Quick Length Changes Research Articles

Single molecule imaging expanded to macroscopic level by computer simulation.

Biological machines work skillfully with extremely low energy cost. For example, a human brain consumes only one watt (difference between working and resting states) and a cell does 1p (1×10−12) watts. On the other hand, a super computer “Kei”, which is the best one in Japan, consumes 20 M (2×107) watts. How can the biological machines function so skillfully? We have studied the mechanism of molecular motor, myosin that is responsible for the muscle contraction. For this, we have developed a single molecule nano-detection technique, which allows us to directly observe and manipulate a single myosin molecule of tens of nanometers in size in solution. The movement of myosin motor was expected to be accurate and mechanistic based on an analogy to a man-made machine. However, our result showed that the myosin motor moves not accurately but fluctuating back and forth. The fluctuating movement is due to what the myosin motor does not avoid but instead uses thermal noise. This is in sharp contrast to a manmade machine that uses a lot of energy to avoid thermal noise and work accurately. Can the muscle contract successfully if the myosin motors move fluctuating back and forth? Recently our computer simulation has answered to this question that the fluctuation of myosin motors is essential for adaptive force generation of muscle when quick length or force change is applied (QBiC: http://www.qbic.riken.jp). Furthermore, the UT-heart project of U. Tokyo has shown by a computer simulation by the 10-Peta (1015) Flops computer “Kei” that the fluctuating movement of myosin motors is important for adaptive beats of human heart (http://www.scls.riken.jp/). Recently, we are working with a soccer player, Neymar. The question is how his brain can function to play brilliant soccer. We have measured his brain activities by fMRI and suggested that his brain also uses spontaneous fluctuation to develop his brilliant soccer play (CiNet: http://cinet.jp). Elucidation of the principle of biology using fluctuations that operates commonly from molecule to brain allows us to develop man-made machines that work skillfully with very low energy cost.

Structural Changes of Actin-Bound Myosin Heads after a Quick Length Change in Frog Skeletal Muscle

Changes in the x-ray diffraction pattern from a frog skeletal muscle were recorded after a quick release or stretch, which was completed within one millisecond, at a time resolution of 0.53 ms using the high-flux beamline at the SPring-8 third-generation synchrotron radiation facility. Reversibility of the effects of the length changes was checked by quickly restoring the muscle length. Intensities of seven reflections were measured. A large, instantaneous intensity drop of a layer line at an axial spacing of 1/10.3 nm −1 after a quick release and stretch, and its partial recovery by reversal of the length change, indicate a conformational change of myosin heads that are attached to actin. Intensity changes on the 14.5-nm myosin layer line suggest that the attached heads alter their radial mass distribution upon filament sliding. Intensity changes of the myosin reflections at 1/21.5 and 1/7.2 nm −1 are not readily explained by a simple axial swing of cross-bridges. Intensity changes of the actin-based layer lines at 1/36 and 1/5.9 nm −1 are not explained by it either, suggesting a structural change in actin molecules.

Mechanisms underlying ischemic diastolic dysfunction: relation between rigor, calcium homeostasis, and relaxation rate.

Increased diastolic chamber stiffness (upward arrow DCS) during ischemia may result from increased diastolic calcium, rigor, or reduced velocity of relaxation. We tested these potential mechanisms during severe ischemia in isolated red blood cell-perfused isovolumic rabbit hearts. Ischemia (coronary flow reduced 83%) reduced left ventricular (LV) contractility by 70%, which then remained stable. DCS progressively increased. When LV end-diastolic pressure had increased 5 mmHg, myofilament calcium responsiveness was altered with 50 mmol/l NH(4)Cl or 10 mmol/l butanedione monoxime. These affected contractility (i.e., a calcium-mediated force) but not upward arrow DCS. Second, quick length changes reversed upward arrow DCS, supporting a rigor mechanism. Third, ischemia increased the time constant of isovolumic pressure decline from 47 +/- 3 to 58 +/- 3 ms (P < 0.02) but concomitantly abbreviated the contraction-relaxation cycle, i.e., pressure dissipation occurred earlier without diastolic tetanization. Finally, to assess any link between rate of relaxation and upward arrow DCS, hearts were exposed to 10 mmol/l calcium. Calcium doubled contractility and accelerated relaxation velocity, but without affecting upward arrow DCS. Thus upward arrow DCS developed during ischemia despite severely reduced contractility via a rigor (and not calcium mediated) mechanism. Calcium resequestration capacity was preserved, and reduced relaxation velocity was not linked to upward arrow DCS.

