In-series fiber architecture in long human muscles

The fiber architecture of adult human sartorius and gracilis muscles was examined using a combination of fiber microdissections and histological methods. Intact fibers were dissected from fascicles of muscle strips that were digested in nitric acid. All of these fibers terminate intrafascicularly by tapering to a fine strand at one or both ends. They measure 4–20 cm after correction for shrinkage. Systematic dissections of 1 cm long blocks sampled at intervals along the muscle length suggest that tapered fiber endings occur at all locations along the muscle but are most common centrally; here they accounted for up to 14% of dissected fibers in each block. Transverse sections of muscle confirm that fiber profiles with small diameters occur at all levels of the muscle but are especially common in sections more than 5 cm from its origin or insertion. The architectural arrangement demonstrated here suggests that long human muscles, like muscles in other species, are composed of relatively short, in-series fibers. This has many implications for the neural activation and force-developing behavior of these muscles that must be considered when paralyzed muscles are reanimated using electrical stimulation. Further, it may predispose long muscles to certain types of neuromuscular damage and dysfunction. © 1993 Wiley-Liss, Inc.     

Cross-linker system between neurofilaments, microtubules and membranous organelles in frog axons revealed by the quick-freeze, deep-etching method

The elaborate cross-connections among membranous organelles (MO), microtubules (MT), and neurofilaments (NF) were demonstrated in unifixed axons by the quick-freeze, deep-etch, and rotary-shadowing method. They were categorized into three groups: NF-associated cross-linker, MT-associated cross-bridges, and long cross-links in the subaxolemmal space. Other methods were also employed to make sure that the observed cross-connections in the unfixed axons were not a result of artifactual condensation or precipitation of soluble components or salt during deep-etching. Axolemma were permeablized either chemically (0.1% saponin) or physically (gentle homogenization), to allow egress of their soluble components from the axon; or else the axons were washed with distilled water after fixation. After physical rupture of the axolemma or saponin treatment, most of the MO remained intact. MT were stabilized by adding taxol in the incubation medium. Axons prepared by these methods contained many longitudinally oriented NF connected to each other by numerous fine cross-linkers (4-6 nm in diameter, 20-50 nm in length). Two specialized regions were apparent within the axons: one composed of fascicles of MT linked with each other by fine cross-bridges; the other was in the subaxolemmal space and consisted of actinlike filaments and a network of long cross-links (50-150 nm) which connected axolemma and actinlike filaments with NF and MT. F-actin was localized to the subaxolemmal space by the nitrobenzooxadiazol phallacidin method. MO were located mainly in these two specialized regions and were intimately associated with MT via fine short (10-20 nm in length) cross-bridges. Cross-links from NF to MO and MT were also common. All these cross-connections were observed after chemical extraction or physical rupture of the axon; however, these procedures removed granular materials which were attached to the filaments in the fresh unextracted axons. The cross-connections were also found in the axons washed with distilled water after fixation. I conclude that the cross- connections are real structures while the granular material is composed of soluble material, probably protein in nature.     

The composite structure of quail pectoralis muscle.

The twitch fibers of the quail pectoralis muscle were found to have one neuromuscular junction each, located in the middle third of the fiber. The length of isolated fibers varied between 8.8 and 33.2 mm, with mean and median values of 16 and 15.6 mm, respectively. The lengths of the fascicles from which the fibers were isolated varied between 30 and 51 mm. The muscle fibers taper at both ends. The neuromuscular junctions, revealed after histochemically reacting the intact muscle for acetyl cholinesterase activity, were arranged in discrete bands, separated by intervals of between 0.94 and 6.70 mm, with a mean value of 3.14 mm. The quail pectoralis muscle is thus composed of discontinuous, tapered muscle fibers, arranged in an overlapping series. It is therefore a muscle in which tension is transmitted laterally between muscle fibers.     

