Effect of stretch and release on equatorial X-ray diffraction during a twitch contraction of frog skeletal muscle.

Time-resolved intensity measurements of the x-ray equatorial reflections were made during twitch contractions of frog skeletal muscles, to which stretches or releases were applied at various times. A ramp stretch applied at the onset of a twitch (duration, 15 ms; amplitude, approximately 3% of muscle length) caused a faster and larger development of contractile force than in an isometric twitch. The stretch accelerated the decrease of the 1.0 reflection intensity (I1,0). The magnitude of increase of the 1,1 reflection intensity (I1,1) was reduced by the stretch, but its time course was also accelerated. A release applied at the peak of a twitch or later (duration, 5 ms; amplitude, approximately 1.5%) caused only a partial redevelopment of tension. The release produced clear reciprocal changes of reflections toward their relaxed levels, i.e., the I1,0 increased and the I1,1 decreased. A release applied earlier than the twitch peak had smaller effects on the reflection intensities. The results suggest that a strength applied at the onset of a twitch causes a faster radial movement of the myosin heads toward actin, whereas a release applied at or later than the peak of a twitch accelerates their return to the thick filament backbone. The results are discussed in the context of the regulation of the myosin head attachment by calcium.     

The Effect of Myofilament Compliance on Kinetics of Force Generation by Myosin Motors in Muscle

We use the inhibitor of isometric force of skeletal muscle N-benzyl-p-toluene sulfonamide (BTS) to decrease, in a dose dependent way, the number of myosin motors attached to actin during the steady isometric contraction of single fibers from frog skeletal muscle (4°C, 2.1 μm sarcomere length). In this way we can reduce the strain in the myofilament compliance during the isometric tetanus (T₀) from 3.54 nm in the control solution (T_0,NR) to ∼0.5 nm in 1 μM BTS, where T₀ is reduced to ∼0.15 T_0,NR. The quick force recovery after a step release (1-3 nm per half-sarcomere) becomes faster with the increase of BTS concentration and the decrease of T₀. The simulation of quick force recovery with a multistate model of force generation, that adapts Huxley and Simmons model to account for both the high stiffness of the myosin motor (∼3 pN/nm) and the myofilament compliance, shows that the increase in the rate of quick force recovery by BTS is explained by the reduced strain in the myofilaments, consequent to the decrease in half-sarcomere force. The model estimates that i), for the same half-sarcomere release the state transition kinetics in the myosin motor are five times faster in the absence of filament compliance than in the control; and ii), the rate of force recovery from zero to T₀ is ∼6000/s in the absence of filament compliance.     

The Pattern of Activation in the Sartorius Muscle of the Frog

The development of isometric twitch tension has been compared with the redevelopment of isometric tension in the fully active frog sartorius muscle following release. At 0°C the rate of rise of isometric twitch tension is the same as that for the muscle in the fully active state at the same tension but not until about 40 msec. after the stimulus and then only for a few milliseconds. The rates of rise of tension in the twitch and in the redevelopment of tension in the fully active muscle appear to be nearly the same at low tensions. Substitution of nitrate for chloride in the Ringer's solution bathing the muscle retards the development of tension during the early part of the contraction phase of the twitch and the effect reaches a maximum within 3 minutes after changing the solutions. These observations have been discussed in connection with some possible patterns of activation and the hypothesis has been advanced that the rate of activation of a sarcomere is determined mainly by the rate at which the transverse component of the link between excitation and contraction is propagated inwards from the periphery to the center of the fiber. This hypothesis has been discussed in relation to others concerning the nature of excitation-contraction coupling.     

Mechanics of cardiac muscle, based on Huxley's model: Mathematical simulation of isometric contraction

Hill's three-component model (Maxwell model) is used to represent the mechanical property of cardiac muscle. The parallel and series elastic elements of the fibres are described according to their non-linear exponential function; and Huxley's sliding-filaments model, together with the activating role of calcium, is applied to the contractile element.

With this composite model, the following responses can be simulated mathematically: isometric twitch at various muscle lengths, tension-length relationships; isometric contraction during quick stretch; and the Bowditch Treppe and tension velocity relationships of the contractile element.     

