4.2 Skeletal Muscle

Skeletal muscle tissue attaches to the skeleton and generates voluntary body movements. Each skeletal muscle is composed of many individual muscle fibers, each of which is multinucleated, and cross striated. Image 4.2a is an electron micrograph scan of skeletal muscle. The connective tissue (fascia) that invests an entire muscle is called the epimysium (labeled "Ep" in image 4.2a). Fascia also penetrates the muscle, separating the muscle fibers into bundles called fasciculi (labeled "Fa" in image 4.2a). This fascia which surrounds each fascicle is called the perimysium, and very thin extensions of the fascia, called the endomysium, envelop the cell membrane of each individual muscle fiber (labeled "Fi" in image 4.2a). Blood vessels (labeled "BV" in image 4.2a) and nerves pass into the muscle near these fascia sheaths in order to reach the individual muscle cells. Beds of capillaries are located among the muscle cells, and each muscle fiber is innervated by its own individual nerve.

Image 4.2b shows a close view of a fasciculus which occupies almost the entire image. Observe the thick white perimysium layer which surrounds the fasciculus and strain your eyes to see the individual muscle fibers, each of which is surrounded by a thin line (pink) of endomysium.

Image 4.2c is a magnified view of the muscle fibers within the fasciculus. Note the thin, lighter-colored line of endomysium located at the tip of the arrow. You can also observe a number of very small purple nuclei which are located just under the membrane of the muscle fibers throughout the entire slide.

Image 4.2d is an example of dermatomyositis, an inflammatory disease of muscle in which the muscle fibers degenerate and die. Untreated, this disease leads to progressive and sporadic weakening of muscles and can present a serious mortality threat.

4.3 Muscle Fibers

When skeletal muscle is viewed microscopically (as seen in image 4.3a), it is composed of many elongated, cylindrical cells called muscle fibers. These fibers run parallel to one another and some fibers may be as long as 12 inches or more in length. Each muscle fiber is enveloped by a plasma membrane called the sarcolemma. Under the sarcolemma is found the cytoplasm of the cell which is called sarcoplasm. Within the sarcoplasm of the muscle fiber, lying close to the sarcolemma, are many nuclei and each muscle fiber contains a large number of mitochondria. Skeletal muscle fibers are multinucleated because of their extreme length and they are packed with numerous mitochondria because of their high ATP demand.

When longitudinally sectioned skeletal muscle fibers are stained with iron hematoxylin, their myofibrils (labeled "Fi" in image 4.3a) exhibit an alternating light and dark banding pattern. The dark A-bands (labeled "A" in image 4.3a) are bisected by a thin, light H-band (labeled "H" in image 4.3a). Adjacent light-staining I-bands (labeled "I" in image 4.3a) are also bisected, but by a thin dark line termed the Z-line (labeled "Z" in image 4.3a). The area between two Z-lines is a unit of muscle organization known as the sarcomere. Nuclei (labeled "Nu" in image 4.3a) of the multinucleate skeletal muscle fibers are located just under the sarcolemma.

Image 4.3b is a light microscope scan where low-power view of skeletal muscle fibers showing their banding patterns can be observed. The arrow in image 4.3b points at an "A" band located on the upper muscle fiber. The border (labeled "a" in image 4.3b) separates two muscle fibers and many flat purple nuclei can be seen along this border.

Image 4.3c is a cross sectional view of muscle fibers. Each fiber is surrounded by a thin layer of endomysium (whitish-blue lines, labeled "b" in image 4.3c) and deep-staining purple nuclei (labeled "a" in image 4.3c) can be seen lying immediately under the cell membrane (sarcolemma). Internal to each fiber numerous dot-like myofibrils can be seen.

4.4 Endomysium

Image 4.4a is an example of endomysium (labeled "En" in image 4.4a) enveloping three, vertically-oriented muscle fibers. Note that the outer cell membrane, the sarcolemma (labeled "Sa" in image 4.4a) of one of the fibers has been ripped away (arrows in image 4.4a) revealing the clearly striated myofibrils (labeled "St" in image 4.4a) inside of the cell. Endomysium is important for stabilizing and linking muscle fibers to one another. It is a rather thin collagenous layer that joins to the thickened perimysium layer which unites muscle fasciculi (bundles of muscle fibers). The perimysium layers, in turn, unite to join the outer epimysium layer which surrounds and supports the entire muscle.

Image 4.4b shows a malignant tumor of fibroblasts, the cell type which secretes the collagen of the endomysium. It is called fibrosarcoma, a connective tissue tumor which is most usually found in the thigh or lower leg (as in this amputated leg specimen), usually near the knee.

Image 4.4c is the microscopic detail of this disease. Note the numerous (dark) misshaped fibroblasts which form densely interlacing bundles and fascicles, showing a typical "herringbone" pattern.