Histological study of motor innervation to long nuclear chain intrafusal fibers in the muscle spindle of the cat

Several muscle spindles of the cat tenuissimus muscle were cut in serial, 1-micron thick transverse sections and stained with toluidine blue in search for long nuclear chain intrafusal muscle fibers. Five complete poles of the long chain fibers were located. Each fiber pole displayed one plate-type motor ending situated in the extracapsular fiber region. The endings were supplied by myelinated motor axons that originated from intramuscular nerve fascicles containing motor axons to extrafusal muscle fibers. One of the endings was innervated by a collateral from a motor axon that supplied an extrafusal end-plate. Ultrastructurally, the long chain endings resembled extrafusal end-plates. They were more complex, in terms of prominence of sole-plate and degree of post-junctional folding, than any other intrafusal ending present in the spindles. The motor endings of the long chain fibers were assumed to be the terminals of static (fast) skeletofusimotor axons, which preferentially innervate the longest nuclear chain fibers of cat muscle spindles.     

Proportions of slow myosin heavy chain-positive fibers in muscle spindles and adjoining extrafusal fascicles, and the positioning of spindles relative to these fascicles.

Chicken leg muscles were examined to calculate the percentages of slow myosin heavy chain (MHC)-positive fibers in spindles and in adjacent extrafusal fascicles, and to clarify how the encapsulated portions of muscle spindles are positioned relative to these fascicles. Unlike mammals, in chicken leg muscles slow-twitch MHC and slow-tonic MHC are expressed in intrafusal fibers and in extrafusal fibers, suggesting a close developmental connection between the two fiber populations. In 8-week-old muscles the proportions of slow MHC-positive extrafusal fibers that ringed muscle spindles ranged from 0-100%. In contrast, proportions of slow MHC-positive intrafusal fibers in spindles ranged from 0-57%. Similar proportions in fiber type composition between intrafusal fibers and surrounding extrafusal fibers were apparent at embryonic days 15 and 16, demonstrating early divergence of extrafusal and intrafusal fibers. Muscle spindles were rarely located within single fascicles. Instead, they were commonly placed where several fascicles converged. The frequent extrafascicular location of spindles suggests migration of intrafusal myoblasts from developing clusters of extrafusal fibers toward the interstitium, perhaps along a neurotrophic gradient established by sensory axons that are advancing in the connective tissue matrix that separates adjoining fascicles.     

Neuromuscular organization of feline anterior sartorius: II. Intramuscular length changes and complex length-tension relationships during stimulation of individual nerve branches.

The feline anterior sartorius is a long strap-like muscle composed of short muscle fibers. Nerve branches that enter this muscle contain the axons of motor units whose constituent muscle fibers are distributed asymmetrically within the muscle. In the present study, twitch and tetanic isometric contractions were evoked by stimulating individual nerve branches while muscle force was recorded and intramuscular length changes were monitored optically by the movement of reflective markers on the muscle. Contractions elicited by stimulating the parent nerve produced little change in the positions of the surface markers. Contractions elicited by stimulating the proximally or distally directed nerve branches caused the muscle to shorten at the end closest to the nerve branch and lengthen at the opposite end. Some muscles were supplied by a centrally directed nerve branch whose stimulation produced variable effects: in some cases a portion of the muscle shortened whereas the rest lengthened, but in other cases, the positions of the surface markers showed little change. The intramuscular length changes produced by stimulating single nerve branches were greater during isometric contractions at short whole-muscle lengths than at long whole-muscle lengths. The twitch and tetanic length-tension relationships obtained by stimulating the individual nerve branches were not congruent with the length-tension relationship produced when the parent nerve was stimulated. At short whole-muscle lengths, stimulation of a single nerve branch generated only a small fraction of the force that could be generated by the muscle when the parent nerve was stimulated. As whole-muscle length increased, an increased fraction of total muscle force could be generated by stimulating a single nerve branch. The results suggest that a complex relationship between passive and active elements contributes to the total muscle force and depends on the distribution of active and passive muscle units throughout the muscle.     

Limb muscle regeneration in peripheral nerve autografts

In a previous study we demonstrated regenerative growth of extraocular muscle within transplanted peripheral nerve autografts. The present study addresses the feasibility of inducing regeneration of limb muscle within autologous peripheral nerve implants in the gluteus medius of beagles. In six anesthetized animals, a 2-cm segment of the left infraorbital sensory nerve was removed from the nose and implanted between the cut ends of several muscle fascicles in the left gluteus medius. After 4 weeks, the nerve grafts were removed and examined by light and electron microscopy. Muscle fibers were seen surrounded by the epineurium of the implanted nerve along its entire length, growing in parallel with the long axis of the nerve. The regenerating fibers were closely associated with the basal lamina of degenerating myelinated and unmyelinated axons. This study suggests that limb muscle, like extraocular muscle, is capable of organized regenerative growth within peripheral nerve autografts.     

