9156334
Velocity, force, power, and Ca2+ sensitivity of fast and slow monkey skeletal muscle fibers.
In this study, we determined the contractile properties of single chemically skinned fibers prepared from the medial gastrocnemius (MG) and soleus (Sol) muscles of adult male rhesus monkeys and assessed the effects of the spaceflight living facility known as the experiment support primate facility (ESOP). Muscle biopsies were obtained 4 wk before and immediately after an 18-day ESOP sit, and fiber type was determined by immunohistochemical techniques. The MG slow type I fiber was significantly smaller than the MG type II, Sol type I, and Sol type II fibers. The ESOP sit caused a significant reduction in the diameter of type I and type I/II (hybrid) fibers of Sol and MG type II and hybrid fibers but no shift in fiber type distribution. Single-fiber peak force (mN and kN/m2) was similar between fiber types and was not significantly different from values previously reported for other species. The ESOP sit significantly reduced the force (mN) of Sol type I and MG type II fibers. This decline was entirely explained by the atrophy of these fiber types because the force per cross-sectional area (kN/m2) was not altered. Peak power of Sol and MG fast type II fiber was 5 and 8.5 times that of slow type I fiber, respectively. The ESOP sit reduced peak power by 25 and 18% in Sol type I and MG type II fibers, respectively, and, for the former fiber type, shifted the force-pCa relationship to the right, increasing the Ca2+ activation threshold and the free Ca2+ concentration, eliciting half-maximal activation. The ESOP sit had no effect on the maximal shortening velocity (Vo) of any fiber type. Vo of the hybrid fibers was only slightly higher than that of slow type I fibers. This result supports the hypothesis that in hybrid fibers the slow myosin heavy chain would be expected to have a disproportionately greater influence on Vo.
28441013
Physiological Adaptations to Weightlessness: Effects on Exercise and Work Performance
The aspiration to travel beyond the atmosphere is like the desire to study the ocean floor, the interior of the Earth's crust, to invent a submarine, to fly through the air, improve life, treat disease, and explore the heavens.
25964954
Skeletal muscle fiber, nerve, and blood vessel breakdown in space‐flown rats
Histochemical and ultrastructural analyses were performed postflight on hind limb skeletal muscles of rats orbited for 12.5 days aboard the unmanned Cosmos 1887 biosatellite and returned to Earth 2 days before sacrifice. The antigravity adductor longus (AL), soleus, and plantaris muscles atrophied more than the non‐weight‐bearing extensor digitorum longus, and slow muscle fibers were more atrophic than fast fibers. Muscle fiber segmental necrosis occurred selectively in the AL and soleus muscles; primarily, macrophages and neutrophils infiltrated and phagocytosed cellular debris. Granule‐rich mast cells were diminished in flight AL muscles compared with controls, indicating that mast cell secretion contributed to interstitial tissue edema. Increased ubiquitination of disrupted myofibrils implicated ubiquitin in myofilament degradation. Mitochondrial content and succinic dehydrogenase activity were normal, except for subsarcolemmal decreases. Myofibrillar ATPase activity of flight AL muscle fibers shifted toward the fast type. Absence of capillaries and extravasation of red blood cells indicated failed microcirculation. Muscle fiber regeneration from activated satellite cells was detected. About 17% of the flight AL end plates exhibited total or partial denervation. Thus, skeletal muscle weakness associated with spaceflight can result from muscle fiber atrophy and segmental necrosis, partial motor denervation, and disruption of the microcirculation.— Riley, D. A.; Ilyina‐Kakueva, E. I.; Ellis, S.; Bain, J. L. W.; Slocum, G. R.; and Sedlak, F. R. Skeletal muscle fiber, nerve, and blood vessel breakdown in space‐flown rats. FASEB J 4: 84‐91; 1990.
22730936
Influence of spaceflight on rat skeletal muscle.
The size, succinate dehydrogenase (SDH) and alpha-glycerolphosphate dehydrogenase (GPD) activities, and alkaline myofibrillar adenosinetriphosphatase (ATPase) staining properties were determined from quantitative histochemical analyses of single fibers from five hindlimb muscles of six male rats exposed to a 7-day National Aeronautics and Space Administration spaceflight mission (SL-3). These same properties were determined in a group of ground-based control rats housed under simulated environmental conditions. The wet weight of each of the flight muscles was significantly reduced relative to control. However, the loss of mass varied from 36% in the soleus to 15% in the extensor digitorum longus. The cross-sectional areas of fibers in the flight muscles also were reduced, except for the dark ATPase fibers in the medial gastrocnemius. The greatest relative fiber atrophy occurred in the muscles with the highest proportion of light ATPase fibers. An increase in the percentage of dark ATPase fibers also was observed in flight muscles with a predominance of light ATPase fibers. Also, there was an increase in the biochemically determined myofibrillar ATPase activity of tissue sections of the flight soleus. No changes in histochemical or biochemical measures of ATPase activity were observed in the flight extensor digitorum longus. In general, the SDH activity of flight muscles was maintained, whereas GPD activity either was maintained or increased. Based on a metabolic profile of ATPase, SDH, and GPD, there was an increase in the proportion of fast oxidative-glycolytic fibers in some muscles.
25653891
Models of disuse: a comparison of hindlimb suspension and immobilization.