A model of muscle contraction based on the Langevin equation with actomyosin potentials

We propose a muscle contraction model that is essentially a model of the motion of myosin motors as described by a Langevin equation. This model involves one-dimensional numerical calculations wherein the total force is the sum of a viscous force proportional to the myosin head velocity, a white Gaussian noise produced by random forces and other potential forces originating from the actomyosin structure and intra-molecular charges. We calculate the velocity of a single myosin on an actin filament to be 4.9–49 μm/s, depending on the viscosity between the actomyosin molecules. A myosin filament with a hundred myosin heads is used to simulate the contractions of a half-sarcomere within the skeletal muscle. The force response due to a quick release in the isometric contraction is simulated using a process wherein crossbridges are changed forcibly from one state to another. In contrast, the force response to a quick stretch is simulated using purely mechanical characteristics. We simulate the force–velocity relation and energy efficiency in the isotonic contraction and adenosine triphosphate consumption. The simulation results are in good agreement with the experimental results. We show that the Langevin equation for the actomyosin potentials can be modified statistically to become an existing muscle model that uses Maxwell elements.     

Modulation of contractile activation in skeletal muscle by a calcium-insensitive troponin C mutant

Calcium controls the level of muscle activation via interactions with the troponin complex. Replacement of the native, skeletal calcium-binding subunit of troponin, troponin C, with mixtures of functional cardiac and mutant cardiac troponin C insensitive to calcium and permanently inactive provides a novel method to alter the number of myosin cross-bridges capable of binding to the actin filament. Extraction of skeletal troponin C and replacement with functional and mutant cardiac troponin C were used to evaluate the relationship between the extent of thin filament activation (fractional calcium binding), isometric force, and the rate of force generation in muscle fibers independent of the calcium concentration. The experiments showed a direct, linear relationship between force and the number of cross-bridges attaching to the thin filament. Further, above 35% maximal isometric activation, following partial replacement with mixtures of cardiac and mutant troponin C, the rate of force generation was independent of the number of actin sites available for cross-bridge interaction at saturating calcium concentrations. This contrasts with the marked decrease in the rate of force generation when force was reduced by decreasing the calcium concentration. The results are consistent with hypotheses proposing that calcium controls the transition between weakly and strongly bound cross-bridge states.     

Some properties of linear relaxation in unfused tetanus of human muscle.

The linear relaxation (LR) was studied in isometric unfused tetanus (UT) of the human flexor digitorum sublimis muscle. With a decrease of the force level, the shoulder on the relaxation curve, as measured from the last stimulus, shifted to the right. The length of the linear portion itself weakly depended on activation level. When steady force changed from 100 to 40-50% of the maximum, the slope of LR decreased only by 15 +/- 4%. At smaller force levels the slope began to increase. LR can probably also be hidden in the twitch. With increased tetanus duration, LR becomes longer and slower at all force levels. LR was markedly diminished in contraction on the steep part of the exponential relaxation after a smooth tetanus. Its full recovery needed a train of 4-5 pulses (near 1 s) at the new stationary level. The form of the response to the additional pulse given during relaxation remained approximately constant during the most of LR portion and differed markedly before and after it. LR did not have direct relation to fatigue: in the first UT LR was always slower and longer than in subsequent ones.     

Activation dependence of stretch activation in mouse skinned myocardium: implications for ventricular function