Muscle fiber and tendon length changes in the human vastus lateralis during slow pedaling.

Muscle fascicle lengths of vastus lateralis (VL) muscle were measured in five healthy men during slow pedaling to investigate the interaction between muscle fibers and tendon. Subjects cycled at a pedaling rate of 40 rpm (98 W). During exercise, fascicle lengths changed from 91 +/- 7 (SE) to 127 +/- 5 mm. It was suggested that fascicles were on the descending limb of their force-length relationship. The average shortening velocity of fascicle was greater than that of muscle-tendon complex in the first half of the knee extension phase and was less in the second half. The maximum shortening velocity of fascicle in the knee extension phase was less than that of muscle-tendon complex by 22 +/- 9%. These discrepancies in velocities were mainly caused by the elongation of the tendinous tissue. It was suggested that the elasticity of VL tendinous tissue enabled VL fascicles to develop force at closer length to their optimal length and kept the maximum shortening velocity of VL fascicles low during slow pedaling.     

In vivo behavior of muscle fascicles and tendinous tissues in human tibialis anterior muscle during twitch contraction

In this study we investigated the time course of length and velocity of muscle fascicles and tendinous tissues (TT) during isometric twitch contraction, and examined how their interaction relates to the time course of external torque and muscle fascicle force generation. From seven males, supra-maximal twitch contractions (singlet) of the tibialis anterior muscle were induced at 30 degrees , 10 degrees and -10 degrees plantar flexed positions. The length and velocity of fascicles and TT were determined from a series of their transverse ultrasound images. The maximal external torque appeared when the shortening velocity of fascicles was zero. The fascicle and TT length, and external torque showed a 10-30 ms delay of each onset, with a significant difference in half relaxation times at -10 degrees . The time course of TT elongation, and fascicle and tendinous velocities did not differ between joint angles. Curvilinear length-force properties, whose slope of quasi-linear part was ranged from -15.0 to -5.9 N/mm for fascicles and 5.4 to 14.3N/mm for TT, and a loop-like pattern of velocity-force properties, in which the mean power was ranged from 0.14 to 0.80 W for fascicles, and 0.14 to 0.81 W for TT were also observed. These results were attributed to the muscle-tendon interaction, depending on the slack and non-linearity of length-force relationship of compliant TT. We conclude that the mechanical interaction between fascicles and TT, are significant determinants of twitch force and time characteristics.     

Location along the muscle's length is a determinant of myofibril size

In a previous study of myofibril size in 'Pale' (fast-twitch-glycolytic) fibers of rabbit extraocular muscle (EOM), it was found that individual long Pale fibers demonstrate a substantial increase in the size of myofibril profiles from their proximal to their distal halves (Davidowitz et al., 1996b). That finding raised the question of whether such proximal-to-distal increase of myofibril size in the Pale fibers is determined by: (1) longitudinal position within the individual muscle fibers themselves or (2) location along the length of the muscle as a whole? This question was tested in the present study by comparing the original group of long Pale fibers, which extend the full length of the muscle, with two groups of short Pale fibers, which are respectively confined to the proximal and distal halves of the muscle. It was found that (a) in the proximal half of the muscle, the short fibers and the adjacent portions of the long fibers have the same smaller size of myofibrils, and (b) in the distal half of the muscle, the short fibers and the adjacent portions of the long fibers have the same larger size of myofibrils. This finding indicates that the proximal-to-distal increase of myofibril-profile size in these EOM Pale fibers is determined by location along the length of the muscle as a whole, and is not related to longitudinal position within the individual fibers themselves.     

Architectural and histochemical analysis of the biceps brachii muscle of the horse.