1P069骨格筋急速筋長変化時のX線回折パターンの変化

1P069骨格筋急速筋長変化時のX線回折パターンの変化

Induced potential model of muscular contraction mechanism and myosin molecular structure

The proposed model is characterized by the constant r (Eq. 2-1), the induced potential (Fig. 1), two attached states of a myosin head (Fig. 1), the nonlinear elastic property of the crossbridge (Eq. 2-7), and the expression of U* (Eqs. 3-8 and 3-9), which led us to the following conclusions. 1. The following various magnitudes of myosin head motion are compatible with each other: about 2 nm of the quantity called power stroke by Irving (27), which is the mean moving distance of myosin head in the isometric tension in our model, 4-5 nm of the displacement of a single myosin head during one ATP hydrolysis cycle (Molloy et al. (20)) or a few tens of nm when the actin and myosin filaments are set parallel (Tanaka et al. (21) and Kitamura et al. (42)), and more than 200 nm of the myosin head displacement in a multi-myosin head system below 22 degrees C (Harada et al. (19)). 2. There is one-to-one coupling between the ATP hydrolysis cycle and the attachment-detachment cycle of a myosin head in accordance with the generally accepted concept of chemical reactions, since the head is trapped in the spatially shifting wide potential well (Fig. 1) until epsilon ATP is exhausted. Here, an actin filament interacts with a myosin head like a single molecule. 3. The calculated tension dependence of muscle stiffness agrees well with the observations by Ford et al. (12), as shown in Fig. 9. 4. The calculated shortening velocity V of muscle as a function of P/P0 agreed very well with experimental results as shown in Fig. 13. The deviation from the Hill equation (34) observed by Edman (32) is related with U* being effectively infinite for f1 < kappa b yc0 (Fig. 10). 5. Calculated energy liberation rate W + H as a function of P/P0 has characteristics almost the same as the Hill equation (33), and agrees well with the experimental results as shown in Fig. 14. 6. The time course of tension recovery after a quick length change is determined by four parameters: kappa f, kappa b, a, and Z0. Among them, kappa f, kappa b (Eq. 2-22) and a (Eq. 4-21) are readily determined by analysis of the steady filament sliding and p0. Calculations of T1/T0 and T2/T0 with these three parameters are in very good agreement with experimental data (Fig. 21). Calculated tension variations by assigning the value in Eq. 4-23 to Z0 agree with the observation (Fig. 17). 7. The model suggests that large fluctuations exist in relative positions between the actin and myosin filaments even when the load on a muscle is kept constant (Fig. 23). Taking this fluctuation into account, the time course of the isotonic velocity transient shown in Fig. 22 becomes understandable referring to Fig. 24. 8. The experimental data of the delta yhs vs. delta P/P0 relationship (Fig. 25) is explained. The delta yhs value at delta P/P0 = 0 (about 5 nm) supports the two-attached-state model and thus indicates that the incremental unit step of a myosin head motion along an actin filament is close to L (5.46 nm).

