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Effects on shortening velocity of rabbit skeletal muscle due to variations in the level of thin-filament activation.

The effect of Ca2+ upon maximum shortening velocity (Vmax) has been investigated in skinned single fibres from rabbit psoas muscles. Vmax was obtained at 15 degrees C by measuring the amounts of time (delta t) required to take up various amounts of slack (delta l) imposed at one end of the fibre. During maximal activation with Ca2+, plots of delta l vs. delta t were well fitted by a single straight line. Calculation of Vmax from the slopes of the fitted lines yielded a Vmax of 4.44 +/- 0.15 (S.E. of mean) muscle lengths per second (m.l./s). However, at lower levels of Ca2+ activation, plots of delta l vs. delta t were biphasic, containing an initial phase of steady high-velocity shortening and a subsequent phase of steady low-velocity shortening. The transition between these two phases occurred following active shortening equivalent to 60-80 nm/half-sarcomere. Vmax during the high-velocity phase was relatively insensitive to Ca2+ concentration between pCas (i.e. -log [Ca2+]) of 4.5 and 6.0; however, Vmax fell to 3.58 +/- 0.40 m.l./s at pCa 6.1 and further to 1.02 +/- 0.30 m.l./s at pCa 6.2. Vmax during the low-velocity phase decreased as Ca2+ was lowered within the entire range of pCas studied to a minimum value of 0.35 +/- 0.09 m.l./s at pCa 6.2. The degree of thin-filament activation at a particular pCa was varied by partial extraction of troponin-C, which resulted in a permanent though reversible inactivation of parts of the thin filaments. Partial extraction of troponin-C altered the plots of delta l vs. delta t obtained at pCa 4.5 to a biphasic form. In addition, Vmax during the high- and low-velocity phases of shortening was reduced at each pCa greater than 4.5. Vmax values obtained in control fibres at low Ca2+ concentrations and extracted fibres were in good agreement when generated isometric tensions were equivalent. This was the case for both the high- and low-velocity phases of shortening. Fibres were also activated in the absence of Ca2+ by partial removal of total troponin complexes. These fibres developed steady tensions less than 30% of maximum and underwent biphasic shortening, indicating that this phenomenon cannot be the result of shortening-induced dissociation of Ca2+ from troponin-C.(ABSTRACT TRUNCATED AT 400 WORDS)

Actin Cytoskeleton↗

C-protein limits shortening velocity of rabbit skeletal muscle fibres at low levels of Ca2+ activation.

1. Effects on maximum shortening velocity (Vmax) due to partial extraction of C-protein were investigated in skinned fibres from rabbit psoas muscles. Up to 80% of endogenous C-protein was extracted, as assessed by sodium dodecyl sulphate-polyacrylamide gel electrophoresis (SDS-PAGE) of fibre segments obtained before and after the extraction protocol. Vmax was obtained at 15 degrees C by measuring the times required to take up various amounts of slack imposed at one end of the fibre. 2. During maximal activation with Ca2+, Vmax in control fibres was 4.26 +/- 0.16 (mean +/- S.E.M., n = 7) muscle lengths per second (ML/s). Following extraction of approximately 40% of endogenous C-protein, Vmax in these same fibres was 4.41 +/- 0.24 ML/s. 3. At sufficiently low levels of submaximal activation, high- and low-velocity phases of unloaded shortening were observed. Partial extraction of C-protein significantly increased Vmax in the low-velocity phase but had no effect on the high-velocity phase. The effect on low-velocity Vmax was fully reversed by re-addition of purified C-protein. 4. At low levels of activation, the amount of shortening to the break-point between the high- and low-velocity phases was not significantly affected by C-protein extraction. Under control conditions the average break-point was 85.6 +/- 3.1 nm/half-sarcomere, while 84.1 +/- 3.1 nm/half-sarcomere was obtained following partial extraction of C-protein. 5. These results are considered in terms of a model in which an internal load slows Vmax at low levels of activation once a given amount of active shortening has occurred. C-protein may contribute to this internal load either by binding to actin and myosin or by influencing mechanical properties of myosin cross-bridges.

Animals↗

Effects of inorganic phosphate analogues on stiffness and unloaded shortening of skinned muscle fibres from rabbit.