The biceps brachii of horses is a complex muscle subdivided into two heads which may subserve distinct functions. The lateral head contains a large percentage of type I myofibers. This region is largely composed of short fibers (5-7 mm long) arranged in a pinnate fashion and heavily invested with connective tissue. The medial head contains fewer type I fibers and is composed of relatively longer myofibers (15-20 mm long), also arranged in a pinnate fashion but less heavily invested with connective tissue. It is hypothesized that the lateral muscle head of biceps brachii contributes to the postural role of the muscle in the forelimb passive stay apparatus. The medial head, with its longer fibers and generally fast fiber population may be most important during dynamic activity such as walking, trotting and running.     

Ultrastructure of striated muscle fibers in the middle third of the human esophagus

Striated muscle fibers and their spatial relationship to smooth muscle cells have been studied in the middle third of human esophagus. Biopsies were obtained from 3 patients during surgery. In both the circular and longitudinal layers, the muscle coat of this transition zone was composed of fascicles of uniform dimension (100-200 microns of diameter); some of these bundles were made up of striated muscle fibers, others were pure bundles of smooth muscle cells and some were of the mixed type. Striated muscle fibers represented three different types, which were considered as intermediate, with certain structural features characteristic of the fast fiber type. Of these, the most frequently-found fibers were most similar to the fast fiber type. Satellite cells were numerous; in mixed fascicles they were gradually replaced by smooth muscle cells. The gap between striated muscle fiber and smooth muscle cells was more than 200 nm wide. It contained the respective basal laminae and a delicate layer of amorphous connective tissue. No specialized junctions were formed between consecutive striated muscle fibers, or between striated muscle fibers and smooth muscle cells. Interstitial cells of Cajal were never situated as close to striated muscle fibers as to smooth muscle cells.     

Architectural properties of distal forelimb muscles in horses,Equus caballus

Articular injuries in athletic horses are associated with large forces from ground impact and from muscular contraction. To accurately and noninvasively predict muscle and joint contact forces, a detailed model of musculoskeletal geometry and muscle architecture is required. Moreover, muscle architectural data can increase our understanding of the relationship between muscle structure and function in the equine distal forelimb. Muscle architectural data were collected from seven limbs obtained from five thoroughbred and thoroughbred-cross horses. Muscle belly rest length, tendon rest length, muscle volume, muscle fiber length, and pennation angle were measured for nine distal forelimb muscles. Physiological cross-sectional area (PCSA) was determined from muscle volume and muscle fiber length. The superficial and deep digital flexor muscles displayed markedly different muscle volumes (227 and 656 cm3, respectively), but their PCSAs were very similar due to a significant difference in muscle fiber length (i.e., the superficial digital flexor muscle had very short fibers, while those of the deep digital flexor muscle were relatively long). The ulnaris lateralis and flexor carpi ulnaris muscles had short fibers (17.4 and 18.3 mm, respectively). These actuators were strong (peak isometric force, Fmax=5,814 and 4,017 N, respectively) and stiff (tendon rest length to muscle fiber length, LT:LMF=5.3 and 2.1, respectively), and are probably well adapted to stabilizing the carpus during the stance phase of gait. In contrast, the flexor carpi radialis muscle displayed long fibers (89.7 mm), low peak isometric force (Fmax=555 N), and high stiffness (LT:LMF=1.6). Due to its long fibers and low Fmax, flexor carpi radialis appears to be better adapted to flexion and extension of the limb during the swing phase of gait than to stabilization of the carpus during stance. Including muscle architectural parameters in a musculoskeletal model of the equine distal forelimb may lead to more realistic estimates not only of the magnitudes of muscle forces, but also of the distribution of forces among the muscles crossing any given joint.     

Contractile behavior of the forelimb digital flexors during steady-state locomotion in horses (Equus caballus): an initial test of muscle architectural hypotheses about in vivo function

The forelimb digital flexors of the horse display remarkable diversity in muscle architecture despite each muscle-tendon unit having a similar mechanical advantage across the fetlock joint. We focus on two distinct muscles of the digital flexor system: short compartment deep digital flexor (DDF(sc)) and the superficial digital flexor (SDF). The objectives were to investigate force-length behavior and work performance of these two muscles in vivo during locomotion, and to determine how muscle architecture contributes to in vivo function in this system. We directly recorded muscle force (via tendon strain gauges) and muscle fascicle length (via sonomicrometry crystals) as horses walked (1.7 m s(-1)), trotted (4.1 m s(-1)) and cantered (7.0 m s(-1)) on a motorized treadmill. Over the range of gaits and speeds, DDF(sc) fascicles shortened while producing relatively low force, generating modest positive net work. In contrast, SDF fascicles initially shortened, then lengthened while producing high force, resulting in substantial negative net work. These findings suggest the long fibered, unipennate DDF(sc) supplements mechanical work during running, whereas the short fibered, multipennate SDF is specialized for economical high force and enhanced elastic energy storage. Apparent in vivo functions match well with the distinct architectural features of each muscle.     

