Temperature and amplitude dependence of tension transients in glycerinated skeletal and insect fibrillar muscle.

1. Quick stretches and releases were applied to small bundles of glycerinated fibres of rabbit psoas and insect fibrillar flight muscle. The resulting tension changes were recorded at various temperatures and amplitudes of length change. The results from the two preparations had many features in common. At temperatures near 0 degrees C the asymmetry of the initial tension recovery after stretch and release originally reported in living frog fibres by Huxley & Simmons (1971 alpha) was very obvious. 2. The complete tension course could be described as an elastic change occurring simultaneously with the length change followed by recovery consisting of the sum of a number of exponential terms. These terms usually corresponded to the phases discernible without curve fitting, but in some cases a monotonic rise or fall of tension was seen to consist of two components only after curve fitting. 3. After either stretch or release there was a phase of rapid tension recovery towards the value before the length change. The rate constant of this phase increased as the amplitude of stretch or release was increased to about 2 nm/half sarcomere. At higher amplitudes it remained nearly constant 4. At temperatures near 0 degrees C there was a second and much slower continuation of the recovery after stretch. The rate constant of this second phase was much more sensitive to temperature than that of the first phase and it became slower with increasing amplitude of stretch. As the temperature was raised the speed of the second phase approached the speed of the first phase so that at room temperatures the initial tension recovery after stretch and release was nearly symmetrical. 5. Under many conditions these processes were followed by a change in the opposite direction, the 'delayed tension' described by earlier workers. This third phase of tension change had about the same temperature sensitivity as the second phase of the recovery seen after stretch. The tension due to stretch activation was not maintained in rabbit muscle, resulting in a fourth possible phase, a recovery of tension towards the value before the length change. This was absent or of low amplitude in insect flight muscle. 6. We interpret these tension changes on the basis of an extension of the non-linear model described by White & Thorson (1972). The elastic tension change and the initial fast recovery are both supposed to be properties of the attached cross-bridges, whilst the slower recovery is considered to be due to the detachment of cross-bridges which happened to be attached at the instant the length change was applied. The delayed tension reflects the approach to equilibrium of the number of attached bridges, changed by an effect of muscle length on the attachment rate. The fact that the delayed tension is not maintained in rabbit psoas muscle may be due to the effect of length on attachment rate being transitory.     

A 10 μs component in the tension transients of isolated intact skeletal muscle fibres of the frog

The transients of the tension and the angle of a first order diffraction line of isolated single intact muscle fibres of theM. lumbricalis IV of the foot resulting from rapid length changes have been measured. Furthermore simulations of the tension transients, taking the resonance frequency of the force transducer, the inertia of the fibre and the influence of the surrounding fluid into account, are presented. The tension transients could be simulated by a system of continuous elements with undamped elasticity —Young's modulusE ₁=1.8×10⁸ N/m² — in series with elasticityE ₂=5.4×10⁷ N/m² parallel to a damping element — coefficient of damping =2,300 Ns/m²; yielding a relaxation time of 10 s. The Young's modulus of the undamped series elasticityE ₁ implies that a sudden shortening of 2 nm per half sarcomere should reduce the active tension to zero. The results obtained from the diffraction pattern recording show that the displacement in the fibre is in accordance with the relative length change caused by the displacement generator. It is therefore believed that the tendon and tendon-muscle junction are sufficiently stiff to allow ascribing the above-mentioned elasticity and damping to the fibre itself.     

The recovery of tension in transients during steady lengthening of frog muscle fibres

The tension transients following step length changes imposed on tetanized muscle fibres during the steady phase of force response to lengthening were determined at different velocities. At low velocities the early partial recovery after a step was smaller and slower than under isometric conditions, while the speed of the final total recovery was faster. The degree of depression of the early recovery and the speed of the final recovery increased with the lengthening velocity. At a given lengthening velocity the speed of the total recovery depended on size and direction of the steps, increasing from the region of the larger releases to that of the larger stretches. The changes in the early partial recovery are explained qualitatively by the theory of Huxley and Simmons (1971), while the changes in the speed of the final recovery are explained by assuming that detachment of cross-bridges is negligible until a certain range of cross-bridge strain is reached, and then increases rapidly. It is also necessary to assume that cross-bridges detached in this way re-attach much more rapidly than when they detach on completion of their cycle during shortening.     