The effects of 1 and 2 wk of hindlimb suspension (HS) on rat skeletal muscle function were determined and the results compared with those obtained previously with hindlimb immobilization (HI). Both models of disuse (HS and HI) primarily affected slow-twitch muscle. Each decreased the isometric twitch duration in the slow-twitch soleus; however, the HS-mediated effect was entirely a result of a shortened contraction time (CT), whereas HI reduced one-half relaxation time (1/2 RT) as well as CT. Soleus muscle mass and peak tetanic tension (Po) declined with disuse. The HS effect on muscle mass and Po was variable, however, for all experiments HS produced atrophy equal to or greater than HI. A major difference existed in the effects of HS and HI on the maximal speed of soleus muscle shortening (Vmax). One and 2 wk of HS produced increases in Vmax to 4.45 +/- 0.34 and 6.83 +/- 0.74 fiber lengths/s, respectively, compared with control velocities of 3.05 +/- 0.08. By contrast over a similar time period, HI had no significant effect on soleus Vmax. The increase in Vmax at 14 days of HS was associated with, and perhaps caused by, the increased expression of a second faster migrating isozyme of myosin. The new native isozyme comigrated with fast myosin, but its light chain subunits contained only LC1s and LC2s. The mechanism responsible for the increase is unknown. One plausible explanation is that the apparent HS-mediated modification in muscle fiber type is dependent on the elimination of loadbearing or isometric contractions, a condition that does not exist during HI.
29373115
Atrophy of the soleus muscle by hindlimb unweighting.
The unweighting model is a unique whole animal model that will permit the future delineation of the mechanism(s) by which gravity maintains contractile mass in postural (slow-twitch) skeletal muscle. Since the origination of the model of rodent hindlimb unweighting almost one decade ago, about half of the 59 refereed articles in which this model was utilized have been published in the Journal of Applied Physiology. Thus the purpose of this review is to provide, for those researchers with an interest in the hindlimb unweighting model, a summation of the data derived from this model to data and hopefully to stimulate research interest in aspects of the model for which data are lacking. The stress response of the animal to hindlimb unweighting is transient, minimal in magnitude, and somewhat variable. After 1 wk of unweighting, the animal exhibits no chronic signs of stress. The atrophy of the soleus muscle, a predominantly slow-twitch muscle, is emphasized because unweighting preferentially affects it compared with other calf muscles, which are mainly fast-twitch muscles. The review considers the following information about the unweighted soleus muscle: electromyogram activity, amount and type of protein lost, capillarization, oxidative capacity, glycolytic enzyme activities, fiber cross section, contractile properties, glucose uptake, sensitivity to insulin, protein synthesis and degradation rates, glucocorticoid receptor numbers, responses of specific mRNAs, and changes in metabolite concentrations.
9778497
Contractile function of single muscle fibers after hindlimb suspension.
The purpose of this investigation was to determine how muscle atrophy produced by the hindlimb suspension (HS) model alters the contractile function of slow- and fast-twitch single muscle fibers. After 2 wk of HS, small bundles of fibers were isolated from the soleus and the deep and superficial regions of the lateral and medial heads of the gastrocnemius, respectively. The bundles were placed in skinning solution and stored at -20 degrees C until studied. Single fibers were isolated and suspended between a motor arm and force transducer, the functional properties were studied, and subsequently the fiber type was established by myosin heavy chain (MHC) analysis on 1-D sodium dodecyl sulfate polyacrylamide gel electrophoresis. After HS, slow-twitch fibers of the soleus showed a significant reduction in fiber diameter (68 +/- 2 vs. 41 +/- 1 micron) and peak tension (1.37 +/- 0.01 vs. 0.99 +/- 0.06 kg/cm2), whereas the maximal shortening speed (Vmax) increased [1.49 +/- 0.11 vs. 1.92 +/- 0.14 fiber lengths (FL)/s]. A histogram showed two populations of fibers: one with Vmax values identical to control slow-twitch fibers and a second with significantly elevated Vmax values. This latter group frequently contained both slow and fast MHC protein isoforms. The pCa-force relation of the soleus slow-twitch fibers was shifted to the right; consequently, the free Ca2+ required for the onset of tension and for 50% of peak tension was significantly higher after HS. Slow-twitch fibers isolated from the gastrocnemius after HS showed a significant reduction in diameter (67 +/- 4 vs. 44 +/- 3 microns) and peak tension (1.2 +/- 0.06 vs. 0.96 +/- 0.07 kg/cm2), but Vmax was unaltered (1.70 +/- 0.13 vs. 1.65 +/- 0.18 FL/s). Fast-twitch fibers from the red gastrocnemius showed a significant reduction in diameter (59 +/- 2 vs. 49 +/- 3 microns) but no change in peak tension or Vmax. Fast-twitch fibers from the white superficial region of the medial head of the gastrocnemius were unaffected by HS. Collectively, these data suggest that the effects of HS on fiber function depend on the fiber type and location. Both slow-twitch type I and fast-twitch type IIa fibers atrophied; however, only slow-twitch fibers showed a decline in peak tension, and the increase in Vmax was restricted to a subpopulation of slow-twitch soleus fibers.
19670313
Force-velocity and power characteristics of rat soleus muscle fibers after hindlimb suspension.
The effects of 1, 2, and 3 wk of hindlimb suspension (HS) on force-velocity and power characteristics of single rat soleus fibers were determined. After 1, 2, or 3 wk of HS, small fiber bundles were isolated, placed in skinning solution, and stored at -20 degrees C until studied. Single fibers were isolated and placed between a motor arm and force transducer, functional properties were studied, and fiber protein content was subsequently analyzed by sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE). Additional fibers were isolated from soleus of control and after 1 and 3 wk of HS, and fiber type distribution and myosin light chain stoichiometry were determined from SDS-PAGE analysis. After 1 wk of HS, percent type I fibers declined from 82 to 74%, whereas hybrid fibers increased from 10 to 18%. Percent fast type II fibers increased from 8% in control and 1 wk of HS to 26% by 3 wk of HS. Most fibers showed an increased unloaded maximal shortening velocity (Vo), but myosin heavy chain remained entirely slow type I. The mechanism for increased Vo is unknown. There was a progressive decrease in fiber diameter (14, 30, and 38%) and peak force (38, 56, and 63%) after 1, 2, and 3 wk of HS, respectively. One week of HS resulted in a shift of the force-velocity curve, and between 2 and 3 wk of HS the curve shifted further such that Vo was higher than control at all relative loads < 45% peak isometric force. Peak absolute power output of soleus fibers progressively decreased through 2 wk of HS but showed no further change at 3 wk.(ABSTRACT TRUNCATED AT 250 WORDS)
21751551
Effect of hindlimb unweighting on single soleus fiber maximal shortening velocity and ATPase activity.
