Quick-release Experiments Research Articles

Mechanical properties of resting and active isolated coronary arteries.

Coronary arterial smooth muscle is myogenically active, is acted upon by a variety of modulating agents, and is subjected, in situ, to compression and distension by the myocardium. Its mechanical properties thus play a key role in the regulation of coronary blood flow. To describe these, we applied phase-plane analysis of shortening velocity vs. length and load clamping to strips of isolated coronary arteries made to contract by an increase in extracellular potassium concentration. At any load, length was larger in relaxed than in "resting" preparations. In addition, preloaded shortening was smaller than would be expected from the relation between active isometric force and muscle length. These suggest the occurrence of stretch-activation and shortening-inactivation. To judge from both phase-plane analysis and quick release experiments, shortening velocity depended on load as well as on time. Velocity decreased with increasing duration of contraction. Shortened coronary arteries resisted lengthening induced by loading and could transiently bear loads that considerably exceeded isometric force. This load-bearing capacity increased with increasing shortening. In conclusion, coronary arterial smooth muscle displays the classical relationship between length, force, and velocity. However, the nature of this relationship changes with duration of activity. In addition, it is greatly affected during changes in length or load, such as expected when the arterial wall is exposed to pulsatile blood flow and is surrounded by mechanically active muscle.

Responses of the relaxed and contracted portal vein to imposed stretch and shortening at graded rates.

Portal veins were subjected to stretch and shortening at rates varying from 0.2 to 47% of initial muscle length/s. The total length change was 33% of initial length which was about 6 mm at the resting preload of 1 mN. Relaxed portal veins (Ca‐free medium) gave curvilinear stress‐strain relations which were practically independent of the rate of the length change and they were similar in their ascending and descending branches (non‐linear parallel elasticity). In contractures produced by K+ or hyperosmolar solution the stress‐strain curves showed pronounced and rate dependent hysteresis. The stretch response in K+ contracture showed initially a steep increase in force in relation to rate of lengthening (dP/dL) up to an apparent break‐point at 2–3% elongation, after which the rise in force was quite independent of the rate of stretch. The break‐point is blunted and shifted to greater length change in portal vein due to large series compliance, but in analogy with interpretations from other types of muscle it may reflect the limit of deformation of attached cross bridges. In hypertonicity contracture dP/dL increased in proportion to rate of stretch. Earlier isotonic quick release experiments showed that the force‐velocity relation of portal vein smooth muscle is markedly different in K+ and hyperosmolality contractures, respectively. Similar differences were found when force‐velocity relations were constructed from the present experiments by plotting active force versus rate of imposed length changes. This indicates that the viscous properties of contracted blood vessels, as revealed in stress‐strain hysteresis, are greatly determined by the force‐velocity characteristics of the smooth muscle actomyosin system. There are, however, certain differences between the force‐velocity curves from isotonic quick release experiments and from the present isovelocity study which remain to be explained.

Force-velocity relation in chemically skinned rat portal vein. Effects of Ca2+ and Mg2+.

Rat portal veins were chemically skinned using Triton X-100 and mounted for isometric and quick release experiments at 20 degrees C. The skinned preparations were activated by Ca2+ (EGTA-buffered) in solutions containing 2mM free-Mg2+ and 1 microM calmodulin. Half maximal isometric force was obtained at pCa = 6.2. Maximal force of the skinned preparations, at pCa = 4.5, was 8.2 +/- 0.8 mN/mm2 (n = 6). Force-velocity relations were determined at varied Ca2+-concentrations. Maximal shortening velocity (Vmax) was 0.10 +/- 0.01 lengths/s at pCa = 4.5. At decreasing Ca2+-levels Vmax decreased (at pCa = 6.25, Vmax = 0.051/s). At pCa = 9 an increased level of free-Mg2+ (15 mM) induces a slow and submaximal increase in tension. Force velocity relations of Mg2+-induced contractures were not different from those of Ca2+-contractures of similar magnitude (pCa = 6.3). The results indicate that the degree of activation of the contractile system, as regulated by Ca2+ and Mg2+, influences the kinetic properties of the actomyosin interaction as well as the force development.

Force-velocity characteristics and active tension in relation to content and orientation of smooth muscle cells in aortas from normotensive and spontaneous hypertensive rats.