Induced potential model of muscular contraction mechanism and myosin molecular structure

The proposed model is characterized by the constant r (Eq. 2-1), the induced potential (Fig. 1), two attached states of a myosin head (Fig. 1), the nonlinear elastic property of the crossbridge (Eq. 2-7), and the expression of U∗ (Eqs. 3-8 and 3-9), which led us to the following conclusions. 1. The following various magnitudes of myosin head motion are compatible with each other: about 2 nm of the quantity called power stroke by Irving (27), which is the mean moving distance of myosin head in the isometric tension in our model, 4–5 nm of the displacement of a single myosin head during one ATP hydrolysis cycle (Molloy et al. (20)) or a few tens of nm when the actin and myosin filaments are set parallel (Tanaka et al. (21) and Kitamura et al. (42)), and more than 200 nm of the myosin head displacement in a multi-myosin head system below 22 °C (Harada et al. (19)). 2. There is one-to-one coupling between the ATP hydrolysis cycle and the attachment-detachment cycle of a myosin head in accordance with the generally accepted concept of chemical reactions, since the head is trapped in the spatially shifting wide potential well (Fig. 1) until εATP is exhausted. Here, an actin filament interacts with a myosin head like a single molecule. 3. The calculated tension dependence of muscle stiffness agrees well with the observations by Ford et al. (12), as shown in Fig. 9. 4. The calculated shortening velocity V of muscle as a function of PP0 agreed very well with experimental results as shown in Fig. 13. The deviation from the Hill equation (34) observed by Edman (32) is related with U∗ being effectively infinite for fJ < κbyc0 (Fig. 10). 5. Calculated energy liberation rate W + H as a function of PP0 has characteristics almost the same as the Hill equation (33), and agrees well with the experimental results as shown in Fig. 14. 6. The time course of tension recovery after a quick length change is determined by four parameters: κf, κb, a, and Z0. Among them, κf, κb (Eq. 2–22) and a (Eq. 4-21) are readily determined by analysis of the steady filament sliding and p0. Calculations of T1T0 and T2T0 with these three parameters are in very good agreement with experimental data (Fig. 21). Calculated tension variations by assigning the value in Eq. 4-23 to Z0 agree with the observation (Fig. 17). 7. The model suggests that large fluctuations exist in relative positions between the actin and myosin filaments even when the load on a muscle is kept constant (Fig. 23). Taking this fluctuation into account, the time course of the isotonic velocity transient shown in Fig. 22 becomes understandable referring to Fig. 24. 8. The experimental data of the δyhs vs. ΔPP0 relationship (Fig. 25) is explained. The δyhs value at ΔPP0 = 0 (about 5 nm) supports the two-attached-state model and thus indicates that the incremental unit step of a myosin head motion along an actin filament is close to L (5.46 nm).

Modulation of Ca2+ transient decay by tension and Ca2+ removal in hyperthyroid myocardium.

We investigated the contribution of sarcoplasmic reticulum (SR) and Na+/Ca2+ exchanger in the tension-dependent change in the decay of the Ca2+ transients (CaT) in euthyroid (Eu) and hyperthyroid (Hy) myocardium. Hy was induced by thyroxine treatment to enhance the rate of SR Ca2+ uptake. With the use of the aequorin method, CaT and tension in twitch contraction were simultaneously measured under various conditions (changing muscle length and Ca2+ concentration in solution). In both groups, the decay time of CaT (DT) showed a significant dependence on the developed tension, but the tension dependence of DT in Hy was significantly less than in Eu. In the presence of caffeine (3 mM), the tension dependence of DT in Hy became apparent as in Eu. Inhibition of Na+/Ca2+ exchanger by replacing Na+ with Li+ did not affect the dependence in Hy. The normalized extra Ca2+, which is the Ca2+ concentration change in response to a quick length change, in Hy was similar to that in Eu. pCa-tension relations of skinned trabeculae measured at different lengths (1.9 and 2.3 micrometer) were nearly identical in both groups. These results indicate that the tension-dependent change in the affinity of troponin C for Ca2+ works in both Eu and Hy myocardium and that the tension-dependent change in DT is influenced by the Ca2+ uptake rate of SR.