This study characterizes the time course of change in single soleus muscle fiber size and function elicited by hindlimb unweighting (HU) and analyzes the extent to which varying durations of HU altered maximal velocity of shortening (Vo), myofibrillar adenosinetriphosphatase (ATPase), and relative content of slow and fast myosin in individual soleus fibers. After 1, 2, or 3 wk of HU, soleus muscle bundles were prepared and stored in skinning solution at -20 degrees C. Single fibers were isolated and mounted between a motor arm and a transducer, and fiber force, Vo, and ATPase activity were measured. Fiber myosin content was determined by one-dimensional sodium dodecyl sulfate- (SDS) polyacrylamide gel electrophoresis. After 1, 2, and 3 wk of HU, soleus fibers exhibited a progressive reduction in fiber diameter (16, 22, and 42%, respectively) and peak force (42, 48, and 72%, respectively). Peak specific tension was significantly reduced after 1 wk of HU (18%) and showed no further change in 2-3 wk of HU. During 1 and 3 wk of HU, fiber Vo and ATPase showed a significant increase. By 3 wk, Vo had increased from 1.32 +/- 0.04 to 2.94 +/- 0.17 fiber lengths/s and fiber ATPase from 291 +/- 16 to 1,064 +/- 128 microM.min-1 x mm-3. The percent fibers expressing fast myosin heavy chain increased from 4% to 29% by 3 wk of HU, and Vo and ATPase activity within a fiber were highly correlated.(ABSTRACT TRUNCATED AT 250 WORDS)
21304946
Effect of hindlimb unloading on rat soleus fiber force, stiffness, and calcium sensitivity.
The purpose of this study was to examine the time course of change in soleus muscle fiber peak force (N), tension (Po, kN/m2), elastic modulus (Eo), and force-pCa and stiffness-pCa relationships. After 1, 2, or 3 wk of hindlimb unloading (HU), single fibers were isolated and placed between a motor arm and a transducer, and fiber diameter, peak absolute force, Po, Eo, and force-pCa and stiffness-pCa relationships were characterized. One week of HU resulted in a significant reduction in fiber diameter (68 +/- 2 vs 57 +/- 1 microns), force (3.59 +/- 0.15 vs. 2.19 +/- 0.12 x 10(-4) N), Po (102 +/- 4 vs. 85 +/- 2 kN/m2), and Eo (1.96 +/- 0.12 vs. 1.37 +/- 0.13 x 10(7) N/m2), and 2 wk of HU caused a further decline in fiber diameter (45 +/- 1 microns), force (1.31 +/- 0.06 x 10(-4) N), and Eo (0.96 +/- 0.09 x 10(7) N/m2). Although the mean fiber diameter and absolute force continued to decline through 3 wk of HU, Po recovered to values not significantly different from control. The Po/Eo ratio was significantly increased after 1 (5.5 +/- 0.3 to 7.1 +/- 0.6), 2, and 3 wk of HU, and the 2-wk (9.5 +/- 0.4) and 3-wk (9.4 +/- 0.8) values were significantly greater than the 1-wk values. The force-pCa and stiffness-pCa curves were shifted rightward after 1, 2, and 3 wk of HU. At 1 wk of HU, the Ca2+ sensitivity of isometric force, assessed by Ca2+ concentration required for half-maximal force, was increased from the control value of 1.83 +/- 0.12 to 2.30 +/- 0.10 microM. In conclusion, after HU, the decrease in soleus fiber Po can be explained by a reduction in the number of myofibrillar cross bridges per cross-sectional area. Our working hypothesis is that the loss of contractile protein reduces the number of cross bridges per cross-sectional area and increases the filament lattice spacing. The increased spacing reduces cross-bridge force and stiffness, but Po/Eo increases because of a quantitatively greater effect on stiffness.
42058730
Time course of soleus muscle myosin expression during hindlimb suspension and recovery.
This study examined the time course of adult rodent soleus muscle myofibril and myosin isoform protein expression after 4, 8, 16, 28, and 56 days of hindlimb unweighting by tail suspension (S). The time course of soleus muscle recovery (R) was also examined after 28 days of hindlimb unweighting with an additional 4, 8, 16, and 28 days of unrestricted cage activity. During suspension, soleus muscle myofibril protein rapidly decreased from 34.3 +/- 3.1 (1.96SE) mg/pair in the control (C) group to 6.9 +/- 1.4 (1.96SE) mg/pair in S (t = 56 days). The calculated first-order degradation rate constant for this loss was kd = 0.17 days-1 [half time (t1/2) = 4.1 days]. The estimated slow myosin (SM) isoform content decreased from 13.4 +/- 2.0 (1.96SE) mg/pair in C to 2.1 +/- 0.2 (1.96SE) mg/pair in S (kd = 0.19 days-1, t1/2 = 3.6 days). The relative proportion of other myosin isoforms was increased at 28 and 56 days of suspension, reflecting an apparent de novo synthesis and the loss of SM. Recovery of contractile protein after 28 days of suspension was slower for both the myofibril protein and the SM isoform (kd = 0.07 days-1, t1/2 = 10 days). These data suggest that loss of weight bearing specifically affected the mechanisms of contractile protein expression reflected in soleus muscle protein degradation processes. In addition, the expression of the myosin isoforms were apparently differentially affected by the loss of weight-bearing activity.