Segments of abdominal aorta from spontaneously hypertensive (SHR) and normotensive Wistar-Kyoto (WKY) rats (20-25 weeks) were compared with respect to force production and dynamic mechanical properties. The preparations were mounted in vitro for determination of optimal length (lo) for active force, and then maximally stimulated (high-K+ solution, 10 mm Ca2+, 10(-5) m noradrenaline), and fixed for electron microscopy. Muscle cellular volume per mm vessel wall was significantly (p less than 0.01) higher in SHR (0.14 +/- 0.009 mm3, n = 7) compared to WKY (0.11 +/- 0.004 mm3, n = 7). Unchanged cell length and unaltered cross-sectional area (17 micrometers2) of nucleus containing cell profiles in SHR suggest an increased number of cells in the media. No difference was found in maximum force per unit cell area between SHR (271 +/- 31, n = 7) and WKY (305 +/- 49 mN/mm2, n = 7). Cell orientation was almost circular in both groups, showing that force was measured in parallel to the cell long axis. Aortic segments were mounted in an apparatus for quick-release experiments. They were maximally stimulated and force steps were imposed at peak of contractions. The series elastic component, characterized by the initial elastic recoils at 0.75 lo, has similar stiffness values in SHR and WKY. Velocities were measured 100 msec after release. The results were fitted to Hill's equation and maximum shortening velocity (Vmax) computed. No difference in Vmax was found at 0.75 lo (WKY: 0.048 +/- 0.005; SHR: 0.042 +/- 0.006 lo/s, n = 6 for both). At 0.85 lo, the data were corrected for passive tension (40% to total). Vmax at 0.85 lo was 0.071 +/- 0.009 lo/s (n = 5) for WKY, and 0.069 +/- 0.007 lo/s (n = 5) for SHR. Similar Vmax and force per cell cross-sectional area suggest similar characteristics of actomyosin interaction in SHR and WKY aorta.

Electrical and mechanical activities of frog heart during energetic deficiency

Metabolic inhibition of frog heart by cyanide (3 × 10−3m) induced a drop in tension before any significant alteration occurred in the action potential or the slow inward current. The decline in electrical activity appeared later. After 20 min in cyanide solution at a time when both tension andIslow were reduced, the addition of adrenaline (5 × 10−7m) greatly increased the slow inward current whereas tension only exhibited a partial and slow recovery. In both types of experiment the absence of correlation between the amount of inward Ca2+ current and the tension suggested the existence of a regulatory mechanism other than this current. It was not possible to correlate the fall in tension (which was parallel to the drop in creatine phosphate) with the reduction in ATP. ATP only decreased during application of both cyanide (CN−) and iodoacetic acid (IAA). Quick-stretch and quick-release experiments, conducted during the tension which developed in the presence of CN− and IAA, showed that the mechanical activity did not result from Ca2+-activated tension but from rigor tension, implying a marked deficiency in local ATP. A shortage of the energy available for myofibrillar ATPase activity might be one explanation for the drop in peak tension during metabolic inhibition and the rise in resting tension observed after more severe inhibition might be a shortage of the energy-rich phosphates available for myofibrillar ATPase activity.

The force-velocity relation of isolated twitch and slow muscle fibres of Xenopus laevis.