Effects of in vivo-like activation frequency on the length-dependent force generation of skeletal muscle fibre bundles

It is known that a range of firing frequencies can be observed during in vivo muscle activity, yet information is lacking as to how different in vivo-like frequencies may affect force generation of skeletal muscle. This study examined the effects of constant (CSF, constant within one contraction) and decreasing stimulation frequencies (DSF) on mean sarcomere length-force characteristics of rat gastrocnemius medialis fibre bundles. The CSF resulted in an optimal mean sarcomere length (l _SO) of 2.30 (SEM 0.02), 2.46 (SEM 0.03), 2.76 (SEM 0.03) and more than 2.99 (SEM 0.07)?μm, for 100, 50, 30 and 15 Hz, respectively. Compared to 100-Hz stimulation, both l _SO and the ascending limb of the relationship significantly shifted to higher lengths with lower frequencies. No shift was encountered for the initial part of the descending limb. The DSF reduced the frequency-induced shift to higher mean lengths [l _SO 2.33 (SEM 0.02), 2.52 (SEM 0.08) and more than 2.92 (SEM 0.10)?μm, respectively, for 50, 30 and 15?Hz]. No effect of activation time on length-force characteristics was observed. It was concluded from these studies that the frequency and history of stimulation is a major determinant of the length-force characteristics of muscle fibre bundles, and should be taken into account when analysing animal and human locomotion. The previously observed frequency-induced shift in whole muscle length-force relationship resides mainly at the level of fibre bundles.     

The control of tension and shortening in cardiac and skeletal muscle

The development of tension and shortening in cardiac and skeletal muscle exhibits phenomena suggestive of control by feedback mechanisms at the molecular level. Two of these, presented here, may have surprising implications for the cross-bridge theory of muscular contraction. It can be argued, from the basis of the cross-bridge theory, that the extended muscle is unstable against further imposed stretch by reason of a corresponding decrease of generated tension. Certain measurements of static tension versus muscle length appear to corroborate this conclusion. On the other hand, there is direct evidence for the dynamic stability of muscle at longer lengths. The cross-bridge theory may be incorrect in this regard, and cross-bridges may contain a negative feedback mechanism; but it may be the case that the contradiction arises out of inappropriately extrapolating the static length-tension data to a dynamic situation. By monitoring the sarcomere length of a contracting muscle, it can be seen that shortening does not proceed smoothly, but in more or less discontinuous jumps. This “stepwise” shortening shows evidence of synchronization across a region of a muscle, and suggests some positive feedback process at work. Some speculations are offered as to possible mechanisms, including load-dependent signalling, diffusion, cooperative effects, and repeated cycles of substrate consumption, concomitant with shortening, alternating with a “catch-up” period of substrate production.     

The consequences of fibre heterogeneity on the force-velocity relation of skeletal muscle

The consequences of fibre heterogeneity on the collective force-velocity properties of bundles of parallel fibres were examined in a simulation model. The model was tested by comparing the actual force-velocity curve of a bundle of three fibres, each of which had been individually characterized, with the force-velocity curve predicted by the model for the bundle based on the individual fibre properties. The predicted and measured force-velocity curves were in excellent agreement. The curvature of the force-velocity relation for a muscle, as indicated by a/P0 in Hill's (1938) hyperbolic equation, increases with increasing heterogeneity in the maximum shortening velocities (Vmax(i] of the individual fibres in the muscle. In a muscle that is heterogeneous with respect to Vmax(i), the maximum shortening velocity determined by the slack test method (V0) can be expected to represent the fastest fibre(s) in the muscle. The maximum velocity of shortening (Vm), determined by extrapolation from a hyperbola that is fitted to force-velocity data at finite loads, is substantially lower than V0. The difference in estimates of V0 and Vm is a function of: (i) the degree of heterogeneity of the muscle with respect to Vmax(i) and the curvature of the force-velocity relationship of the individual fibres, and (ii) the force range used to establish the hyperbola from which Vm is derived. The ratio of Vm to V0 can be used as an index to estimate the degree of variability in the maximum velocity of shortening among individual fibres in a muscle.     