24931035
Spaceflight and growth effects on muscle fibers in the rhesus monkey.
Spaceflight causes considerable atrophy in hindlimb muscles of the rat. The purpose of this study was to investigate the effect of a 14-day spaceflight (COSMOS 2044) on selected morphological and metabolic properties of single muscle fibers in a nonhuman primate, Macaca mulatta. Biopsies were taken from the soleus (Sol), medial gastrocnemius (MG), and tibialis anterior (TA) muscles of two rhesus monkeys 107 days before flight and 24 h after return from flight. Muscle biopsies were taken from two independent sites in each muscle by use of a small (3-mm OD) Bergstrom biopsy needle. The biopsies weighed 8-14 mg and contained 100-200 fibers, of which an average of 40 fibers were acceptable for metabolic and size analyses. The 14-day spaceflight had little effect on fiber size in the Sol and MG muscles, whereas there appeared to be a slight decrease in size in the TA. In each of the flight animals, the mean fiber size in the postflight biopsies increased relative to preflight values. An increase in fiber size over the same period of time was also observed in four control monkeys that were the same age and approximately the same weight as the flight monkeys. The relative increase in size was related to the body weight of the monkey at the time of the pre- and postflight biopsies. The mean fiber succinate dehydrogenase activity appeared to decrease in the MG, whereas there was no apparent effect of spaceflight on the Sol and TA muscles.(ABSTRACT TRUNCATED AT 250 WORDS)
2293869
Testing primates with joystick-based automated apparatus: Lessons from the Language Research Center’s Computerized Test System
Nonhuman primates provide useful models for studying a variety of medical, biological, and behavioral topics. Four years of joystick-based automated testing of monkeys using the Language Research Center’s Computerized Test System (LRC-CTS) are examined to derive hints and principle for comparable testing with other species-including humans. The results of multiple parametric studies are reviewed, and reliability data are presented to reveal the surprises and pitfalls associated with video-task testing of performance.
7129796
Architectural and fiber type distribution properties of selected rhesus leg muscles: feasibility of multiple independent biopsies.
In experiments involving primates, e.g. before and after spaceflight, needle biopsies were thought to be a logical and feasible means of obtaining metabolic and morphological information from skeletal muscles. However, the feasibility of obtaining consistent, repeatable biopsies from individual muscles had to be demonstrated prior to the acceptance of this procedure. To study this approach, the architectural properties and the fiber type distributions at three levels and two regions along the proximo-distal axis of the soleus, medial gastrocnemius and tibialis anterior of adult rhesus monkeys were determined. In each muscle, biopsies were taken from specific regions where the fiber type distribution was determined. Within each region of each muscle, the fiber type populations were similar at the three levels studied. The percentage of fast or oxidative fibers in the biopsies and in the regions of the same muscle were highly correlated, i.e. r = 0.98 for both comparisons. In addition, based on normalized values (z scores), 25/26 and 22/26 biopsies were within the 95% confidence interval, i.e. the biopsies were a representative sample of the mean fiber type population of that region of the muscle. In all muscles, the mean fiber lengths were no more than one third the length of the muscle. Together, these data indicate the feasibility of obtaining independent, repeated biopsies having similar fiber types from each of the muscles studied.
24967832
Soleus fiber force and maximal shortening velocity after non-weight bearing with intermittent activity.
This study examined the effectiveness of intermittent weight bearing (IWB) as a countermeasure to non-weight-bearing (NWB)-induced alterations in soleus type I fiber force (in mN), tension (Po; force per fiber cross-sectional area in kN/m-2), and maximal unloaded shortening velocity (Vo, in fiber lengths/s). Adult rats were assigned to one of the following groups: normal weight bearing (WB), 14 days of hindlimb NWB (NWB group), and 14 days of hindlimb NWB with IWB treatments (IWB group). The IWB treatment consisted of four 10-min periods of standing WB each day. Single, chemically permeabilized soleus fiber segments were mounted between a force transducer and position motor and were studied at maximal Ca2+ activation, after which type I fiber myosin heavy-chain composition was confirmed by sodium dodecyl sufate-polyacrylamide gel electrophoresis. NWB resulted in a loss in relative soleus mass (-45%), with type I fibers displaying reductions in diameter (-28%) and peak isometric force (-55%) and an increase in Vo (+33%). In addition, NWB induced a 16% reduction in type I fiber Po, a 41% reduction in type I fiber peak elastic modulus [Eo, defined as (delta force/delta length) x (fiber length/fiber cross-sectional area] and a significant increase in the Po/Eo ratio. In contrast to NWB, IWB reduced the loss of relative soleus mass (by 22%) and attenuated alterations in type I fiber diameter (by 36%), peak force (by 29%), and Vo (by 48%) but had no significant effect on Po, Eo, or Po/Eo. These results indicate that a modest restoration of WB activity during 14 days of NWB is sufficient to attenuate type I fiber atrophy and to partially restore type I peak isometric force and Vo to WB levels. However, the NWB-induced reductions in Po and Eo, which we hypothesize to be due to a decline in the number and stiffness of cross bridges, respectively, are considerably less responsive to this countermeasure treatment.
24069773
Effects of calcium and ionic strength on shortening velocity and tension development in frog skinned muscle fibres.