Recent evidence suggests that ventricular ejection is partly powered by a delayed development of force, i.e., stretch activation, in regions of the ventricular wall due to stretch resulting from torsional twist of the ventricle around the apex-to-base axis. Given the potential importance of stretch activation in cardiac function, we characterized the stretch activation response and its Ca2+ dependence in murine skinned myocardium at 22 degrees C in solutions of varying Ca2+ concentrations. Stretch activation was induced by suddenly imposing a stretch of 0.5-2.5% of initial length to the isometrically contracting muscle and then holding the muscle at the new length. The force response to stretch was multiphasic: force initially increased in proportion to the amount of stretch, reached a peak, and then declined to a minimum before redeveloping to a new steady level. This last phase of the response is the delayed force characteristic of myocardial stretch activation and is presumably due to increased attachment of cross-bridges as a consequence of stretch. The amplitude and rate of stretch activation varied with Ca2+ concentration and more specifically with the level of isometric force prior to the stretch. Since myocardial force is regulated both by Ca2+ binding to troponin-C and cross-bridge binding to thin filaments, we explored the role of cross-bridge binding in the stretch activation response using NEM-S1, a strong-binding, non-force-generating derivative of myosin subfragment 1. NEM-S1 treatment at submaximal Ca2+-activated isometric forces significantly accelerated the rate of the stretch activation response and reduced its amplitude. These data show that the rate and amplitude of myocardial stretch activation vary with the level of activation and that stretch activation involves cooperative binding of cross-bridges to the thin filament. Such a mechanism would contribute to increased systolic ejection in response to increased delivery of activator Ca2+ during excitation-contraction coupling.     

Temperature dependence of mammalian muscle contractions and ATPase activities

Isolated rat and mouse extensor digitorum longus (EDL) and soleus muscles were studied under isometric and isotonic conditions at temperatures from approximately 8 degrees -38 degrees C. The rate constant for the exponential rise of tension during an isometric tetanus had a Q10 of approximately 2.5 for all muscles (corresponding to an enthalpy of activation, delta H = 66 kJ/mol, if the rate was determined by a single chemical reaction). The half-contraction time, contraction time, and maximum rate of rise for tension in an isometric twitch and the maximum shortening velocity in an isotonic contraction all had a similar temperature dependence (i.e., delta H approximately 66 kJ/mol). The Mg++ ATPase rates of myofibrils prepared from rat EDL and soleus muscles had a steeper temperature dependence (delta H = 130 kJ/mol), but absolute rates at 20 degrees C were lower than the rate of rise of tension. This suggests that the Mg++ ATPase cycle rate is not limiting for force generation. A substantial fraction of cross-bridges may exist in a resting state that converts to the force-producing state at a rate faster than required to complete the cycle and repopulate the resting state. The temperature dependence for the rate constant of the exponential decay of tension during an isometric twitch or short tetanus (and the half-fall time of a twitch) had a break point at approximately 20 degrees C, with apparent enthalpy values of delta H = 117 kJ/mol below 20 degrees C and delta H = 70 kJ/mol above 20 degrees C. The break point and the values of delta H at high and low temperatures agree closely with published values for the delta H of the sarcoplasmic reticulum (SR) Ca++ ATPase. Thus, the temperature dependence for the relaxation rate of a twitch or a short tetanus is consistent with that for the reabsorption rate of Ca++ into the SR.     

Residual force enhancement following eccentric induced muscle damage

During lengthening of an activated skeletal muscle, the force maintained following the stretch is greater than the isometric force at the same muscle length. This is termed residual force enhancement (RFE), but it is unknown how muscle damage following repeated eccentric contractions affects RFE. Using the dorsiflexors, we hypothesised muscle damage will impair the force generating sarcomeric structures leading to a reduction in RFE. Following reference maximal voluntary isometric contractions (MVC) in 8 young men (26.5±2.8y) a stretch was performed at 30°/s over a 30° ankle excursion ending at the same muscle length as the reference MVCs (30° plantar flexion). Surface electromyography (EMG) of the tibialis anterior and soleus muscles was recorded during all tasks. The damage protocol involved 4 sets of 25 isokinetic (30°/s) lengthening contractions. The same measures were collected at baseline and immediately post lengthening contractions, and for up to 10min recovery. Following the lengthening contraction task, there was a 30.3±6.4% decrease in eccentric torque (P<0.05) and 36.2±9.7% decrease in MVC (P<0.05) compared to baseline. Voluntary activation using twitch interpolation and RMS EMG amplitude of the tibialis anterior remained near maximal without increased coactivation for MVC. Contrary to our hypothesis, RFE increased (～100-250%) following muscle damage (P<0.05). It appears stretch provided a mechanical strategy for enhanced muscle function compared to isometric actions succeeding damage. Thus, active force of cross-bridges is decreased because of impaired excitation-contraction coupling but force generated during stretch remains intact because force contribution from stretched sarcomeric structures is less impaired.     