1. We examined the effects of aluminofluoride (AlFx) and orthovanadate (Vi), tightly binding analogues of orthophosphate (Pi), on the mechanical properties of glycerinated fibres from rabbit psoas muscle. Maximum Ca(2+)-activated force, stiffness, and unloaded shortening velocity (Vus) were measured under conditions of steady-state inhibition (up to 1 mM of inhibitor) and during the recovery from inhibition. 2. Stiffness was measured using either step or sinusoidal (1 kHz) changes in fibre length. Sarcomere length was monitored continuously by helium-neon laser diffraction during maximum Ca2+ activation. Stiffness was determined from the changes in sarcomere length and the corresponding changes in force. Vus was measured using the slack test method. 3. AlF chi and Vi each reversibly inhibited force, stiffness and Vus. Actively cycling cross-bridges were required for reversal of these inhibitory effects. Recovery from inhibition by AlF chi was 3- to 4-fold slower than that following removal of V1. 4. At various degrees of inhibition, AlF chi and Vi both inhibited steady-state isometric force more than either Vus or stiffness. For both AlF chi and Vi, the relatively greater inhibition of force over stiffness persisted during recovery from steady-state inhibition. We interpret these results to indicate that the cross-bridges with AlF chi or Vi bound are analogous to those which occur early in the cross-bridge cycle.

Aluminum↗

Accuracy of muscle moment arms estimated from MRI-based musculoskeletal models of the lower extremity.

OBJECTIVE: Biomechanical models that compute the lengths and moment arms of soft tissues are broadly applicable to the treatment of movement abnormalities and the planning of orthopaedic surgical procedures. The goals of this study were to: (i) develop methods to construct subject-specific biomechanical models from magnetic resonance (MR) images, (ii) create models of three lower-extremity cadaveric specimens, and (iii) quantify the accuracy of muscle-tendon lengths and moment arms estimated using these models. MATERIALS AND METHODS: Models describing the paths of the medial hamstrings and psoas muscles for a wide range of body positions were developed from MR images in one joint configuration by defining kinematic models of the hip and knee, and by specifying "wrapping surfaces" that simulate interactions between the muscles and underlying structures. Our methods for constructing these models were evaluated by comparing hip and knee flexion moment arms estimated from models of three specimens to the moment arms determined experimentally on the same specimens. Because a muscle's moment arm determines its change in length with joint rotation, these comparisons also tested the accuracy with which the models could estimate muscle-tendon lengths over a range of hip and knee motions. RESULTS: Errors in the moment arms calculated with the models, averaged over functional ranges of hip and knee flexion, were less than 4 mm (within 10% of experimental values). CONCLUSION: The combination of MR imaging and graphics-based musculoskeletal modeling provides an accurate and efficient means of estimating muscle-tendon lengths and moment arms in vivo.

Biomechanical Phenomena↗

S100A1 modulates skeletal muscle contraction by desensitizing calcium activation of isometric tension, stiffness and ATPase.

S100, a subfamily of the EF-hand type calcium sensing proteins, is implicated in many cellular functions including muscle contractility. Two isoforms, S100A1 and S100B, at 2-10 microM significantly inhibit active tension, stiffness and ATPase of skinned single rabbit psoas muscle fibers at sub-maximal (pCa approximately 6.1-5.6), but not at maximal levels of activation (pCa 4.0). S100A1 is a more potent inhibitor than S100B. Hill analysis of the ATPase-pCa and tension-pCa curves indicates that these proteins reduce calcium sensitivity and enhance the cooperativity toward calcium. We propose S100A1, and perhaps S100B, are viable candidates as physiological modulators of muscle contraction.

Adenosine Triphosphatases↗

Psoas abscess: case report and review of the literature.

Psoas muscle abscess is rare and presents a diagnostic challenge requiring a high index of suspicion. We report an unusual case of primary psoas abscess caused by Proteus mirabilis. Primary psoas abscess is most commonly present in children and is usually caused by Staphylococcus aureus. A total of 434 cases of psoas abscess has been reported. The subject is discussed noting the differences between primary and secondary cases, and emphasizing the importance of ultrasound and computerized tomography guided drainage of psoas abscess. A flow chart for the evaluation and management of primary and secondary abscess is presented, taking into consideration the success rate of the various methods of treatment described in the literature.