4.5 Muscle Vascularization

Image 4.5a is a scanning electron micrograph of a blood vessel cast taken from skeletal muscle. Since skeletal muscle is such an ATP-demanding tissue during contraction, an adequate blood flow is critical for the delivery of oxygen and oxidative metabolism. Further, since muscle fibers forcibly shorten, the extensive vascular network must be able to deliver blood as the individual fibers lengthen and contract. Note the functional organization of the blood vessel network in the slide. Blood arrives via the relatively large artery to the right. This process to subdivide to ultimately form arterioles that feed an extensive capillary plexus. The capillary plexus is seen in the lower center of the image as a series of horizontal, parallel vessels with numerous vertical interconnections. The major capillaries show regular spacing (which corresponds to the diameter of individual muscle fibers) so that they run parallel and in between the muscle fibers. Should one of these parallel vessels get pinched during contraction, the interconnections, called transverse capillary loops, can maintain the critical blood flow. You must also remember that this muscle vascular system is very dynamic: regular exercise will enhance the vascularity (develop more capillaries and anastomoses) and functionally deliver more blood to the exercised muscle.

Image 4.5b is an example of a tumor that arises from the endothelial cells of blood vessels. These benign vascular tumors, called hemangiomas, can occur in any organ, including muscle. Capillary hemangiomas which infiltrate between muscle fibers have been well described in the clinical literature. Cavernous hemangiomas, such as the one seen in the arm of this baby, can become quite large. Histologically, these lesions are characterized by numerous blood-filled spaces lined by benign endothelial cells. Image 4.5c is a microscopic example of this disease.

4.6 Myofibrils

Muscle fibers (which are cells) are made up of smaller units (organelles) called fibrils. These fibrils, which can number from hundreds to thousands per fiber, run longitudinally through the muscle cell and consist of even smaller structures called filaments. The myofilaments of a myofibril do not extend the entire length of a muscle fiber, but instead they are separated into compartments that are called sarcomeres. Sarcomeres are separated from one another by very narrow zones of dense protein material called "Z" lines. In a relaxed muscle fiber, that is, one that is not contracting, the thin and thick myofilaments overlap and form a dark, dense band called the anisotropic band, or A band. The A band in image 4.6a shows vertically-oriented myofibrils (labeled "Mf" in image 4.6a) showing their repeating sarcomeres (labeled "Sa" in image 4.6a). The horizontal A bands alternate with light-colored, less dense areas called isotropic bands, or I bands (labeled "I" in image 4.6a) which are composed of thin myo- filaments only. This combination of alternating dark and light A & I bands gives the muscle fiber its striped appearance. A narrow H zone (labeled "H" in image 4.6a) within each A band contains thick myofilaments only. In the center of each H zone is the "M" line, a series of fine threads that appear to connect the middle sections of the thick filaments. Between the myofibrils numerous mitochondria (labeled "M" in image 4.6a) can be seen.

In image 4.6b, the numerous myofibrils (one runs between the arrows in image 4.6b) are running obliquely from top left to lower right. The structure labeled "A" in image 4.6b is the nucleus inside of the cell and note that each myofibril is formed of repeating numbers of sarcomeres. Each sarcomere is bounded by Z-line partitions (seen as very dark black lines running across each fibril) which enclose the myofilaments that contract the muscle.

Image 4.6c is an example of Duchenne muscular dystrophy. This disease, which is a sex-linked genetic disorder, weakens muscles progressively, usually making patients wheelchair- bound by 10 years old. Histologically, the muscle fibers degenerate as the internal fibrils are destroyed, producing the classic cross-sectional picture as seen in image 4.6c. The darker green fibers are degenerating, and you can see an "endomysial fibrosis" (light areas surrounding the cells) as collagen deposits around the individual muscle fibers.

Image 4.6d is an example of neurogenic atrophy of muscle fibers. When motor neurons are damaged and the innervating fibers degenerate, then sarcoplasmic elements, including the myofibrils, undergo destruction, and the cell shrinks, becoming non-functional. These atrophic cells are seen in this cross-sectional slide as small, dark orange cells undergoing "angular atrophy".

4.7 Sarcomere

Image 4.7a is a high-resolution transmission electron micrograph of four fibrils, illustrating the detailed structure of the sarcomere (between the arrows in image 4.7a). Note that the arrows in image 4.7a point to the Z-lines, which are the demarcations of each sarcomeric unit. The structure labeled "A" in image 4.7a is the sarcoplasmic reticulum, a network of flattened membrane tubules which release calcium to the sarcomeres. It is this calcium that acts as the immediate trigger for muscle contraction.