1. The influence of Ca2+ concentration and ionic strength on the maximum velocity of shortening (Vmax) and the tension generating capability of frog skinned muscle fibres has been studied at temperatures between 1 and 10 degrees C. 2. Fibre segments were mounted between a force transducer and servo motor, where they could be viewed and photographed through a microscope. Segments in which the striations became non‐uniform during activation were discarded. 3. Velocity was obtained as a function of load by stepping the tension to values less than the steady isometric tension. Vmax was then determined by an extrapolation technique. Vmax was also obtained using a second, independent method by measuring the times required to take up various amounts of slack imposed on the segments. 4. Vmax was significantly influenced by the Ca2+ concentration, decreasing by about one half when the Ca2+ concentration was reduced to give steady tensions less than half‐maximal. 5. Vmax was not influenced by changes in ionic strength, in the range 0.09‐0.18 M. Steady tension was found to increase as ionic strength was decreased in the same range. 6. These results indicate that the effect of changes in ionic strength is to alter the numbers or stiffness of attached cross‐bridges, while there is no apparent influence of ionic strength on the steady‐state kinetics of the actin‐myosin interaction during unloaded shortening. The mechanism responsible for the influence of Ca2+ on Vmax is unknown, though possible sites of action for Ca2+ are discussed.
9381888
Shortening velocity in single fibers from adult rabbit soleus muscles is correlated with myosin heavy chain composition.
Extensive variations exist in the heavy and light chain components of myosin in vertebrate striated muscles. In the present study, we have characterized a specific contractile property, velocity of shortening, and protein subunit composition of single fibers from adult rabbit soleus muscles. Maximum velocity of shortening (Vmax) was measured using the slack test method, and the myosin composition of these same fibers was determined using an ultrasensitive sodium dodecyl sulfate-polyacrylamide gel electrophoresis system. While most fibers were found to have velocities between 0.5 and 1.0 muscle length/s, several had velocities distributed between 1.33 and 2.99 muscle length/s. The fibers in the slower group had myosin subunits that were solely of the slow type; however, those in the faster group contained both fast and slow heavy chains and light chains. The velocity of shortening measured in fibers having both myosin types was highly correlated with the myosin heavy chain composition, with velocity increasing as the proportion of fast-type heavy chain increased. Variations in light chain composition, particularly fast and slow myosin light chain 1, appeared to occur independently of the variations in heavy chain composition, suggesting that some myosin molecules consist of mixtures of slow- and fast-type subunits.
41398237
Improved methodology for analysis and quantitation of proteins on one-dimensional silver-stained slab gels.
A sodium dodecyl sulfate-discontinuous polyacrylamide gel electrophoresis system for separation and quantitation of low-molecular-weight (75 to 10K Da) proteins from single muscle fibers is described. Slab gels (0.75 mm thick) were stained using an improved silver-stain technique which does not require photographic fixers in order to achieve low-level background staining. The modified staining procedure uses continuous flow washing to minimize the handling of gels. The procedure has high sensitivity and gave a linear response between approximately 2 and 70 ng of protein per band. In addition, a convenient method for mounting slab gels for photography, scanning, and long-term storage has been developed.
42722093
Muscle Mechanics: Adaptations with Exercise‐Training
Based on the MHC isoform pattern, adult mammalian limb skeletal muscles contain two and, in some species, three types of fast fibers (Type IIa, IIx, and IIb), and one slow fiber (Type I). Slow muscles, such as the soleus, contain primarily the slow Type I fiber, whereas fast-twitch muscles are composed primarily of a mixture of the fast myosin isozymes. Force generation involves cross-bridge interaction and transition from a weakly bound, low-force state (AM-ADP-P(i)) to the strongly bound, high-force state (AM-ADP). This transition is thought to be rate limiting in terms of dP/dt, and the high-force state is the dominant cross-bridge form during a peak isometric contraction. Intact fast and slow skeletal muscles generate approximately the same amount of peak force (Po) of between 200 and 250 kN.m-2. However, the rate of transition from the low- to high-force state shows Ca2+ sensitivity and is 7-fold higher in fast-twitch, as compared to slow-twitch, skeletal muscle fibers. Fiber Vo or the maximal cross-bridge cycle rate is highly correlated with and thought to be dependent on the specific activity of the myosin or myofibrillar ATPase. The hierarchy for Vo is the Type IIb > IIx > IIa > I. This functional difference for the fast fiber types explains the higher Vo observed in the predominantly Type IIb SVL vs. the mixed fast Type IIa and IIb EDL muscle. A plot of Vo vs. species size demonstrates that an inverse relationship exists between Vo and body mass. From the standpoint of work capacity, the important property is power output. An analysis of individual muscles indicates that peak power is obtained at loads considerably below 50% of Po. Individuals with a high percentage of fast-twitch fibers generate a greater torque and higher power at a given velocity than those with predominantly slow-twitch fibers. In humans, mean peak power occurred in a ratio of 10:5:1 for the Type IIb, IIa, and I fibers. The in vivo measurement of the torque-velocity relationship and Vmax in human muscle is difficult because of limitations inherent in the equipment used and the inability to study the large limb muscles independently. Nevertheless, the in vivo torque-velocity relationships are similar to those measured in vitro in animals. This observation suggests that little central nervous system inhibition exists and that healthy subjects are able to achieve maximal activation of their muscles. Although peak isometric tension is not dependent on fiber type distribution, a positive correlation exists between the percentage of fast fibers and peak torque output at moderate-to-high angular isokinetic velocities. Consequently, peak power output is substantially greater in subjects possessing a predominance of fast fibers. The mechanical properties of slow and fast muscles do adapt to programs of regular exercise. Endurance exercise training has been shown to increase the Vo of the slow soleus by 20%. This increase could have been caused by either a small increase in all, or most, of the fibers, or to a conversion of a few fibers from slow to fast. Recently, the increase was shown to be caused by the former, as the individual slow Type I fibers of the soleus showed a 20% increase in Vo, but there was little or no change in the percentage of fast fibers. The increased Vo was correlated with, and likely caused by, an increased fiber ATPase. We hypothesize that the increased ATPase and cross-bridge cycling speed might be attributable to an increased expression of fast MLCs in the slow Type I fibers (Fig. 14.10). This hypothesis is based on the fact that light chains have been shown to be involved in the power stroke, and removal of light chains depresses force and velocity. Regular endurance exercise training had no effect on fiber size, but with prolonged durations of daily training it depressed Po and peak power. When the training is maintained over prolonged periods, it may even induce atrophy of the slow Type I and fast Type IIa fibers. (ABSTRACT TRUNCATED)
10464596
Contractile, biochemical, and histochemical properties of thyrotoxic rat soleus muscle.