157 Effect of a low osmolality nonionic contrast medium on thrombin generation and fibrinolysis in patients subjected to coronary angiography

The proposed model is characterized by the constant r (Eq. 2-1), the induced potential (Fig. 1), two attached states of a myosin head (Fig. 1), the nonlinear elastic property of the crossbridge (Eq. 2-7), and the expression of U∗ (Eqs. 3-8 and 3-9), which led us to the following conclusions. 1.1. The following various magnitudes of myosin head motion are compatible with each other: about 2 nm of the quantity called power stroke by Irving (27), which is the mean moving distance of myosin head in the isometric tension in our model, 4–5 nm of the displacement of a single myosin head during one ATP hydrolysis cycle (Molloy et al. (20)) or a few tens of nm when the actin and myosin filaments are set parallel (Tanaka et al. (21) and Kitamura et al. (42)), and more than 200 nm of the myosin head displacement in a multi-myosin head system below 22 °C (Harada et al. (19)).2.2. There is one-to-one coupling between the ATP hydrolysis cycle and the attachment-detachment cycle of a myosin head in accordance with the generally accepted concept of chemical reactions, since the head is trapped in the spatially shifting wide potential well (Fig. 1) until εATP is exhausted. Here, an actin filament interacts with a myosin head like a single molecule.3.3. The calculated tension dependence of muscle stiffness agrees well with the observations by Ford et al. (12), as shown in Fig. 9.4.4. The calculated shortening velocity V of muscle as a function of PP0 agreed very well with experimental results as shown in Fig. 13. The deviation from the Hill equation (34) observed by Edman (32) is related with U∗ being effectively infinite for fJ < κbyc0 (Fig. 10).5.5. Calculated energy liberation rate W + H as a function of PP0 has characteristics almost the same as the Hill equation (33), and agrees well with the experimental results as shown in Fig. 14.6.6. The time course of tension recovery after a quick length change is determined by four parameters: κf, κb, a, and Z0. Among them, κf, κb (Eq. 2–22) and a (Eq. 4-21) are readily determined by analysis of the steady filament sliding and p0. Calculations of T1T0 and T2T0 with these three parameters are in very good agreement with experimental data (Fig. 21). Calculated tension variations by assigning the value in Eq. 4-23 to Z0 agree with the observation (Fig. 17).7.7. The model suggests that large fluctuations exist in relative positions between the actin and myosin filaments even when the load on a muscle is kept constant (Fig. 23). Taking this fluctuation into account, the time course of the isotonic velocity transient shown in Fig. 22 becomes understandable referring to Fig. 24.8.8. The experimental data of the δyhs vs. ΔPP0 relationship (Fig. 25) is explained. The δyhs value at ΔPP0 = 0 (about 5 nm) supports the two-attached-state model and thus indicates that the incremental unit step of a myosin head motion along an actin filament is close to L (5.46 nm).

Induced Potential Model for Muscular Contraction Mechanism, Including Two Attached States of Myosin Head

The model for myosin head motion along an actin filament as proposed by Mitsui & Chiba [(1996).J. theor. Biol.182,147–159] is here modified so that it can explain the isometric tension and isotonic velocity transients having the same parameter values as the stationary filament sliding. The modified model differs in that a myosin head forms a complex with two actin molecules in an actin filament and has two attached states in the complex instead of three. Thus an incremental step in the myosin head motion is equal to the F-actin monomer repeat (5.46 nm). Muscle properties concerning the stationary filament sliding are calculated with new parameters in a manner similar to that of Mitsui–Chiba, with the results being qualitatively similar to theirs.In studying the transient phenomena, a quantitative expression is given for the potential energy of the myosin head in the complex, and two rate constants are applied to the kinetics of the head. The time course of tension recovery after a quick length change is determined by calculating the statistical distribution of the head in the two attached states, which conforms to experimental observations by Fordet al. [(1977).J. Physiol.269,441–515]. The tension variationsT1/T0andT2/T/0calculated with parameters determined from the analysis of stationary filament slidings are in fairly good agreement with the experimental data by Fordet al. The model suggest that a large fluctuation exists in the relative position between the actin and myosin filaments even when the load on a muscle is kept constant. Taking this fluctuation into account explains the characteristics of the isotonic velocity transient observed by Civan & Podolsky [(1966).J. Physiol.184,511–534].