1. A study has been made of the relation between force and speed of shortening, or lengthening, in isolated twitch and slow muscle fibres, dissected from the iliofibularis muscle of Xenopus laevis. Both after-loaded and quick-release contractions were studied. Twitch fibres were stimulated electrically to give tetanic contractions (5-20 degrees C); slow fibres were activated by a rapid change to solutions with high K concentration (30-75 mM; experiments at 21-24 degrees C).2. The velocity of slow fibres was constant during shortening over 10% length change in after-loaded contractions, except at forces exceeding about 0.8 of isometric tension, P(0). In quick-release experiments, shortening velocity was found to depend not only on the relative load, P/P(0), but also on the instant when the release was made. With increasing time after onset of contraction the initial rate of shortening decreased; also, a progressive fall in speed during shortening became more marked.3. The fall in initial shortening speed with time of release from the onset of a contracture was more pronounced at high [K](o) than at low.4. The relation between the relative force, P/P(0), and shortening velocity, V, in after-loaded contractions (75 mM-K) and quick-release contractions (45 mM-K, early releases) in slow fibres could be represented by a hyperbola with the constants a = 0.10P(0), b = 0.11 lengths/sec; extrapolated V(max.) was 1.10 lengths/sec.5. Isometric tension and maximum shortening velocity in slow fibres were very nearly constant between 32 and 75 mM-K. a/P(0), however, was clearly reduced at 32 mM-K, representing a more curved P-V relation.6. Force-velocity data for twitch fibres (quick-release contractions, 20 degrees C) were reasonably well fitted by a hyperbola (a = 0.38P(0), b = 1.97 lengths/sec, V(max.) = 5.20 lengths/sec), but a systematic deviation was observed for forces exceeding 0.6P(0).7. a/P(0) for twitch fibres was found to be independent of temperature in the range 5-20 degrees C. Q(10) for b was 2.24 (10-20 degrees C), and 2.86 (5-10 degrees C).8. V(max.) for twitch fibres was calculated to be 6.34 lengths/sec at 22.5 degrees C, the average temperature in the slow fibre experiments. The maximum shortening velocity in twitch fibres is thus 6 times higher than in slow fibres.9. When loads in the range 1.1-1.4P(0) were quickly applied to an actively contracting slow fibre, lengthening of the fibre occurred in two phases, an initial rapid phase, followed by a phase of extremely slow lengthening. In corresponding experiments on twitch fibres lengthening was rapid at first and then gradually became slower.10. Factors affecting the shape of the force-velocity curve are discussed. Calculations based on A. F. Huxley's (1957) model for muscle contraction indicated that cross-bridge turnover rate is about 15 times lower in slow than in twitch fibres.

Responses of smooth muscle to quick load change studied at high time resolution.

Quick-release and quick-stretch experiments have been performed on preparations of smooth muscle from rat portal vein and rabbit urinary bladder. The low equivalent mass of the isotonic lever (8 mg) implied that inertial oscillations were limited to the first 5-10 msec after the load step. The high time resolution achieved in this way enabled us to separate three components in the length response to a step change in force: (1) an immediate passive elastic recoil, (2) an isotonic velocity transient lasting 50-75 msec and (3) shortening of the contractile element after its full adjustment to the new load. The maximal series elastic recoil was about 10% of the total muscle length in portal vein but only some 3% in urinary bladder. Stiffness of series elasticity increased in proportion to force and was about 3 times higher in bladder than in portal vein at any force level. Force-velocity relations for loads less than Po could be fitted to Hill's equation; Vmax in 4 AC-stimulated portal veins was 0.53 +/- 0.03 muscle lengths/sec and in 8 K+-activated bladder preparations 0.18 +/- 0.01 muscle lengths/sec. Application of loads greater than Po produced rates of lengthening greater than expected from an extrapolation of Hill's hyperbola. The nature of the transient component is discussed in the light of recent studies of force and velocity transients in skeletal muscle.

A critical analysis of myocardial force-velocity relations obtained from damped quick-release experiments

On the basis of two different models of damped release experiments, the passive component of the all-over shortening velocity throughout an abrupt unloading of cardiac muscle is calculated by treating the muscle functionally as a non-linear spring. It can be shown that: According to these results, the criterion, that force-velocity curves of hyperbolic shape are necessarily true with regard toHill's definition, must be considered as notsufficient. In damped quick-release experiments on isolated cat capillary muscles the derived formulae could sufficiently be justified.

Contraction in venous smooth muscle induced by hypertonicity. Calcium dependence and mechanical characteristics.

AbstractA tonic increase in tension was observed in the isolated rat portal vein in response to large increases in extracellular tonicity (1.5–3 times normal). The purpose of this study was to find out whether this tension increase was due to active contraction or passive shrinkage. The metabolic dependence of the response and its mechanical characteristics, as revealed by quick release experiments, lead to the conclusion that active contraction is the predominant mechanism. The maximal force developed in the hypertonicity contracture amounted to 31 per cent of the maximal K+ contracture obtained in isotonic medium. The hypertonicity contracture occurred independent of the level of membrane potential and in the absence of external Ca2+ (below 10‐‐9 M). It is thus elicited through mechanisms which differ in these respects from the electromechanical cbupling operating in the K+ contracture. The speed of shortening against a given afterload was found to be lower in a hypertonicity contracture than in a K' contracture of comparable isometric force. Explanations for observed differences between the two types of contracture are discussed.