The effects of thyrotoxicosis on the contractile properties of soleus muscle were examined in rats given 3 mg of T4 and 1 mg of T3 per kg of diet for 6--8 wk. Thyrotoxicosis induced significant decreases in isometric twitch contraction time (CT), one-half relaxation time, and peak twitch tension. The Ca2+ uptake activity of the sarcoplasmic reticulum (SR) was increased in the thyrotoxic muscles; this adaptation in the SR provides a possible mechanism for the alterations in isometric contractile properties. Thyrotoxicosis induced a large increase in fibers classified as type 2, on the basis of an alkali-stable histochemical reaction for ATPase, in the soleus. Although this reaction is commonly interpreted as indicating that a muscle is fast, maximum shortening velocity (Vmax) and Mg2+ activated actomyosin ATPase activity were unaffected in the thyrotoxic soleus. Our findings provide evidence that CT and Vmax can vary independently and that the histochemical ATPase reaction may not always reflect the biochemical properties that make myosin fast or slow.
8803371
Structure-function relationship of soleus muscle fibres from the rhesus monkey
Functional and structural properties of rhesus monkey skinned fibres were studied in order to examine the relationship between calcium/strontium (Ca/Sr) activation characteristics and protein composition. The fibres were classified according to their Ca/Sr affinity into slow (61%) and fast groups (39%). According to the myosin isoform composition, two additional hybrid types were defined. Thus, four profiles were characterized: two corresponding to slow (S) and fast (F) isoforms and two corresponding to a mixed proportion of slow and fast isoforms. They were called hybrid slow (HS) or hybrid fast (HF) based on the predominant myosin isoform. Tension/pCa parameters and maximal shortening velocities were determined. S fibres showed a higher pCa threshold and affinity as well as shallower slopes of their tension/pCa curve than did F fibres. HS and HF fibres exhibited tension/pCa curves which were positioned close to those of S and F fibres, respectively. No significant difference was observed between S and HS fibres or between F and HF fibres. Maximal shortening velocity values were higher for fibres expressing predominantly fast myosin isoforms. We suggest than when both S and F isoforms of myofibrillar proteins are expressed in a muscle fibre, the functional properties are mainly governed by the predominant isoform.
23158799
Fiber‐type composition of selected hindlimb muscles of a primate (cynomolgus monkey)
The distribution of fiber types in selected leg and thigh muscles of three male Cynomolgus monkeys were determined. Almost all fibers could be classified as fast‐glycolytic (FG), fast‐oxidative glycolytic (FOG), or slow‐oxidative (SO) according to the qualitative histochemical staining scheme described by Peter et al. (1972). Most muscles showed regional variations in fiber‐type distributions, i.e., the percent SO was higher and the percent FG was lower in the deep, compared to the superficial, regions of the muscle. Exceptions were the soleus and plantaris muscles, which contained similar distributions of fiber types throughout their cross sections. In the extensor compartment of the leg, a layering of fiber types from deep to superficial were evident in the triceps surae and plantaris complex with the deepest muscle, the soleus, having primarily SO fibers. A similar layering arrangement was observed in the extensor compartment of the thigh, with the deepest muscle, the vastus intermedius, having a much larger proportion of SO fibers than the other muscles in the quadriceps complex. These results indicate that Cynomolgus monkey hindlimb muscles, unlike human leg muscles (Saltin and Gollnick: Handbook of Physiology, L.D. Peachey, ed. American Physiological Society, MD, pp. 55–631, 1983) have a regional distribution of fiber types similar to that observed in many subprimate mammals. Further, the presence of compartmentalization of fiber types within the cross section of several of the muscles studied is suggestive of structure‐function interrelationships related to motor control.
33258052
Contractile properties of rat, rhesus monkey, and human type I muscle fibers.
It is well known that skeletal muscle intrinsic maximal shortening velocity is inversely related to species body mass. However, there is uncertainty regarding the relationship between the contractile properties of muscle fibers obtained from commonly studied laboratory animals and those obtained from humans. In this study we determined the contractile properties of single chemically skinned fibers prepared from rat, rhesus monkey, and human soleus and gastrocnemius muscle samples under identical experimental conditions. All fibers used for analysis expressed type I myosin heavy chain as determined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Allometric coefficients for type I fibers from each muscle indicated that there was little change in peak tension (force/fiber cross-sectional area) across species. In contrast, both soleus and gastrocnemius type I fiber maximal unloaded shortening velocity (Vo), the y-intercept of the force-velocity relationship (Vmax), peak power per unit fiber length, and peak power normalized for fiber length and cross-sectional area were all inversely related to species body mass. The present allometric coefficients for soleus fiber Vo (-0.18) and Vmax (-0.11) are in good agreement with published values for soleus fibers obtained from common laboratory and domesticated mammals. Taken together, these observations suggest that the Vo of slow fibers from quadrupeds and humans scale similarly and can be described by the same quantitative relationships. These findings have implications in the design and interpretation of experiments, especially those that use small laboratory mammals as a model of human muscle function.
45401511
Effects of tension and stiffness due to reduced pH in mammalian fast‐ and slow‐twitch skinned skeletal muscle fibres.