去负荷小鼠比目鱼肌收缩特性和纤维类型转化

目的:探讨去负荷后小鼠比目鱼肌的收缩特性与骨骼肌纤维类型转化之间的关系。方法:采用离体肌肉灌流技术和电刺激方法,在小鼠后肢去负荷28 d引起骨骼肌萎缩后,观察比目鱼肌单收缩、强直收缩能力和肌疲劳指标等收缩特性的改变,同时利用组织免疫荧光染色和实时定量聚合酶链式反应(real-time PCR)等技术检测去负荷后比目鱼肌快慢肌纤维组成和纤维类型转化的变化。结果:去负荷28 d后,小鼠比目鱼肌单收缩力、强直收缩能力和疲劳指数(fatigue index)均有显著性下降,同时伴有快肌纤维亚型的增加和慢肌纤维亚型的减少。结论:去负荷28 d后小鼠比目鱼肌收缩特性的改变和快慢肌纤维类型的转化有关。     

The Three Filament Model of Skeletal Muscle Stability and Force Production

Ever since the 1950s, muscle force regulation has been associated with the cross-bridge interactions between the two contractile filaments, actin and myosin. This gave rise to what is referred to as the "two-filament sarcomere model". This model does not predict eccentric muscle contractions well, produces instability of myosin alignment and force production on the descending limb of the force-length relationship, and cannot account for the vastly decreased ATP requirements of actively stretched muscles. Over the past decade, we and others, identified that a third myofilament, titin, plays an important role in stabilizing the sarcomere and the myosin filament. Here, we demonstrate additionally how titin is an active participant in muscle force regulation by changing its stiffness in an activation/force dependent manner and by binding to actin, thereby adjusting its free spring length. Therefore, we propose that skeletal muscle force regulation is based on a three filament model that includes titin, rather than a two filament model consisting only of actin and myosin filaments.     

Effect of transient stretch on intracellular Ca2+ during triggered propagated contractions in intact trabeculae

Transient stretch of cardiac muscle during a twitch contraction may dissociate Ca2+ from myofilaments into the cytosol at the moment of quick release of the muscle. We studied the effect of stretch and quick release of trabeculae on changes in intracellular Ca2+ ([Ca2+]i) during triggered propagated contractions (TPCs). Trabeculae were dissected from the right ventricle of 9 rat hearts. [Ca2+]i was measured using electrophoretically injected fura-2. Force was measured using a silicon strain gauge and sarcomere length was measured using laser diffraction techniques. Reproducible TPCs (n = 13) were induced by trains of electrical stimuli (378 +/- 19 ms interval) for 7.5 s at [Ca2+]o of 2.0 mM (27.9 +/- 0.2 degrees C). The latency of the TPC force and the underlying increase in [Ca2+]i was calculated from the time (TimeF) between the last stimulus and the peak of TPC force (PeakF), or the time (TimeCa) between the last stimulus and the peak of the increase in [Ca2+]i during the TPCs (PeakCa). As a result of a 10% increase in muscle length for 150-200 ms during the last stimulated twitches, TimeF and TimeCa decreased and PeakF and PeakCa increased significantly (n = 13). In addition, transient stretch sometimes induced a twitch contraction subsequent to the accelerated TPC and its underlying increase in [Ca2+]i. These results suggest that Ca2+ binding and dissociation from the myofilaments by the stretch and quick release of muscle may modulate the TPC force and the underlying increases in [Ca2+]i and play an important role in the induction of arrhythmias.     

Stretch induced tension rise in a molluscan smooth muscle skinned by freeze drying

Summary The anterior byssus retractor muscle (ABRM) ofMytilus edulis was skinned by freeze drying. Tension transients in response to quick length steps were recorded during isometric contraction induced in ATP salt solution containing 2×10^–6 M Ca²⁺. These transients consisted of four phases similar to those described by Huxley (1974) in skeletal muscle. Under certain conditions (stretch amplitude not larger than 0.6% L_O), and in particular in the presence of cyclic AMP, we observed a delayed tension rise following a quick stretch (stretch activation) which appears to be similar to the stretch activation of insect flight muscle (Jewell and Rüegg 1966).     