Aged↗

Toxicological studies with 6-mercaptopurine in neonates.

In previous studies, we found that Sprague-Dawley rats injected with 6-MP monohydrate at 2 mg base/kg sc daily from 2 to 22 days of age had atrophy of thigh and sublumbar muscles when killed at 16 months of age. The first sign of this muscle atrophy was detected grossly (flattened croup with or without paresis) at 12 months of age. In one experiment of the present work, using the same treatment in rats as above, we found that the earliest onset of muscle atrophy observed by light microscopy occurred at 2 months of age. By 4 months the atrophy could be detected grossly. The atrophy did not uniformly involve all muscles of the hindquarters; the thigh (especially the semitendinosus), leg (soleus but not the extensor carpi group), and lumbar vertebral (including the psoas) muscles were involved. Foreleg (biceps), intercostal, and tongue muscles as well as the sciatic nerve and internal organs appeared unaffected. In another experiment, weanling Sprague-Dawley rats given large daily doses of 6-MP from 25 to 45 days of age had normal muscles when killed at 8 months. In a third experiment, Wistar rats injected with 6-MP (2 mg base/kg sc) daily from 2 to 22 days of age and killed at 6 months had muscle atrophy similar to that seen in Sprague-Dawley rats. In the last experiment, mice and hamsters given large daily doses of 6-MP from 2 to 22 days of age had normal muscles when killed at 10 months. It appears from these results that the 6-MP-induced muscle atrophy occurs only after treatment during the neonatal period and that the atrophy may be species-specific.

Animals↗

Myofibrillar interaction of blot immunoaffinity-purified antibodies against native titin as studied by direct immunofluorescence and immunogold staining.

Titin (also called connectin), a major but so far highly elusive myofibrillar component in striated muscle was purified from glycerinated chicken breast muscle in its native state by use of a similar purification procedure as recently introduced for purification of native titin from rabbit psoas muscle. Low-angle rotary shadowing reveals highly convoluted, long and slender strands, sometimes more extended and with nodules, but also an aggregation into filamentous bundles and reticular networks. Antisera were raised against the purified native molecule and monospecific titin antibodies prepared by a rapid nitrocellulose blot immunoaffinity-purification procedure. Titin antibodies bound to the nitrocellulose immobilized native antigen were directly conjugated with fluorescein isothiocyanate. Titin specifity of purified antibodies was checked by immunoblotting. Direct immunofluorescence of glycerinated myofibrils revealed a uniform doublet staining pattern within the sarcomeres by labelling the region of the A-I junctions and some diffuse staining in the region of the myosin filaments. The same myofibrils examined by indirect immunoelectron microscopy revealed the gold particles highly concentrated at the A-I junctions with considerable labelling within the A-bands, except in their centers. Residual I-bands and Z-lines are free of label. In overstretched myofibrils immunogold staining labelled the gap filaments in the space between I- and A-bands. Isolated native thick filaments showed gold labelling of coiled superthin filaments at the ends of the thick filaments (end-filaments) and at their sides, respectively. The colloidal gold technique in combination with an affinity-purified titin antibody raised against the native molecule adds further evidence for the existence and distribution of an endosarcomeric superthin cytoskeletal filament lattice with titin as a major component.

Animals↗

Further studies on 6-mercaptopurine-induced muscle atrophy in rats, mice, and hamsters treated as neonates.

In previous studies, we found that Sprague-Dawley rats injected with 6-mercaptopurine monohydrate (6-MP) at 2 mg base/kg sc daily from 2 to 22 days of age had atrophy of thigh and sublumbar muscles when killed at 16 months of age. The first sign of this muscle atrophy was detected grossly (flattened croup with or without paresis) at 12 months of age. In one experiment of the present work, using the same treatment in rats as above, we found that the earliest onset of muscle atrophy observed by light microscopy occurred at 2 months of age. By 4 months the atrophy could be detected grossly. The atrophy did not uniformly involve all muscles of the hindquarters; the thigh (especially the semitendinosus), leg (soleus but not the extensor carpi group), and lumbar vertebral (including the psoas) muscles were involved. Foreleg (biceps), intercostal, and tongue muscles as well as the sciatic nerve and internal organs appeared unaffected. In another experiment, weanling Sprague-Dawley rats given large daily doses of 6-MP from 25 to 45 days of age had normal muscles when killed at 8 months. In a third experiment, Wistar rats injected with 6-MP (2 mg base/kg sc) daily from 2 to 22 days of age and killed at 6 months had muscle atrophy similar to that seen in Sprague-Dawley rats. In the last experiment, mice and hamsters given large daily doses of 6-MP from 2 to 22 days of age had normal muscles when killed at 10 months. It appears from these results that the 6-MP-induced muscle atrophy occurs only after treatment during the neonatal period and that the atrophy may be species specific.