A critical analysis of myocardial force-velocity relations obtained from damped quick-release experiments.

On the basis of two different models of damped release experiments, the passive component of the all-over shortening velocity throughout an abrupt unloading of cardiac muscle is calculated by treating the muscle functionally as a non-linear spring. It can be shown that: According to these results, the criterion, that force-velocity curves of hyperbolic shape are necessarily true with regard toHill's definition, must be considered as notsufficient. In damped quick-release experiments on isolated cat capillary muscles the derived formulae could sufficiently be justified.

Mechanical properties of the stimulated papillary muscle in quick-release experiments

The behavior of heart muscle in the stimulated state is a subject of much controversy. A brief summary of the conclusions drawn by some investigators is given to bring the conflicts to focus. In 1970 Fung formulated the general problem incorporating Hill's three-elements model and Huxley's sliding element theory. This formalism is of general validity and is adopted here to study the papillary muscle behavior. Preliminary results obtained using quick-release technique are presented.

Calcium and the Control of Contraction and Relaxation in a Molluscan Catch Muscle

The anterior byssal retractor muscle (ABRM) is a molluscan smooth muscle that when appropriately stimulated can be made to perform two distinct types of contraction: (1) a phasic contraction that relaxes upon cessation of stimulation, and (2) a contraction known as “catch” that continues long after stimulation has ended. The latter type is marked by an energy output much smaller than that during phasic contraction (Nauss and Davies, 1969; Baguet and Gillis, 1968); during catch there are neither signs of active state as measured by quick-release experiments (Jewell, 1959; Johnson and Twarog, 1960) nor electrical activity as measured by intracellular or extracellular experiments (Twarog, 1967a).

Cardiac Muscle Models

Quick-stretch and quick-release experiments were performed on right ventricular cat papillary muscles to test the applicability of the Hill model to cardiac mechanics. Series elastic component (SEC) force-length curves were calculated from stretches and releases carried out at various times during the contractile cycle. At any SEC force, the SEC elastic modulus depended on the time during the contractile cycle at which it was measured. When measured at the same time and at the same SEC force, elastic moduli obtained by releases of less than 1% of muscle length differed from those obtained by corresponding stretches. Larger stretches, in fact, appeared to yield negative elastic moduli. Thus, a unique SEC modulus could not be identified at any level of SEC force. It is concluded that the concept of the SEC as a passive elasticity appears unsatisfactory and, as a consequence, that the quantitative validity of the Hill model for cardiac muscle is questionable. Moreover, since an anatomical counterpart of the SEC has not been identified, the Hill model also appears unsatisfactory from a structural point of view.

Feedback interaction of mechanical and electrical events in the isolated mammalian ventricular myocardium (cat papillary muscle).

Measurements of transmembrane potentials were performed under different contractile conditions on isolated cat papillary muscles. It was found that the duration of the action potential (within limits of about 20%) depends on the mode of contraction. Isotonic shortening tends to prolong, isometric tension development tends to shorten the duration of the action potential. As a result of the action potential alterations negative or positive inotropic mechanical transients are observed during 5–10 subsequent beats. The decrease in action potential duration is roughly proportional to the force development, and the increase of action potential duration is related to the shortening velocity. By applying a controlled stretch the shortening velocity of the contractile element (V CE ) was reduced below its value during purely isometric conditions. A further decrease of the action potential duration was observed. IncreasingV CE by release experiments increased the action potential duration beyond that observed under lightly loaded isotonic contractions. A quick release taking place after repolarization is complete produces a new distinct wave of depolarization (10–15 mV) which can sometimes initiate a new action potential. The quick release experiments fascilitated the estimation of the time delay of the feedback interaction which is less than 10 msecs. The possibility that passive geometrical changes of the plasma membrane is a causitive factor of the described phenomenon was experimentally excluded. Alternative explanations are discussed. It seems likely that a controlling parameter of this excitation contraction feedback system is contained in the force velocity relation of the contractile element influencing the internal Ca++-transients by its mode of contraction.