Localization of alpha 7 integrins and dystrophin suggests potential for both lateral and longitudinal transmission of tension in large mammalian muscles

Non-primate mammalian muscles with fascicles above 35 mm in length are composed predominantly of arrays of short, non-spanning muscle fibres, which terminate within the belly of the muscle fascicle at one or both ends. We have previously described the morphological form of various muscle-to-muscle and muscle-to-matrix junctions which are likely involved in tension transmission within one such muscle - the guinea pig sternomastoid muscle (Young et al. 2000). Here, we use immunohistochemistry to investigate the cell adhesion molecules present at these junctions. We find strong immunoreactivity against the alpha 7B integrin subunit and dystrophin, and slight reactivity against the alpha 7A integrin at all intrafascicular fibre terminations (IFTs), as well as at the muscle-tendon junction (MTJ). Tenascin, the sole ligand for alpha 9 beta 1 integrin, was absent from IFTs but present at the MTJ, suggesting the two sites are molecularly distinct. In addition to their expression at junctional sites, alpha 7B integrin and dystrophin were also expressed ubiquitously along the non-junctional sarcolemma, suggesting potential involvement in diffuse lateral transmission of tension between adjacent fibres. We conclude that the distribution of alpha 7 beta 1 integrins and dystrophin in series-fibred muscles suggests they are involved in transmission of tension from intrafascicularly terminating fibres to neighbouring fibres lying both in-series and in-parallel, via the extracellular matrix (ECM).     

Motor control of the mandible closer muscle in ants

Despite their simple design, ant mandible movements cover a wide range of forces, velocities and amplitudes. The mandible is controlled by the mandible closer muscle, which is composed of two functionally distinct subpopulations of muscle fiber types: fast fibers (short sarcomeres) and slow ones (long sarcomeres). The entire muscle is controlled by 10-12 motor neurons, 4-5 of which exclusively supply fast muscle fibers. Slow muscle fibers comprise a posterior and an antero-lateral group, each of which is controlled by 1-2 motor neurons. In addition, 3-4 motor neurons control all muscle fibers together. Simultaneous recordings of muscle activity and mandible movement reveal that fast movements require rapid contractions of fast muscle fibers. Slow and subtle movements result from the activation of slow muscle fibers. Forceful movements are generated by simultaneous co-activation of all muscle fiber types. Retrograde tracing shows that most dendritic arborizations of the different sets of motor neurons share the same neuropil in the subesophageal ganglion. In addition, fast motor neurons and neurons supplying the lateral group of slow closer muscle fibers each invade specific parts of the neuropil that is not shared by the other motor neuron groups. Some bilateral overlap between the dendrites of left and right motor neurons exists, particularly in fast motor neurons. The results explain how a single muscle is able to control the different movement parameters required for the proper function of ant mandibles.     

Reports of the length dependence of fatigue are greatly exaggerated.