Biochemical, Mechanical and Energetic Characterization of Right Ventricular Hypertrophy in the Ferret Heart

Ferret right ventricular hypertrophy is characterized by a decreased and prolonged isometric contraction, associated with altered intracellular calcium (Ca2+) regulation. However myofibrillar composition, cross-bridge function and/or energy transfer may also be involved in these contractile disturbances. Therefore, mechanical properties of myofibrils have been studied with Triton X-100-skinned fibres and troponin (Tn) T and I composition has been examined. Mitochondrial function and functional activity of creatine kinase (CK) isoforms have been studied in saponin-skinned fibres of control (C) and hypertrophied (H) ferret right ventricle, to check for a possible mismatch between energy production and utilization. Our results show that neither TnT nor TnI isoform expression, nor myofibrillar Ca2+responsiveness (similar apparent Ca2+ sensitivity and Hill coefficient) were affected by pressure-overload. Similarly, maximal tension and stiffness, as well as cross bridge cycling rate (v) - assessed by quick length changes - were not significantly altered. Importantly, passive stiffness was dramatically increased (163 ± 30mN/mm2/μm for C v 500 ± 121mN/mm2/μm for H; P < 0.02). Moreover, there was a significant correlation between passive stiffness and cross-bridge cycling rate, indicating that a factor involved in the passive stiffness may affect cross-bridge kinetics. Oxidative capacity (normalized to ventricular dry weight), reflecting mitochondrial ATP production and mitochondrial CK efficacy, as well as myofibrillar CK efficacy (assessed by the shift of MgATP-rigor tension curves before and after phosphocreatine addition), were similar in both groups. These results demonstrate that ferret right ventricular pressure-overload was accompanied by a development of myofibrils and a parallel increase of energy production capacity, transfer and utilization. Decreased compliance, probably linked to an increase in the collagen fraction and/or alterations of the cytoskeletal architecture of the overloaded ventricle, could contribute to the slower time course and decreased amplitude of the isometric twitch.

Effects of acidosis and alkalosis on mechanical properties of hypertrophied rat heart fiber bundles

Effects of alkalosis (pH 7.4) or acidosis (pH 6.8) on the intrinsic mechanical properties of control and pressure-overloaded rat hearts were studied in Triton X-100-treated left ventricular fiber bundles. In control bundles, Ca sensitivity [pCa required for one-half maximal response (pCa50)] was 5.520 +/- 0.012 at pH 7.1. Alkalosis increased it by 0.357 +/- 0.018 pCa unit, whereas acidosis decreased it by 0.365 +/- 0.014 pCa unit with no change in Hill coefficient. Maximal tension was decreased by acidic pH and increased by alkaline pH. Stiffness was measured by the response to quick length changes. Acidosis decreased maximal stiffness but increased the stiffness-to-force ratio, whereas alkalosis increased maximal stiffness but had no effect on stiffness-to-force ratio, suggesting that acidosis decreased the force generated per cross bridge. Alkalosis increased the time constant of tension recovery following a quick stretch from 10.6 +/- 0.66 to 17.45 +/- 1.83 ms, suggesting a decreased cross-bridge cycling rate. Pressure overload induced by thoracic aortic stenosis for 4-6 wk led to a 200% cardiac hypertrophy associated with a shift from fast to slow ventricular myosin. pCa50 of hypertrophied bundles was not different from control (5.541 +/- 0.012). Ca sensitivity was increased by 0.383 +/- 0.008 in alkaline medium and decreased by 0.325 +/- 0.009 in acidic medium. Stiffness-to-force ratio was decreased in acidic pH, and the time constant of tension recovery was increased from 31.0 +/- 0.4 to 34.9 +/- 0.25 ms by alkalosis. In hypertrophied bundles, maximal tension was decreased by acidic pH but not changed by alkalosis. These results show that in the small pH range of our study 1) pH changes have symmetrical effects on Ca sensitivity in both control and hypertrophied bundles, 2) a decrease or an increase in H+ concentration does not have symmetrical effects on the mechanics of the cross bridges, and 3) changes in the phenotype of contractile proteins induced by aortic stenosis do not influence Ca sensitivity, only moderately influence the response to pH changes, and mainly affect the cross-bridge cycling rate.