The age-related loss of skeletal muscle mass and function: Measurement and physiology of muscle fibre atrophy and muscle fibre loss in humans

Age-related loss of skeletal muscle mass and function, sarcopenia, is associated with physical frailty and increased risk of morbidity (chronic diseases), in addition to all-cause mortality. The loss of muscle mass occurs incipiently from middle-age (∼1%/year), and in severe instances can lead to a loss of ∼50% by the 8–9th decade of life. This review will focus on muscle deterioration with ageing and highlight the two underpinning mechanisms regulating declines in muscle mass and function: muscle fibre atrophy and muscle fibre loss (hypoplasia) – and their measurement. The mechanisms of muscle fibre atrophy in humans relate to imbalances in muscle protein synthesis (MPS) and breakdown (MPB); however, since there is limited evidence for basal alterations in muscle protein turnover, it would appear that “anabolic resistance” to fundamental environmental cues regulating diurnal muscle homeostasis (namely physical activity and nutrition), underlie age-related catabolic perturbations in muscle proteostasis. While the ‘upstream’ drivers of the desensitization of aged muscle to anabolic stimuli are poorly defined, they most likely relate to impaired efficiency of the conversion of nutritional/exercise stimuli into signalling impacting mRNA translation and proteolysis. Additionally, loss of muscle fibres has been shown in cadaveric studies using anatomical fibre counts, and from iEMG studies demonstrating motor unit loss, albeit with few molecular investigations of this in humans. We suggest that defining countermeasures against sarcopenia requires improved understandings of the co-ordinated regulation of muscle fibre atrophy and fibre loss, which are likely to be inextricably linked.     

Metabolic characteristics of fibre types in human skeletal muscle.

Muscle biopsy samples were obtained from healthy subjects in order to evaluate quantitative differences in single fibres of substrate (glycogen and triglyceride) and ion concentrations (Na+ and K+) as well as enzyme activity levels (succinate-dehydrogenase, SDH; phosphofructokinase, PFK; 3-hydroxyacyl-CoA-dehydrogenase, HAD; myosin ATPase) between human skeletal muscle fibre types. After freeze drying of the muscle specimen fragments of single fibres were dissected out and stained for myofibrillar-ATPase with preincubations at pH's of 10.3, 4.6, 4.35. Type I ("red") and II A,B, and C ("white") fibres could then be identified. Glycogen content was the same in different fibres, whereas triglyceride content was highest in Type I fibres (2-3 X Type II). No significant differences were observed for Na+ and K+ between fibre types. The activity for the enzymes studied were quite different in the fibre types (SDH and HAD, Type I is approximately 1.5 X Type II; PFK Type I is approximately 0.5 X Type II, Myosin ATPase Type I is approxiamtely 0.4 X Type II). The subgroups of Type II fibres were distinguished by differences in both SDH and PFK activities (SDH, Type II C is greater than A is greater than B; PFK, Type II B is greater than A is approximately C). It is concluded that contractile and metabolic characteristics of human skeletal fibres are very similar to many other species. One difference, however, appears to be than no Type II fibres have an oxidative potential higher than Type I fibres.     

Contractile properties of skeletal muscle fibre bundles from mice deficient in carbonic anhydrase II

The function of cytosolic carbonic anhydrase (CA) isozyme II is largely unknown in skeletal muscle. Because of this, we compared the in vitro contractile properties of extensor digitorum longus (EDL) and soleus (SOL) fibre bundles from mice deficient in CA II (CAD) to litter mate controls (LM). Twitch rise, 1/2 relaxation time and peak twitch force at 22°C of fibre bundles from CAD EDL [28.4±1.4 ms, 31.2±2.3 ms, 6.2±1.0 Newton/cm² (N/cm²), respectively] and CAD SOL (54.2±7.5 ms, 75.7±13.8 ms, 2.9±0.5 N/cm², respectively) were significantly higher compared to LM EDL (20.5±2.2 ms, 21.9±3.7 ms, 4.5±0.2 N/cm²) and LM SOL (42.8±3.5 ms, 51.4±2.4 ms, 2.1±0.4 N/cm²). However, in acidic Krebs–Henseleit solution, mimicking the pH, PCO₂, and HCO₃ ^– of arterial blood from CAD mice, twitch rise, 1/2 relaxation time, and peak twitch force of fibre bundles from CAD EDL (19.3±0.7 ms, 19.7±2.3 ms, 4.8±0.8 N/cm²) and CAD SOL (41.4±3.6 ms, 51.9±5.5 ms, 2.2±0.7 N/cm²) were not significantly different from LM fibre bundles in normal Krebs–Henseleit solution (EDL: 19.7±1.1 ms, 21.6±0.6 ms, 4.7±0.2 N/cm²; SOL: 42.5±3.1 ms, 51.8±2.6 ms, 1.8±0.3 N/cm²). A higher pH_i during exposure to acidic bathing solution was maintained by CAD EDL (7.37±0.02) and CAD SOL (7.33±0.05) compared to LM EDL (7.28±0.04) and LM SOL (7.22±0.02). This suggests that the skeletal muscle of CAD mice possesses an improved defense of pH_i against elevated pCO₂. In support of this, apparent non-bicarbonate buffer capacity (in mequiv H⁺ (pH unit)^–1 (kg cell H₂O)^–1) as determined by pH microelectrode was markedly increased in CAD EDL (75.7±4.1) and CAD SOL (85.9±3.3) compared to LM EDL (39.3±4.7) and LM SOL (37.5±3.8). Both latter phenomena may be related to the slowed rate of intracellular acidification seen in CAD SOL in comparison with LM SOL upon an increase in PCO₂ of the bath. In conclusion, skeletal muscle from mice deficient in CA II exhibits altered handling of acid–base challenges and shows normal contractile behavior at normal intracellular pH.     