In image 4.7b, more internal detail of the sarcomere can be seen. Note the thick A band (labeled "A" in image 4.7a) which is the area in which actin and myosin filaments overlap and the central H band (labeled "H" in image 4.7a) which is a lighter area filled only with myosin filaments.

Image 4.7c is another more detailed view of the sarcomere with the Z-line is labeled "Z" in image 4.7c; the I band (labeled "I" in image 4.7c) is half of the light-colored zone see between striations; the A band (labeled "A" in image 4.7c) is half of the striation band itself; the H zone (labeled "H" in image 4.7c) is the lighter-colored area within the A band that contains only myosin filaments; and the M-line (labeled "M" in image 4.7c), a darker line within the H zone that reveals interconnections that stabilize the myosin filaments.

Image 4.7d is an example of how muscular dystrophy affects the sarcomere structure. Note the disorganization of the sarcomeres in the upper left section of the slide as the myofilaments are undergoing dissolution.

4.8 Filaments

Image 4.8a is a diagram of the structure of the two major filaments that comprise skeletal muscle: actin (pink structure in image 4.8a) and myosin (blue structure in image 4.8a). The myosin filaments are normally much thicker than the actin filaments, but here the actins have been magnified in size to reveal their component molecules. The actins are bonded to the Z-line which demarcates the end of the sarcomere. Each actin consists of a double twisted strand of F actin, each of which is a long necklace of globular G actin molecules (pink spheres in image 4.8a). Lying in the grooves formed by the twisting of the F actin chains are the repeating tropomyosin molecules which guard the actin filament and prevent the heads of the myosin cross bridges from attaching to the filament. Associated with each tropomyosin molecule is a smaller troponin complex. When calcium is released into the sarcomere, it binds to the troponin complex causing it to pull the tropomyosin away from the actin filament, exposing a binding site for the cross bridge of the myosin filament to attach and pull. The myosin filaments are a bundle of long myosin proteins, many of which have specialized cross bridges projecting out at their ends. When the tropomyosin has been removed from the binding site, the head of the myosin cross bridge attaches to actin, forming actinomyosin which causes the head to pull the actin filament. It can do this repeatedly, causing the actin filament to slide over the myosin and thereby producing a shortening of the sarcomere and generating overall muscle cell contraction.

Image 4.8b show the orderly arrangement of actin filaments (labeled "A" in image 4.8b) and the myosin filaments (labeled "B" in image 4.8b) within the sarcomeres. Observe the attachment of the actin filaments to the dark black Z-line and also note the relatively thicker dimensions of the myosin filaments compared to the actins.

4.9 Smooth Muscle

Smooth muscle tissue is common throughout the body, occurring in many of the soft organ systems. For example, in the wall of the alimentary canal it provides the motive power for mechanically mixing the food and propelling it through the digestive tract. Smooth muscle is also found in the walls of blood vessels where it allows for vasomotor control (constriction and dilation), in the walls of respiratory passages where it can open or close the air passageways, and in the urinary and reproductive ducts where it functions to expel urine or semen. In the human female the uterus is primarily composed of smooth muscle which serves as the engine for birth. Unlike skeletal muscle, smooth muscle is under autonomic (involuntary) nervous control and the tissue is quite distinct from that of striated muscle. Smooth muscle fibers are long, spindle-shaped cells that contain a single nucleus and lack striations (as seen in image 4.9a).

Image 4.9b is a the light microscope image of smooth muscle. Note the flattened purple nuclei located in the middle of spindle-shaped cells. Although it is difficult to see the borders of these cells, you can see that they are tightly packed, which allows the transmission of excitatory muscle potentials directly from one cell to another.

Image 4.9c is a cross-sectional view of smooth muscle tissue (bottom right of image 4.9c). Observe that the arrow points to a centrally-located nucleus in a smooth muscle cell. The upper left of image 4.9c is smooth muscle cells in a longitudinal view.

4.10 Cardiac Muscle

Cardiac muscle tissue composes the middle layer of the wall of the heart. It is characterized by branching muscle fibers, a centrally positioned nucleus, and tight-fitting cell boundaries called intercalated discs. These discs help to hold adjacent cells together and transmit the force of contraction from cell to cell. Like skeletal muscle, cardiac muscle is striated, but unlike skeletal muscle it generates rhythmical autogenic contractions. Image 4.10a is a low-power light microscope slide of cardiac muscle. Note the faint striations in the muscle fibers which lie horizontally in this slide (note also the flattened dark blue nuclei). The arrow in image 4.10a points to an intercalated disc that serves not only as a bonding site between cardiac cells but as an electrotonic junction that allows the passage of the muscle membrane potential from one cardiocyte to another.