Mechanical properties of rat cardiac skinned fibers are altered by chronic growth hormone hypersecretion.

Chronic growth hormone (GH) hypersecretion in rats leads to increased isometric force without affecting the unloaded shortening velocity of isolated cardiac papillary muscles, despite a marked isomyosin shift toward V3. To determine if alterations occurred at the level of the contractile proteins in rats bearing a GH-secreting tumor (GH rats), we examined the mechanical properties of skinned fibers to eliminate the early steps of the excitation-contraction coupling mechanism. We found that maximal active tension and stiffness at saturating calcium concentrations (pCa 4.5) were markedly higher in GH rats than in control rats (tension, 52.9 +/- 5.2 versus 38.1 +/- 4.6 mN.mm-2, p < 0.05; stiffness, 1,105 +/- 120 versus 685 +/- 88 mN.mm-2.microns-1, p < 0.01), whereas values at low calcium concentrations (pCa 9) were unchanged. In addition, the calcium sensitivity of the contractile proteins was slightly but significantly higher in GH rats than in control rats (delta pCa 0.04, p < 0.001). The crossbridge cycling rate, reflected by the response to quick length changes, was lower in GH rats than in control rats (62.0 +/- 2.6 versus 77.4 +/- 6.6 sec-1, p < 0.05), in good agreement with a decrease in the proportion of alpha-myosin heavy chains in the corresponding papillary muscles (45.5 +/- 2.0% versus 94.6 +/- 2.4%, p < 0.001). The changes in myosin heavy chain protein phenotype were paralleled by similar changes of the corresponding mRNAs, indicating that the latter occurred mainly at a pretranslational level. These results demonstrate that during chronic GH hypersecretion in rats, alterations at the myofibrillar level contribute to the increase in myocardial contractility observed in intact muscle.

The Potentiating Effect of Prestretch on the Contractile Performance of Rat Gastrocnemius Medialis Muscle During Subsequent Shortening and Isometric Contractions

The aim of the present study was to investigate the effect of an active stretch during the onset of a muscle contraction on subsequent active behaviour of the contractile machinery within an intact mammalian muscle-tendon complex. Muscle length and shortening velocity were studied because they may be important variables affecting this so-called prestretch effect. Seven gastrocnemius medialis (GM) muscles of the rat were examined. Tetanic, isovelocity shortening contractions from 3 mm above muscle optimum length (l0) to l0 - 2 mm, at velocities of 10-50 mm s-1 (dynamic experiments), were preceded by either an isometric contraction (PI) or an active stretch (PS). By imposing quick length decreases between the prephase and the concentric phase, all excess force generated in the prephase was instantaneously eliminated. This procedure only allowed small force changes during subsequent shortening (caused by the intrinsic properties of the contractile machinery). In this way, the influence of series elastic structures on subsequent muscle performance was minimized. Experiments were also performed at lengths ranging from l0 + 2.5 mm to l0 - 1.5 mm, keeping the length constant after the initial quick length changes (isometric experiments). For the dynamic experiments, enhancement of the performance of the contractile machinery (potentiation) was calculated as the ratio of the average force level over each millimetre of shortening during PS to that during PI conditions (PS/PI). For the isometric experiments, the PS/PI force ratio after 300 ms of stimulation was used. The main result of the present study confirmed results reported in the literature and experiments on isolated muscle fibres. For all conditions, a potentiation effect was found, ranging from about 2 to 16%. Muscle length appeared to have a large positive effect on the degree of potentiation. At the greatest lengths potentiation was largest, but at lengths below optimum a small effect was also found. A negative influence of shortening velocity was mainly present at increased muscle lengths (l0 + 2.5 mm and l0 + 1.5 mm). For the dynamic experiments, no interaction was found between the effects of muscle length and shortening velocity on potentiation. However, there was a clear difference between the isometric and dynamic responses: the dependence of potentiation on muscle length was significantly greater for the isometric contractions than for the dynamic ones. These isometric-dynamic differences indicate that the processes underlying prestretch effects operate differently under isometric and dynamic conditions.(ABSTRACT TRUNCATED AT 400 WORDS)