The effect of muscle fatigue on the isometric contractile characteristics of skeletal muscle in the cat

The rise time of an isometric twitch, the tetanic tension, the twitch tetanus ratio, the frequency-tension relationship, and the height of the MUAP (motor unit action potential) were measured in fast twitch (medial gastrocnemius) and slow twitch (soleus) muscles of the cat immediately before, in the middle, and immediately after fatiguing isometric contractions at tensions of 30, 50 and 80% of each muscle's initial strength (tetanic tension recorded from the unfatigued muscle). Although the twitch-tetanus ratio was always less for the soleus than for the medial gastrocnemius muscles, the twitch-tetanus ratio for any one muscle was constant throughout the duration of fatiguing isometric contractions at any of the tensions examined. In contrast, the twitch tension and tetanic tension of the muscles were both less after the contractions, the largest reduction occurring for both muscles during contractions sustained at the lowest isometric tensions. The time to peak tension of an isometric twitch was prolonged for both muscles following the contractions. This was associated with a corresponding shift in the frequency tension relationship such that at the point of muscular fatigue, the muscles tetanized at lower frequencies of stimulation than did the unfatigued muscle. In contrast, the amplitude of the MUAP showed only a modest reduction throughout the duration of the fatiguing contractions.     

Regulatory mechanism of length-dependent activation in skinned porcine ventricular muscle: role of thin filament cooperative activation in the Frank-Starling relation

Cardiac sarcomeres produce greater active force in response to stretch, forming the basis of the Frank-Starling mechanism of the heart. The purpose of this study was to provide the systematic understanding of length-dependent activation by investigating experimentally and mathematically how the thin filament “on–off” switching mechanism is involved in its regulation. Porcine left ventricular muscles were skinned, and force measurements were performed at short (1.9 µm) and long (2.3 µm) sarcomere lengths. We found that 3 mM MgADP increased Ca²⁺ sensitivity of force and the rate of rise of active force, consistent with the increase in thin filament cooperative activation. MgADP attenuated length-dependent activation with and without thin filament reconstitution with the fast skeletal troponin complex (sTn). Conversely, 20 mM of inorganic phosphate (Pi) decreased Ca²⁺ sensitivity of force and the rate of rise of active force, consistent with the decrease in thin filament cooperative activation. Pi enhanced length-dependent activation with and without sTn reconstitution. Linear regression analysis revealed that the magnitude of length-dependent activation was inversely correlated with the rate of rise of active force. These results were quantitatively simulated by a model that incorporates the Ca²⁺-dependent on–off switching of the thin filament state and interfilament lattice spacing modulation. Our model analysis revealed that the cooperativity of the thin filament on–off switching, but not the Ca²⁺-binding ability, determines the magnitude of the Frank-Starling effect. These findings demonstrate that the Frank-Starling relation is strongly influenced by thin filament cooperative activation.     

Velocity transients and viscoelastic resistance to active shortening in cat papillary muscle.

When isotonic force steps were applied to activated papillary muscles, the velocity was almost never constant. Early rapid shortening associated with the step persisted for 2-7 ms after the step ends. The early rapid shortening is attributed to lightly damped series elastic recoil and velocity transients of the contractile elements. In most steps, the subsequent velocity declines progressively with shortening, and most of the decline in velocity can be accounted for by compression of a viscoelastic element in parallel with the contractile elements. To demonstrate this, the time course of isotonic velocity was compared with a model in which the force-velocity characteristics of the contractile element were assumed to be constant, and the decline in velocity was due to increasing compression of the viscoelastic element. This model predicted the observed results except that the predicted velocities rose progressively above the measured values for steps to light loads applied late in the twitch, and fell below the velocity trace for heavy loads applied early in the twitch. These deviations would occur if rapid shortening caused deactivation late in the twitch, and if activation were rising early in the twitch. A conditioning step applied to the muscle during the rise of force depressed both isometric force and maximum velocity measured at the peak of force; isometric force was more depressed with later conditioning steps than with earlier steps, while maximum velocity was depressed by about the same extent with both early and late steps. This difference between the effects on isometric force and maximum velocity are explained by a combination of deactivation and viscoelastic load.     