Image 4.10b is another low-power light micrograph of cardiac muscle. Note here the highly branching, elongate muscle fibers with their dark blue nuclei. Close examination of this image reveals thin purple lines crossing the fibers. These are the intercalated discs which represent the tight anatomical and physiological junctions between the cardiocytes.

Image 4.10c is a comparative example of normal cardiac tissue (center section of image 4.10c) to hypertrophic tissue (left and right sections of image 4.10c). Note that the hypertrophic cardiac fibers are much thicker than the normal cells. This is due to the proliferation of sarcoplasmic elements, such as myofilaments.

Image 4.10d is a view into the cytoplasm of a cardiocyte. Running from top to bottom are rod-like myofibrils, crossed over by horizontal tubules that arise from sarcoplasmic reticulum tubules (lighter colored tubes running beside and on top of the myofibrils). Note the numerous mitochondria (labeled "Mi" in image 4.10d) located in between the myofibrils. So many mitochondria are required since cardiac muscle demands large amounts of ATP to fuel the continuous, periodic contraction of the heart.

4.11 Myoneural Junction

Image 4.11a show the general features of the myoneural (or neuromuscular) junction. The motor neuron sends a long process, the axon, to synapse with the muscle membrane. The actual junction is the location where the axon terminal (an expanded bulbous ending of the axon--sometimes called the button) makes functional contact with the muscle membrane (sarcolemma). The axolemma of the nerve does not actually contact the sarcolemma of the muscle fiber here, but is separated from it by a synaptic cleft. Also, the sarcolemma is highly modified in this area, being twisted into juctional folds and having numerous receptors for the synaptic transmitter substance (acetylcholine) that will be neurosecreted from the axon terminal. Within the axon terminal numerous synaptic vesicles which contain the transmitter molecules can be observed, as well as numerous mitochondria which supply the required energy for biochemical events of synaptic transmission. Briefly, the myoneural junction functions as a trigger point for the initiation of a muscle membrane potential that will stimulate the entire muscle fiber to contract. The nerve impulse that arrives at the myoneural junction causes a number of synaptic vesicles to move to the axolemma and neurosecrete acetylcholine molecules into the synaptic cleft. These molecules quickly diffuse across the cleft and combine with receptor sites located along the junctional folds of the sarcolemma. When sufficient numbers of these receptors are "keyed" by the acetylcholine molecules, the muscle sarcolemma is depolarized to its electrical threshold and a propagated muscle membrane potential is generated in all directions away from the myoneural junction.

Image 4.11b is a highly magnified electron micrograph of an actual myoneural junction. The nerve (axon terminal) is seen at the top image 4.11b and the muscle component at the bottom of image 4.11b. Note the numerous synaptic vesicles (labeled "B" in image 4.11b) in the nerve terminus, along with some darker-staining mitochondria (labeled "A" in image 4.11b). Toward the bottom of image 4.11b, the synaptic cleft at the arrow (labeled "C" in image 4.11b) is just below the basal lamina. Note also the highly infolded and thicker appearance of the muscle membrane (junctional folds) just below the synaptic cleft.

4.12 Tendon

Image 4.12a is a scanning electron micrograph of regular dense connective tissue that composes a tendon. Dense connective tissue, which is principally composed of bundles of collagen fibers, can be divided into regular or irregular types by the way in which the collagen bundles are arranged. If they are highly interwoven and oriented in all directions, then the tissue is called irregular dense connective tissue and resists stress in all directions. This is characteristic of the dermis of the skin and the dura mater of the brain. But if the collagen bundles are oriented in parallel in the same direction, then the tissue is termed regular dense connective tissue and it will resist stress very well in the direction of the parallel fibers. This type is very useful in tendons (and ligaments) where the strong force of muscle contraction is generated in a definitive direction and the tendon must resist tearing--either from the bone, internally, or from the muscle itself. Note the dense packing of the collagen fibers in the tendon and the parallel, unidirectional arrangement of the bundles.

Image 4.12b is a light micrograph of regular dense connective tissue of a tendon. Note the dense pink collagen fibers (labeled "a" in image 4.12b) and the scattered fibroblast cells which secrete them. These fibroblast cells (labeled "b" in image 4.12b) can be located by identifying their flat, horizontal, purple nuclei.

Image 4.12c is another example of normal tendon (labeled "a" in image 4.12c), here stained green. Again, note the flat fibroblast cells (with light blue nuclei) and contrast the appearance of this typical tendon structure to that of typical skeletal muscle tissue (seen as cross-banded, dark-purple fibers seen at the top of the image).

Image 4.12d is a comparative example of a pathological condition of tendon sheaths. This disease is called clear cell tumor of the tendon sheath. As can easily be seen, there is a striking multiplication of cellular masses ("clear cells") and a severe reduction in the amount of supporting collagen fibers.