Effects of Volatile Anesthetics on Mechanical Properties of Rat Cardiac Skinned Fibers

Volatile anesthetics were demonstrated to decrease calcium sensitivity and maximal developed force of detergent-treated rat cardiac skinned fibers. To further investigate the possible mechanisms involved in the decrease of force production, stiffness measurements were performed at defined levels of activation with the use of quick length changes of 0.3 to 4% of initial muscle length in the absence and in the presence of 2 MAC of halothane, enflurane, or isoflurane. The results of various series of experiments suggest that these anesthetics have multiple sites of action on cardiac myofibrillar proteins: 1) they decreased active stiffness indicating a decreased number of attached force-generating cross-bridges; 2) they increased the stiffness/force ratio suggesting that the individual force developed by each cross-bridge was decreased during anesthetic exposure; and 3) they increased the time constant of force recovery, which is consistent with the decreased rate of ATP hydrolysis described by others. These changes in cross-bridges kinetics and efficiency may result from conformational changes in all the protein systems involved in force production, and especially actin-myosin attachment and detachment. However, the changes observed were small despite a relatively high concentration of anesthetics; therefore, they will probably participate only to a moderate extent in the overall negative inotropic effect of these agents.

Pressure response to quick volume changes in tetanized isolated ferret hearts.

Observed pressure responses to quick volume changes in the isolated tetanized heart of ferrets were compared with previously reported tension responses to quick length changes in isolated cardiac muscle. Hearts were isolated from ferrets, perfused with ryanodine solution, and stimulated rapidly (50 ms between stimulations) to produce repeated 4-s intervals of tetanus. During each tetanus interval, volume increments of different amplitudes were rapidly removed and then reinfused into the left ventricular chamber. The pressure responses to these volume changes were evaluated for differences between withdrawals and infusions and for dependence on the amplitude of the volume change. It was found for both withdrawal and infusion that the response could be divided into three phases: 1) an immediate phase coincident with volume change, 2) a fast-recovery phase, and 3) a slow-recovery phase. The amplitude of the immediate phase was linearly dependent on the volume change so that a single regression line fit all the data (withdrawal and infusion). The fast recovery phase was 2.5 times faster for infusion than for withdrawal and generated a rebound effect with the pressure going below the initial pressure in the response to infusion. The pressure never went above the initial pressure in the response to withdrawal. The slow-recovery phases in infusion and withdrawal did not differ. These responses in the isolated heart bear striking similarities to tension responses to quick length changes in isolated constantly activated cardiac muscle. We concluded that muscle fiber dynamics were being faithfully transformed to left ventricular (LV) chamber dynamics without appreciable distortion because of the many intervening factors between the wall muscle fiber and the LV chamber.(ABSTRACT TRUNCATED AT 250 WORDS)

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.

Reversible MM-creatine kinase binding to cardiac myofibrils.

Skinned rat papillary muscles and purified preparations of rat cardiac myofibrils were used to study the nature of the interaction of creatine kinase with cardiac myofibrils. High activity of creatine kinase (2 IU/mg protein in fibers and 0.9 IU/mg in purified myofibrils) was due mostly to reversibly bound enzyme. This activity could be removed and rebound. The process of creatine kinase rebinding was characterized by apparent Km value of 0.14 mg/ml (approximately equal to 2 X 10(6) M). Rebinding of creatine kinase to cardiac myofibrils restored the phenomenon of functional compartmentation of adenine nucleotides in myofibrillar space and restored the ability of phosphocreatine to decrease the rigor tension in the presence of MgADP. The physiological experiments with quick length changes showed that rebinding of creatine kinase to skinned papillary muscle also restored Ca sensitivity, increased maximal tension development, decreased stiffness, and restored the tension recovery after quick length changes in muscle under condition of inhibition of endogenous creatine kinase by 1-fluoro-2,4-dinitrobenzene. It is concluded that creatine kinase reversibly bound to cardiac myofibrils is involved in the energy supply for cardiac contraction.

Role of creatine kinase in force development in chemically skinned rat cardiac muscle.