A model of force production that explains the lag between crossbridge attachment and force after electrical stimulation of striated muscle fibers.

Whereas the mechanical behavior of fully activated fibers can be explained by assuming that attached force-producing crossbridges exist in at least two configurations, one exerting more force than the other (Huxley A. F., and R. M. Simmons. 1971. Nature [Lond.]. 233:533-538), and the behavior of relaxed fibers can be explained by assuming a single population of weakly binding rapid-equilibrium crossbridges (Schoenberg, M. 1988. Biophys. J. 54:135-148), it has not been possible to explain the transition between rest and activation in these terms. The difficulty in explaining why, after electrical stimulation of resting intact frog skeletal muscle fibers at 1-5 degrees C, force development lags stiffness development by more than 15 ms has led a number of investigators to postulate additional crossbridge states. However, postulation of an additional crossbridge state will not explain the following three observations: (a) Although the lag between force and stiffness is very different after stimulation, during the redevelopment of force after an extended period of high velocity shortening, and during relaxation of a tetanus, nonetheless, the plots of force versus stiffness in each of these cases are approximately the same. (b) When the lag between stiffness and force during the rising phase of a twitch is changed nearly fourfold by changing temperature, again the plot of force versus stiffness remains essentially unchanged. (c) When a muscle fiber is subjected to a small quick length change, the rate constant for the isometric force recovery is faster when the length change is applied during the rising phase of a tenanus than when it is applied on the plateau. We have been able to explain all the above findings using a model for force production that is similar to the 1971 model of Huxley and Simmons, but which makes the additional assumption that the force-producing transition envisioned by them is a cooperative one, with the back rate constant of the force-producing transition decreasing as more crossbridges attach.     

Critical sarcomere extension required to recruit a decaying component of extra force during stretch in tetanic contractions of frog skeletal muscle fibers

29 single frog skeletal muscle fibers were stretched during fused tetanic contractions. The force increase during stretch exhibited a breakpoint at a critical length change (average: 16.6 nm per one-half sarcomere) that was independent of velocity of stretch and of sarcomere length between 1.8 and 2.8 microns. After stretch there was an early decaying force component with a force-extension curve similar to that during stretch, which disappeared over approximately 2 s. This component was removed by a small, quick release, leaving a longer- lasting component. The critical amplitude of release required to produce this result was found by clamping the fiber to a load at which there was zero velocity of shortening. This amplitude increased with time up to the angle in the force record during stretch, was constant for the remainder of the stretch, and decreased with time after the end of stretch; it was consistently less than the critical amplitude of stretch required to reach the breakpoint of force enhancement during stretch but was also independent of sarcomere length. The force drop accompanying the critical release showed a small increase up to an optimum magnitude at 2.4--2.7 microns sarcomere length, with a decrease at longer lengths.     

Diazepam reveals different rate-limiting processes in rat skeletal muscle contraction

The action of the tranquilizer diazepam on rat skeletal muscle showed that relaxation of isometric twitches is controlled by different processes in extensor digitorum longus (fast-twitch) and soleus (slow-twitch) muscles. Diazepam caused an increase in the amplitude of twitches in fibres from both muscles but increased the twitch duration only in soleus. The amplitude of fused tetani were reduced in both muscles and the rate of relaxation after the tetanus slowed by as much as 34% when the amplitude of the tetanus was reduced by only 11%. The slower tetanic relaxation indicated that calcium uptake by the sarcoplasmic reticulum was slower than normal in slow- and fast-twitch fibres. We conclude therefore that calcium uptake by the sarcoplasmic reticulum is rate limiting for twitch relaxation in slow-twitch but not fast-twitch fibres and suggest that calcium binding to parvalbumin controls relaxation in the fast fibres.