1. The pH dependence of the Ca2+ sensitivities of isometric tension and stiffness was investigated at 10 and 15 degrees C in skinned single fibres from rat and rabbit fast‐ and slow‐twitch skeletal muscles. Stiffness was determined by recording the tension responses to sinusoidal length changes (3.3 kHz); peak‐to‐peak amplitude of the length change was monitored by laser diffraction and averaged approximately 1.3 nm (half‐sarcomere)‐1. We have assumed that stiffness provides a measure of the number of cross‐bridge attachments to actin. 2. At maximal Ca2+ activation, stiffness was depressed by 22 +/‐ 2% (mean +/‐ S.E.M.) in fast‐twitch fibres but was unchanged in slow‐twitch fibres when pH was lowered from 7.00 to 6.20. As reported previously, maximum tension was depressed by 20‐45% at low pH, with the effect being greater in fast‐twitch compared to slow‐twitch fibres. 3. In fast‐twitch fibres at 10 and 15 degrees C the Ca2+ concentrations for half‐maximal activation of tension and stiffness were increased at low pH. In slow‐twitch fibres, similar effects were observed at 15 degrees C, but at 10 degrees C there were no changes in the Ca2+ sensitivities of tension and stiffness when pH was lowered. 4. Linear relationships between relative tension and relative stiffness were obtained at all temperatures, with slopes of 1.04 +/‐ 0.01 at pH 7.00 and 0.76 +/‐ 0.01 at pH 6.20 in fast‐ and slow‐twitch fibres, indicating that force per cross‐bridge attachment is similarly reduced at low pH in both types of fibres. 5. In both fast‐ and slow‐twitch fibres, rigor tension (no added ATP or creatine phosphate; pCa 9.0) was depressed by 35 +/‐ 7% and stiffness by 12 +/‐ 4% when pH was reduced from 7.00 to 6.20. Since cross‐bridge cycling is inhibited in rigor the effect of low pH to depress rigor tension suggests that pH directly modulates the strength of the bond formed between actin and myosin. 6. The effect of low pH to reduce the apparent number of cross‐bridge attachments during maximal Ca2+ activation in fast‐ but not slow‐twitch fibres could account for the greater H(+)‐induced depression of maximum force in fast‐twitch fibres. In both fibre types, the decrease in the number of cross‐bridge attachments at submaximal concentrations of Ca2+ may in part account for the loss in Ca2+ sensitivity of tension at low pH, due perhaps to a reduction in co‐operative activation of the thin filament by bound cross‐bridges.
25480373
Isometric force and maximal shortening velocity of single muscle fibers from elite master runners.
Single chemically permeabilized gastrocnemius fibers from six elite endurance-trained master runners (RUN group) and five age-matched sedentary controls (SED group) were mounted between a force transducer and a position motor, studied under conditions of maximal and submaximal Ca2+ activation, and subsequently electrophoresed on 5% polyacrylamide gels to determine myosin heavy chain (MHC) composition. For the SED group, peak isometric tension (Pzero) averaged 143 +/- 3, 156 +/- 4, and 170 +/- 4 kN/m2 and maximal shortening velocity (Vzero) averaged 0.43 +/- 0.01, 1.90 +/- 0.08, and 5.59 +/- 0.40 fiber lengths/s for fibers expressing type I, IIa, and IIx MHC, respectively (all comparisons, P < 0.05). Hill plot analysis of relative forces during submaximal Ca2+ activation indicated no SED vs. RUN differences in Ca2+ sensitivity or in the cooperativity of Ca2+ activation. However, at maximal Ca2+ activation, RUN type I and IIa fibers produced 15% less peak absolute force than SED fibers (P < 0.05). This reduction in fiber force was a direct result of the smaller diameter of the RUN fibers (P < 0.05), because Pzero, peak elastic modulus (Ezero), and Pzero/Ezero were not different between SED and RUN groups. RUN type I fibers also displayed a mean Vzero that was 19% higher than the average Vzero of the SED type I fibers (P < 0.05). In separate experiments, quantification of relative myosin light chain (MLC) isoform content revealed a 28% greater ratio of MLC3 to MLC2 in single type I fibers from the RUN group (P < 0.05), suggesting that the elevated Vzero of the RUN type I fibers was related to a greater expression of MLC3. In conclusion, the single fibers from the elite master runners displayed specific morphological and contractile properties that may enhance the performance of these athletes during prolonged muscular activity.
32346161
Force‐velocity relations and myosin heavy chain isoform compositions of skinned fibres from rat skeletal muscle.
1. This study was performed to assess whether muscle contractile properties are related to the presence of specific myosin heavy chain (MHC) isoforms. 2. Force‐velocity relations and MHC isoform composition were determined in seventy‐four single skinned muscle fibres from rat soleus, extensor digitorum longus and plantaris muscles. 3. Four groups of fibres were identified according to their MHC isoform composition determined by monoclonal antibodies: type 1 (slow), and types 2A, 2B and 2X (fast). 4. With respect to maximum velocity of shortening (V0), the fibres formed a continuum between 0.35 and 2.84 L/s (muscle lengths per second) at 12 degrees C. V0 in type 1 fibres (slow fibres) was between 0.35 and 0.95 L/s (0.639 +/‐ 0.038 L/s; mean +/‐ S.E. of mean). V0 in type 2 fibres (fast fibres) was consistently higher than 0.91 L/s. Ranges of V0 in the three fast fibre types mostly overlapped. Type 2A and 2X fibres had similar mean V0 values (1.396 +/‐ 0.084 and 1.451 +/‐ 0.066 L/s respectively); type 2B fibres showed a higher mean V0 value (1.800 +/‐ 0.109 L/s) than type 2A and 2X fibres. 5. Mean values of a/P0, an index of the curvature of force‐velocity relations, allowed us to identify two groups of fibres: a high curvature group comprised of type 1 (mean a/P0, 0.066 +/‐ 0.007) and 2A (0.066 +/‐ 0.024) fibres and a low curvature group comprised of type 2B (0.113 +/‐ 0.013) and 2X (0.132 +/‐ 0.008) fibres. 6. Maximal power output was lower in slow fibres than in fast fibres, and among fast fibres it was lower in type 2A fibres than in type 2X and 2B. 7. Force per unit cross‐sectional area was less in slow fibres than in fast fibres. There was no relation between fibre type and cross‐sectional area. 8. The results suggest that MHC composition is just one of the determinants of shortening velocity and of other muscle contractile properties.