Activation in a Skeletal Muscle Contraction Model with a Modification for Insect Fibrillar Muscle

A sliding filament model for muscle contraction is extended by including an activation mechanism based on the hypothesis that the binding of calcium by a regulating protein in the myofibrils must occur before the rate constant governing the making of interactions between cross-bridges and thin filament sites can take on nonzero values. The magnitude of the rate constant is proportional to the amount of bound calcium. The model's isometric twitch and rise of force in an isometric tetanus are similar to the curves produced by real muscles. It redevelops force after a quick release in an isometric tetanus faster than the initial rise. Quick release experiments on the model during an isometric twitch show that the "active state" curve produced is different from the postulated calcium binding curve. The force developed by the model can be increased by a small quick stretch delivered soon after activation to values near the maximum generated in an isometric tetanus. Following the quick stretch, the force remains near the tetanic maximum for a long time even though the calcium binding curve rises to a peak and subsequently decays by about 50%. The model satisfies the constraint of shortening with a constant velocity under a constant load. Modifications can be made in the model so that it produces the delayed force changes following step length changes characteristic of insect fibrillar muscle.

Delay of twitch relaxation induced by stress and stress relaxation

The triceps surae of male rats was prepared in situ for isometric and isotonic recording and for stimulation through the cut sciatic nerve. Relaxation in isometric twitches is markedly delayed in muscle showing high resting tension. The time course of the delayed twitch relaxation is similar to that of stress relaxation in quickly stretched resting muscle. Delay of twitch relaxation is exaggerated in experiments with quick release and foreloading. The findings led to the following conclusions: a) the parallel elastic elements, in muscle with resting tension, lose stress during shortening of contractile elements and regain stress during relaxation of contractile elements. b) The exaggerated delay of twitch relaxation, produced in quick release experiments, is due to increased stress and stress relaxation made possible by enhanced release from stress in the preceding contraction phase of the twitch. c) Stress relaxation in parallel elastic elements accounts for the observed delay of twitch relaxation in muscle with resting tension. Submitted on January 30, 1961

On the tropomyosin-paramyosin system in relation to the viscous tone of lamellibranch ‘catch’ muscle

Some lamellibranch muscles are able to maintain tension for prolonged periods with very little energy expenditure, possibly by means of a special ‘catch' mechanism. Whilst actomyosin seems to be associated with active contraction, the 'catch' might well be linked with tropomyosin A ( TM A ) which is more abundant in these muscles than actomyosin ( AM ) and located in special filaments known as paramyosin. There is in fact a correlation of tropo­myosin content and ‘tonic’ properties in three different portions of Pecten maximus adductor muscle, the striated adductor ( AM/TM = 9:1), the translucent portion of the smooth adductor ( AM/TM = 2:1), and the opaque portion of the latter ( AM/TM = 1:2). A typical ‘catch’ can be demonstrated in the glycerinated fibres of the tropomyosin-rich but not in the tropomyosin-poor type by measuring fibre rigidity in presence of adenosine triphosphate ( ATP ) and Mg ions. Quick release and stretch experiments have shown that, unlike active contraction, rigidity in presence of ATP is not dependent on a functional AM system, which can be inactivated with Salyrgan, denatured with ethanol or plasticized with pyrophosphate. Instead, a change in fibre rigidity is always found to be associated with a change in the colloidal state of TM A when the latter is selectively modified in situ , as, for example, by varying the pH between 6⋅5 and 8 or the ratio of the Mg- ATP chelate to 'free' ATP . The latter increases the solubility and the electrophoretic mobility of TM A and it plasticizes artificial tropomyosin threads or the tropomyosin system in situ . Its action is best explained in terms of a decrease in protein-protein interaction caused by an increase in net charge due to binding, an effect which it counteracted by divalent cations which combine with ATP . Extra­-polating these findings to the behaviour of living muscle one might tentatively suggest that the high rigidity of glycerinated tonic adductor muscle fibres in presence of Mg- ATP corresponds to the high muscle ‘viscosity’ of tonically contracted living molluscan muscle and that free ATP acts as a physiological plasticizer, inducing changes in muscle ‘viscosity’ by affecting the electrostatic tropomyosin-tropomyosin interaction either within or between the para­myosin (tropomyosin) filaments.