The influence of phosphocreatine in the presence or absence of MgATP and MgADP was studied in Triton X-100-treated thin papillary muscles and ventricular strips of the rat heart. The pCa/tension relationships, the pMgATP/tension relationships, and the tension responses to quick length changes were analyzed. The results show three major consequences of the reduction of the phosphocreatine concentration in the presence of millimolar concentrations of the MgATP. (a) The resting tension and the maximal Ca2+-activated tension were increased, and the pCa/tension relationship was shifted toward higher pCa values and its steepness was decreased; these effects were enhanced by the inclusion of MgADP. (b) The time constant of tension recoveries after quick stretches applied during maximal activation was increased, while the extent of these recoveries was decreased. (c) The study of pMgATP/tension relationships in low Ca concentrations showed that the decrease in phosphocreatine induced a shift toward higher MgATP values with no changes in maximal rigor tension or the slope coefficient; these effects were increased by the increase in MgADP and were independent of the preparation diameter. Thus, modifications of the apparent Ca sensitivity and resting and maximal tension when phosphocreatine is decreased seem to be due to an increasing participation of rigor-like or slowly cycling cross-bridges spending more time in the attached state. These results suggest that endogenous creatine kinase is able to ensure maximal efficiency of myosin ATPase by producing a local high MgATP/MgADP ratio.

Tonic force maintenance after decay of active state in brachiopod smooth adductor muscle

1. Contraction of smooth adductor muscles (SmA) in the brachiopod,Laqueus californicus, is followed by very slow relaxation (R1/2=3 min). The period during which the cross-bridges are in the active state extends from the time of activation to a short time after peak tension is reached. The delayed relaxation phase represents tension maintenance in the absence of the active state. 2. The contraction characteristics of these muscles were compared to those during the catch contractions of mollusc adductor muscles. The immediate elasticity of brachiopod SmA, defined as the change in tension in response to quick length changes, is the same during the contraction and relaxation phases. The immediate elasticity of mollusc catch muscles is greater during the tonic relaxation (catch) than the active contraction phase (Pfitzer and Ruegg 1982). 3. When exposed to CO2, the SmA, whether relaxed or during contraction, enter a reversible rigor-like state of stretch resistance. The immediate elastic modulus during these conditions was the same as during normal contractions. 4. The results indicate that the same physical linkages, probably actomyosin cross-bridges, are responsible for all phases of the contraction/relaxation cycle and also during CO2 induced rigor-like stiffness.

The effect of MgATP on forming and breaking actin-myosin linkages in contracted skinned insect flight muscle fibres.

At neutral pH, fully Ca2+ -activated glycerinated dorsal longitudinal fibre bundles from Lethocerus indicus contract under isometric conditions and respond to release by deactivation, i.e. quick release causes a delayed tension fall. At slightly alkaline pH, the release-induced deactivation becomes a transient phenomenon, i.e. a delayed tension fall is followed by a slow tension recovery. This enabled us to study the effect of MgATP concentration on the phases of deactivation and slow recovery. Reduction of the MgATP concentration slows down the tension response to a quick length change and increases the time constants of the delayed deactivation phase and of the slow recovery phase. The rate constants depend on the ATP concentration according to the Michaelis-Menten law yielding apparent dissociation constants (Km) of 2 mM and 0.09 mM and maximal rate constants of 700 s-1 and 20 s-1 for the deactivation phase (crossbridge detachment) and slow recovery phase (crossbridge reattachment) respectively. The rate of MgATP hydrolysis is also hyperbolically related to the MgATP concentration (Km = 0.14 mM, maximal MgATP turnover rate 1.2 s-1. It is concluded that the effect of MgATP on the deactivation phase, in which crossbridges dissociate strain dependent from the actin, is controlled by at least two mechanisms: (1) fast equilibrium transitions within attached crossbridge states which augment MgATP dissociation from crossbridges with discharged elastic elements; and (2) a crossbridge strain-dependent isomerization of the ternary actin-myosin-MgATP complex which determines crossbridge detachment from the actin.