45351036
Force-velocity and force-power properties of single muscle fibers from elite master runners and sedentary men.
Gastrocnemius muscle fiber bundles were obtained by needle biopsy from five middle-aged sedentary men (SED group) and six age-matched endurance-trained master runners (RUN group). A single chemically permeabilized fiber segment was mounted between a force transducer and a position motor, subjected to a series of isotonic contractions at maximal Ca2+ activation (15 degrees C), and subsequently run on a 5% polyacrylamide gel to determine myosin heavy chain composition. The Hill equation was fit to the data obtained for each individual fiber (r2 > or = 0.98). For the SED group, fiber force-velocity parameters varied (P < 0.05) with fiber myosin heavy chain expression as follows: peak force, no differences: peak tension (force/fiber cross-sectional area), type IIx > type IIa > type I; maximal shortening velocity (Vmax, defined as y-intercept of force-velocity relationship), type IIx = type IIa > type I; a/Pzero (where a is a constant with dimensions of force and Pzero is peak isometric force), type IIx > type IIa > type I. Consequently, type IIx fibers produced twice as much peak power as type IIa fibers, whereas type IIa fibers produced about five times more peak power than type I fibers. RUN type I and IIa fibers were smaller in diameter and produced less peak force than SED type I and IIa fibers. The absolute peak power output of RUN type I and IIa fibers was 13 and 27% less, respectively, than peak power of similarly typed SED fibers. However, type I and IIa Vmax and a/Pzero were not different between the SED and RUN groups, and RUN type I and IIa power deficits disappeared after power was normalized for differences in fiber diameter. Thus the reduced absolute peak power output of the type I and IIa fibers from the master runners was a result of the smaller diameter of these fibers and a corresponding reduction in their peak isometric force production. This impairment in absolute peak power production at the single fiber level may be in part responsible for the reduced in vivo power output previously observed for endurance-trained athletes.
11795416
Variations in contractile properties of rabbit single muscle fibres in relation to troponin T isoforms and myosin light chains.
1. The maximal velocity of shortening (Vmax), tension‐pCa relationships and the contractile and regulatory protein composition were determined in single, chemically skinned fibres from adult rabbit plantaris muscles. 2. Three groups of fibres were identified based on their protein compositions. One group had exclusively the slow‐type myosin heavy chain (MHC) and myosin light chains (LC) and had low velocities. Another group of fibres had mixtures of fast‐type and slow‐type MHCs and LCs and had intermediate shortening velocities. The third group of fibres had fast‐type myosin heavy and light chains and high velocities. 3. The low‐velocity fibres had a mean velocity (+/‐ S.E.M.) of 0.86 +/‐ 0.03 muscle lengths/s (ML/s) at 15 degrees C. The remaining fibres formed a continuum with respect to Vmax from 1.37 to 3.94 ML/s. These results indicate that a much greater diversity exists among single fibres from adult mammalian skeletal muscle than previously recognized. The intermediate‐ and high‐velocity fibres formed a continuum (from slow to fast) with respect to the amount of myosin light chain 3 (LC3). That is, Vmax increased with the relative LC3 content in single fibres in the intermediate‐ and high‐velocity groups in a quantitative, statistically significant manner. 4. Three isoforms of fast‐type troponin T were identified among the intermediate‐ and high‐velocity fibres. These fibres also contained fast‐type troponin C and troponin I. As was the case with the relative LC3 content, these fibres also formed a continuum with respect to the relative proportions of the three isoforms of fast‐type troponin T. It appears that different isoforms of troponin T are responsible for a slightly higher Ca2+ sensitivity of tension development in the high‐velocity fibres compared to the intermediate fibres. The continuum in troponin T isoform composition paralleled an increase in Vmax among these fibres. 5. The low‐velocity fibres had the highest Ca2+ sensitivity of the three groups and had exclusively the slow‐type isoforms of the regulatory proteins in the troponin complex. 6. The co‐ordinated variations in troponin T and LC3 compositions among the intermediate‐ and high‐velocity fibres are discussed as a possible means for the further differentiation of the contractile properties of the fibres in these two groups, beyond that provided by myosin heavy chain isoforms alone.
32914784
Altered Ca2+ dependence of tension development in skinned skeletal muscle fibers following modification of troponin by partial substitution with cardiac troponin C.
Binding of Ca2+ to the troponin C (TnC) subunit of troponin is necessary for tension development in skeletal and cardiac muscles. Tension was measured in skinned fibers from rabbit skeletal muscle at various [Ca2+] before and after partial substitution of skeletal TnC with cardiac TnC. Following substitution, the tension-pCa relationship was altered in a manner consistent with the differences in the number of low-affinity Ca2+-binding sites on the two types of TnC and their affinities for Ca2+. The alterations in the tension-pCa relationship were for the most part reversed by reextraction of cardiac TnC and readdition of skeletal TnC into the fiber segments. These findings indicate that the type of TnC present plays an important role in determining the Ca2+ dependence of tension development in striated muscle.