Relative force depression associated with muscle fatigue is reported to be greater when assessed at short vs. long muscle lengths. This appears to be due to a rightward shift in the force-length relationship. This rightward shift may be caused by stretch of in-series structures, making sarcomere lengths shorter at any given muscle length. Submaximal force-length relationships (twitch, double pulse, 50 Hz) were evaluated before and after repetitive contractions (50 Hz, 300 ms, 1/s) in an in situ preparation of the rat medial gastrocnemius muscle. In some experiments, fascicle lengths were measured with sonomicrometry. Before repetitive stimulation, fascicle lengths were 11.3 +/- 0.8, 12.8 +/- 0.9, and 14.4 +/- 1.2 mm at lengths corresponding to -3.6, 0, and 3.6 mm where 0 is a reference length that corresponds with maximal active force for double-pulse stimulation. After repetitive stimulation, there was no change in fascicle lengths; these lengths were 11.4 +/- 0.8, 12.6 +/- 0.9, and 14.2 +/- 1.2 mm. The length dependence of fatigue was, therefore, not due to a stretch of in-series structures. Interestingly, the rightward shift that was evident when active force was calculated in the traditional way (subtraction of the passive force measured before contraction) was not seen when active force was calculated by subtracting the passive force that was associated with the fascicle length reached at the peak of the contraction. This calculation is based on the assumption that passive force decreases as the fascicles shorten during a fixed-end contraction. This alternative calculation revealed similar postfatigue absolute active force depression at all lengths. In relative terms, a length dependence of fatigue was still evident, but this was greatly diminished compared with that observed when active force was calculated with the traditional method.     

SNAREs in native plasma membranes are active and readily form core complexes with endogenous and exogenous SNAREs

Skeletal muscles display a remarkable diversity in their arrangement of fibers into fascicles and in their patterns of innervation, depending on functional requirements and species differences. Most human muscle fascicles, despite their great length, consist of fibers that extend continuously from one tendon to the other with a single nerve endplate band. Other mammalian muscles have multiple endplate bands and fibers that do not insert into both tendons but terminate intrafascicularly. We investigated whether these alternate structural features may dictate different modes of cell hypertrophy in two mouse gracilis muscles, in response to expression of a muscle-specific insulin-like growth factor (IGF)-1 transgene (mIGF-1) or to chronic exercise. Both hypertrophic stimuli independently activated GATA-2 expression and increased muscle cross-sectional area in both muscle types, with additive effects in exercising myosin light chain/mIGF transgenic mice, but without increasing fiber number. In singly innervated gracilis posterior muscle, hypertrophy was characterized by a greater average diameter of individual fibers, and centralized nuclei. In contrast, hypertrophic gracilis anterior muscle, which is multiply innervated, contained longer muscle fibers, with no increase in average diameter, or in centralized nuclei. Different modes of muscle hypertrophy in domestic and laboratory animals have important implications for building appropriate models of human neuromuscular disease.     

Architecture and consequent physiological properties of the semitendinosus muscle in domestic goats

Morphological and physiological analyses confirm that the semitendinosus muscle of goats contains two separate compartments in series, each with distinct innervation. These compartments of the muscle are in turn composed of short fibers (approximately four fibers in series in the proximal compartment and seven to eight fibers in the distal compartment) which overlap each other for more than 30% of their length, with much of the overlapping portions consisting of slender tails that terminate at one-tenth of the midfiber diameter. Groups of fibers are associated into relatively narrow bands that run end-to-end in each compartment. The data suggest that the maximum length of muscle fibers may be limited; even the fibers of parallel-fibered muscles may not scale with the dimension of the animal.     

Optimal muscle fascicle length and tendon stiffness for maximising gastrocnemius efficiency during human walking and running

Muscles generate force to resist gravitational and inertial forces and/or to undertake work, e.g. on the centre of mass. A trade-off in muscle architecture exists in muscles that do both; the fibres should be as short as possible to minimise activation cost but long enough to maintain an appropriate shortening velocity. Energetic cost is also influenced by tendon compliance which modulates the timecourse of muscle mechanical work. Here we use a Hill-type muscle model of the human medial gastrocnemius to determine the muscle fascicle length and Achilles tendon compliance that maximise efficiency during the stance phase of walking (1.2 m/s) and running (3.2 and 3.9 m/s). A broad range of muscle fascicle lengths (ranging from 45 to 70 mm) and tendon stiffness values (150-500 N/mm) can achieve close to optimal efficiency at each speed of locomotion; however, efficient walking requires shorter muscle fascicles and a more compliant tendon than running. The values that maximise efficiency are within the range measured in normal populations. A non-linear toe-region region of the tendon force-length properties may further influence the optimal values, requiring a stiffer tendon with slightly longer muscle fascicles; however, it does not alter the main results. We conclude that muscle fibre length and tendon compliance combinations may be tuned to maximise efficiency under a given gait condition. Efficiency is maximised when the required volume of muscle is minimised, which may also help reduce limb inertia and basal metabolic costs.