Animals↗

Electrophoretic differentiaion f myofibrillar proteins in the pig.

1. Starch-gel electrophoretograms of myosin and tropomyosin preparations in 8m-urea, from longissimus dorsi and psoas muscles of the pig, were characterized by laser densitometry. 2. The typical pattern for freshly prepared myosin from both muscles was similar, there being five electrophoretically distinct components. 3. The number of electrophoretically distinct components in both muscles increased after freeze-drying, but the effect of freeze-drying was more marked in longissimus dorsi. 4. Extraction with 8m-urea containing 2% beta-mercaptoethanol decreased the number of major electrophoretically distinct components of the fresh myosin of both muscles to four. 5. Although there was also some simplification of the patterns after freeze-drying the greater susceptibility of the myosin from longissimus dorsi was still evident. 6. The typical pattern for freshly prepared tropomyosin in 8m-urea differed in the two muscles: in each case it was more complex than that of the corresponding myosins. 7. The pattern of tropomyosin from neither longissimus dorsi nor psoas was altered significantly after freeze-drying. 8. Electrophoretograms of pig longissimus dorsi tropomyosin in 8m-urea differed from those of longissimus dorsi tropomyosin from sheep, ox and rabbit. 9. Extraction of the tropomyosins in 8m-urea and 2% beta-mercaptoethanol simplified the electrophoretic pattern to two major components with samples from pig, sheep and ox, and to one major component with samples from rabbit. 10. It was concluded that classification of skeletal muscles as ;red' or ;white' is insufficient to account for the degree of functional specialization which the electrophoretograms suggest.

Animals↗

Nitric oxide impairs Ca2+ activation and slows cross-bridge cycling kinetics in skeletal muscle.

The effects of the nitric oxide (NO) donor spermine NONOate (Sp-NO, 1.0 mM) on cross-bridge recruitment and cross-bridge cycling kinetics were studied in permeabilized rabbit psoas muscle fibers. Fibers were activated at various Ca2+ concentrations (pCa, negative logarithm of Ca2+ concentration), and the pCa at which force was maximal (pCa 4.0) and approximately 50% of maximal (pCa50 5.6) were determined. Fiber stiffness was determined using 1-kHz sinusoidal length perturbations, and the fraction of cross bridges in the force-generating state was estimated by the ratio of stiffness during maximal (pCa 4.0) and submaximal (pCa 5.6) Ca2+ activation to stiffness during rigor (at pCa 4.0). Cross-bridge cycling kinetics were evaluated by measuring the rate constant for force redevelopment after quick release (by 15% of optimal fiber length, L(o)) and restretch of the fiber to L(o). Exposing fibers to Sp-NO for 10 min reduced force and the fraction of cross bridges in the force-generating state at maximal and submaximal (pCa50) Ca2+ activation. However, the effects of Sp-NO were more pronounced during submaximal Ca2+ activation. Sp-NO also reduced the rate constant for force redevelopment but only during submaximal Ca2+ activation. We conclude that Sp-NO reduces Ca2+ sensitivity by decreasing the number of cross bridges in the strongly bound state and also impairs cross-bridge cycling kinetics during submaximal activation.

Algorithms↗

The effect of phosphate and calcium on force generation in glycerinated rabbit skeletal muscle fibers. A steady-state and transient kinetic study.