Force and force transients in skeletal muscle fibres of the frog skinned by freeze-drying

A freeze-drying method is described by which single skinned skeletal muscle fibres or fibre bundles can readily be obtained. Skinned fibre segments of the ileofibularis and semitendinous muscles of the frog — activated by means of a rapid increase in the Ca-concentration — showed very stable and reproducible contractions. Complete activation occurred at a Ca-concentration of 1.6·10^–6 M and the mid-point of the pCa-tension curve occurred at 6.3·10^–7 M. Addition of phosphate (10^–2 M) had a depressing effect on the speed of the Ca-activated tension development as well as on the maximum tension reached.Addition of caffeine (10^–2 M) had no effect on the tension generation, indicating that the sarcoplasmic reticulum, if present, was not active. The force responses due to rapid length changes applied to the Ca-activated fibre preparations were found to be qualitatively similar to the force responses on intact tissue. This skinning technique might be employed on human biopsies, enabling the measurement of physiological parameters such as for example force and shortening velocity.     

The intracellular Ca2+ transient and tension in frog skeletal muscle fibres measured with high temporal resolution.

1. The purpose of this study was to determine, with high temporal resolution, the relationship between the intracellular Ca2+ transient (ICT) and the mechanical responses of intact, single skeletal muscle fibres of frogs following stimulation by a single, brief depolarization. 2. The time course of the ICT was monitored using the Ca(2+)-sensitive fluorescent dyes mag-fura-2 (furaptra) and mag-fura-5. The mag-fura dyes have a low affinity for Ca2+ and have been shown to track the ICT with no appreciable kinetic delay. Continuous records of mag-fura fluorescence, tension and stiffness responses were obtained simultaneously at high time resolution at a sarcomere length of 2.9 microns. Experimental temperature was 3 degrees C. 3. When a delay of 0.4 ms due to the low-pass filter associated with the photodetector was included, the onset of the fluorescence response preceded the onset of latency relaxation (the small fall in tension that precedes positive tension generation) by 3.1 +/- 0.2 ms (mean +/- S.E.M., n = 8). After its onset, the mag-fura fluorescence signal continued to change rapidly (indicating increasing intracellular [Ca2+]) to an extreme level that occurred 1.5 +/- 0.5 ms (mean +/- S.E.M., n = 7) before tension had recovered to its resting level following latency relaxation. The time delay from the extreme of the fluorescence signal to the peak of the tension signal was 239 +/- 27 ms (mean +/- S.E.M., n = 6).(ABSTRACT TRUNCATED AT 250 WORDS)     

Optical rotation signals recorded from a single skeletal muscle fibre of a frog

The optical rotation signal of a single frog skeletal muscle fibre was recorded and its properties were examined. To reduce movement of the fibre associated with its excitation, the fibre was immersed in Ringer solution made of 90–95% D2O, stretched, and lightly pressed from above with a cylindrical lens. After these procedures no mechanical movements were recognizable under the dissection microscope, and the optical rotation signal appeared consistently as a brief transient increase in dextrorotation with a duration of about 23 ms. The shape and polarity of the signal did not change among preparations and remained unchanged when the direction of impulse conduction along the fibre was reversed. The influence of the birefringence change associated with excitation was found to be small in most preparations. Ryanodine (10M) reduced the amplitude of the signal to about half that of the control, and increased the duration slightly. Nitrendipine (10μM) reduced the duration of the signal to about half that of the control, and reduced the amplitude to about two-thirds of the control. The effects of the chemicals took place within 10 minutes and remained constant for the periods examined. The intracellular action potentials were not appreciably altered by these chemicals. It is concluded that the optical rotation signal of the single muscle fibre reports the molecular conformational change of proteins during the early stage of the contraction process. This revised version was published online in July 2006 with corrections to the Cover Date.