Before and after culture the number of sarcomeres was determined by laser diffraction and fibre cross-sectional area (CSA) was determined by microscopy. For five fibres twitch tension increased during culture and tetanic tension was stable for periods varying from 8 to 14 days (‘stable fibres’), after which fibres were removed from culture for analysis. Fibre CSA and the number of sarcomeres in series remained constant during culture. Five other fibres showed a substantial reduction in twitch and tetanic tension within the first five days of culture (‘unstable fibres’). After 7–9 days of culture, three of these fibres died. For two of the unstable fibres, after the substantial force reduction, twitch and tetanic tension increased again. Finally at day 14 and 18 of culture, respectively, the tensions attained values higher than their original values. For stable fibres, twitch contraction time, twitch half-relaxation time and tetanus 10%-relaxation time increased during culture. For unstable fibres these parameters fluctuated. For all fibres the stimulus threshold fluctuated during the first two days, and then remained constant, even for the fibres that were cultured for at least two weeks. It is concluded that the present culture system for mature muscle fibres allows long-term studies within a well-defined medium. Unfortunately, initial tetanic and twitch force are poor predictors of the long-term behaviour of the fibres

Intrinsic cardiac adrenergic (ICA) cell density and MAO-A activity in failing rat hearts

The efficiency (work/oxygen consumption) of isolated papillary muscles from failing hearts is reduced. We investigated whether this can be due to an increase of intrinsic cardiac adrenergic (ICA) cell density. The number of ICA cells in the septum and both ventricular walls was determined by tyrosine hydroxylase immunohistochemistry in rats with monocrotaline-induced pulmonary hypertension. We found that the number of ICA cells is about 200,000 per ,rat heart. ICA cell density was significantly lower in right ventricular myocardium of hypertrophied hearts (P<0.01). MAO-A enzyme histochemistry and inhibition experiments with clorgyline in papillary muscles were performed to localize the enzyme and to determine its oxygen consumption. Upregulation of MAO-A was found in the right ventricular wall and papillary muscles of failing hearts (P = 0.018). A positive correlation between ICA cell density and MAO-A activity was absent. Clorgyline (2 μM) decreased the basal rate of oxygen consumption of right ventricular papillary muscles by 65 μM O2/s (P = 0.027). This rate can only be maintained for several seconds judging from the catecholamine content of the preparations reported previously. High ICA cell activity rather than density and/or recycling of oxidized catecholamines are discussed as alternative explanations for the low myocardial efficiency in experimental pulmonary hypertension

On-off asymmetries in oxygen consumption kinetics of single Xenopus laevis skeletal muscle fibres suggest higher-order control

The mechanisms controlling skeletal muscle oxygen consumption (V(o)₂) during exercise are not well understood. We determined whether first-order control could explain V(o)₂kinetics at contractions onset (V(o)₂(on)) and cessation (V(o)₂off)) in single skeletal muscle fibres differing in oxdidative capacity, and across stimulation intensities up to V(o)₂(max). Xenopus laevis fibres (n = 21) were suspended in a sealed chamber with a fast response P(o)₂ electrode to measure V(o)₂ every second before, during and after stimulated isometric contractions. A first-order model did not well characterize on-transient V(o)₂ kinetics. Including a time delay (TD) in the model provided a significantly improved characterization than a first-order fit without TD (F-ratio; P < 0.05), and revealed separate 'activation' and 'exponential' phases in 15/21 fibres contracting at V(o)₂(max) (mean ± SD TD: 14 ± 3s). On-transient kinetics (τV(o)₂(on)) was weakly and linearly related to V(o)₂(max) (R² = 0.271, P = 0.015). Off-transient kinetics, however, were first-order, and τV(o)₂(off) was greater in low-oxidative (V(o)₂max < 0.05 nmol mm⁻³s⁻¹ than high-oxidative fibres (V(o)₂(max > 0.10 nmol mm ⁻³ s⁻¹; 170 ± 70 vs. 29 ± 6 s, P < 0.001). 1/ τV(o)₂(off) was proportional to V(o)₂(max) (R² = 0.727, P < 0.001), unlike in the on-transient. The calculated oxygen deficit was larger (P < 0.05) than the post-contraction volume of consumed oxygen at all intensities except V(o)₂(max). These data show a clear dissociation between the kinetic control of V(o)₂at the onset and cessation of contractions and across stimulation intensities. More complex models are therefore required to understand the activation of mitochondrial respiration in skeletal muscle at the start of exercise

Skeletal muscle capillarization and oxidative metabolism in healthy smokers

We investigated whether the lower fatigue resistance in smokers than in nonsmokers is caused by a compromised muscle oxidative metabolism. Using calibrated histochemistry, we found no differences in succinate dehydrogenase (SDH) activity, myoglobin concentration, or capillarization in sections of the vastus lateralis muscle between smokers and nonsmokers. The relationship between fatigue resistance and SDH activity in nonsmokers (r = 0.93; p = 0.02) is absent in smokers. This indicates that the lower muscle fatigue resistance of smokers can likely be attributed to causes other than differences in oxidative metabolism and capillarization. © 2008 NRC