The effect of varying concentrations of Pi and Ca2+ on isometric force and on the rate of force development in skinned rabbit psoas muscle fibers has been investigated. Steady-state results show that the three parameters that define the force-pCa relation (Po, pK, and n) all vary linearly with log [Pi]. As [Pi] increases, Po and pK decrease while n increases. The kinetics of force generation in isometrically contracting fibers were studied by laser flash photolysis of caged phosphate. The observed rate of the resulting tension transient, kPi, is 23.5 +/- 1.7 s-1 at 10 degrees C, 0.7 mM Pi, and is independent of [Ca2+] over the range pCa 4.5-7.2. By contrast, kTR, the rate of tension redevelopment following a period of isotonic shortening, is sensitive to [Ca2+] and is slower than kPi (kTR = 13.6 +/- 0.2 s-1 at pCa 4.5, 0.7 mM Pi). The results show that [Ca2+] does not directly affect the Pi release or force-generating steps of the cross-bridge cycle and show that the observed rate of force development depends on how the measurement is made. The data can be interpreted in terms of a model in which strong cross-bridges activate the thin filament, this activation being modulated by Ca2+ binding to troponin.

Actomyosin↗

Fluctuations in tension during contraction of single muscle fibers.

We have searched for fluctuations in the steady-state tension developed by stimulated single muscle fibers. Such tension "noise" is expected to be present as a result of the statistical fluctuations in the number and/or state of myosin cross-bridges interacting with thin filament sites at any time. A sensitive electro-optical tension transducer capable of resolving the expected fluctuations in magnitude and frequency was constructed to search for the fluctuations. The noise was analyzed by computing the power spectra and amplitude of stochastic fluctuations in the photomultiplier counting rate, which was made proportional to muscle force. The optical system and electronic instrumentation together with the minicomputer software are described. Tensions were measured in single skinned glycerinated rabbit psoas muscle fibers in rigor and during contraction and relaxation. The results indicate the presence of fluctuations in contracting muscles and a complete absence of tension noise in eith rigor or relaxation. Also, a numerical method was developed to simulate the power spectra and amplitude of fluctuations, given the rate constants for association and dissociation of the cross-bridges and actin. The simulated power spectra and the frequency distributions observed experimentally are similar.

Animals↗

Successful medical management of multifocal psoas abscess following cesarean section: report of a case and review of the literature.

The psoas abscess is a rare complication in obstetric and gynaecology. Two types of psoas abscess are recognized. The primary psoas abscess is generally following haematogenous dissemination of an infectious agent and the source is usually occult. The most frequently isolated pathogen is Staphylococcus aureus. On the other hand, the secondary abscess is the result of local extension of an infectious process near the psoas muscle. We report the case of a patient who develops a bacteremia from an infected cesarean section wound. The complications were thigh and psoas abscesses with left sacroiliitis. Medical management with prolonged antibiotherapy permit clinical, biological and radiological improvement. Although it required a long hospital stay, medical treatment alone was effective. More experience is required to determine which therapeutic option: medical treatment and/or surgery, is the best choice for this type of complication.

Adult↗

[Ultrasonics in the diagnosis of deep hematomas in hemophilia].

The authors emphasize the clinical importance of US in evaluating the extent and the topographic relationships of haematomas. Seven haemorrhagic episodes occurred in this group of patients have been studied using US. For retrobulbar haematomas we have utilized a single water bath transducer operating at 5 and 7.5 MHz, while for soft tissue (muscle) we have utilized linear array transducer operating at 3.5 MHz. The haemorrhagic complications consisted of two deep haematomas involving the ileo-psoas muscle, with different evolution; two relapsing haematomas of the soleus muscle followed by different clinical course; two post-traumatic relapsing retrobulbar haematomas which gave rise to severe complications; two haematomas which occurred simultaneously in the right iliac fossa. Our experience suggests that US can give both accurate diagnostic information and useful clues concerning the solution and therapeutic management of haematomas. This leads to a better prognosis, mostly when the instrumental investigation is carried out early.

Adolescent↗

Orientational dynamics of indane dione spin-labeled myosin heads in relaxed and contracting skeletal muscle fibers.

We have used electron paramagnetic resonance (EPR) spectroscopy to study the orientation and rotational motions of spin-labeled myosin heads during steady-state relaxation and contraction of skinned rabbit psoas muscle fibers. Using an indane-dione spin label, we obtained EPR spectra corresponding specifically to probes attached to Cys 707 (SH1) on the catalytic domain of myosin heads. The probe is rigidly immobilized, so that it reports the global rotation of the myosin head, and the probe's principal axis is aligned almost parallel with the fiber axis in rigor, making it directly sensitive to axial rotation of the head. Numerical simulations of EPR spectra showed that the labeled heads are highly oriented in rigor, but in relaxation they have at least 90 degrees (Gaussian full width) of axial disorder, centered at an angle approximately equal to that in rigor. Spectra obtained in isometric contraction are fit quite well by assuming that 79 +/- 2% of the myosin heads are disordered as in relaxation, whereas the remaining 21 +/- 2% have the same orientation as in rigor. Computer-simulated spectra confirm that there is no significant population (> 5%) of heads having a distinct orientation substantially different (> 10 degrees) from that in rigor, and even the large disordered population of heads has a mean orientation that is similar to that in rigor. Because this spin label reports axial head rotations directly, these results suggest strongly that the catalytic domain of myosin does not undergo a transition between two distinct axial orientations during force generation. Saturation transfer EPR shows that the rotational disorder is dynamic on the microsecond time scale in both relaxation and contraction. These results are consistent with models of contraction involving 1) a transition from a dynamically disordered preforce state to an ordered (rigorlike) force-generating state and/or 2) domain movements within the myosin head that do not change the axial orientation of the SH1-containing catalytic domain relative to actin.

Animals↗

The effect of altered temperature on Ca2(+)-sensitive force in permeabilized myocardium and skeletal muscle. Evidence for force dependence of thin filament activation.

The effect of changes in temperature on the calcium sensitivity of tension development was examined in permeabilized cellular preparations of rat ventricle and rabbit psoas muscle. Maximum force and Ca2+ sensitivity of force development increased with temperature in both muscle types. Cardiac muscle was more sensitive to changes in temperature than skeletal muscle in the range 10-15 degrees C. It was postulated that the level of thin filament activation may be decreased by cooling. To investigate this possibility, troponin C (TnC) was partially extracted from both muscle types, thus decreasing the level of thin filament activation independent of temperature and, at least in skeletal muscle fibers, decreasing cooperative activation of the thin filament as well. TnC extraction from cardiac muscle reduced the calcium sensitivity of tension less than did extraction of TnC from skeletal muscle. In skeletal muscle the midpoint shift of the tension-pCa curve with altered temperature was greater after TnC extraction than in control fibers. Calcium sensitivity of tension development was proportional to the maximum tension generated in cardiac or skeletal muscle under all conditions studied. Based on these results, we conclude that (a) maximum tension-generating capability and calcium sensitivity of tension development are related, perhaps causally, in fast skeletal and cardiac muscles, and (b) thin filament activation is less cooperative in cardiac muscle than in skeletal muscle, which explains the differential sensitivity of the two fiber types to temperature and TnC extraction. Reducing thin filament cooperativity in skeletal muscle by TnC extraction results in a response to temperature similar to that of control cardiac cells. This study provides evidence that force levels in striated muscle influence the calcium binding affinity of TnC.

Actin Cytoskeleton↗

Effect of joule temperature jump on tension and stiffness of skinned rabbit muscle fibers.

The effects of a temperature jump (T-jump) from 5-7 degrees C to 26-33 degrees C were studied on tension and stiffness of glycerol-extracted fibers from rabbit psoas muscle in rigor and during maximal Ca2+ activation. The T-jump was initiated by passing an alternating current pulse (30 kHz, up to 2.5 kV, duration 0.2 ms) through a fiber suspended in air. In rigor the T-jump induces a drop of both tension and stiffness. During maximal activation, the immediate stiffness dropped by (4.4 +/- 1.6) x 10(-3)/1 degree C (mean + SD) in response to the T-jump, and this was followed by a monoexponential stiffness rise by a factor of 1.59 +/- 0.14 with a rate constant ks = 174 +/- 42 s-1 (mean +/- SD, n = 8). The data show that the fiber stiffness, determined by the cross-bridge elasticity, in both rigor and maximal activation is not rubber-like. In the activated fibers the T-jump induced a biexponential tension rise by a factor of 3.45 +/- 0.76 (mean +/- SD, n = 8) with the rate constants 500-1,000 s-1 for the first exponent and 167 +/- 39 s-1 (mean +/- SD, n = 8) for the second exponent. The data are in accordance with the assumption that the first phase of the tension transient after the T-jump is due to a force-generating step in the attached cross-bridges, whereas the second one is related to detachment and reattachment of cross-bridges.

Animals↗