ELECTROMECHANICAL COUPLING IN TUBULAR MUSCLE FIBERS : I. The Organization of Tubular Muscle Fibers in the Scorpion Leiurus quinquestriatus

The tubular fibers of the claw-closer muscle of the scorpion have a central core containing nuclei and mitochondria. The myofibrils have the shape of thin lamellae (1 µ) extending radially from the core to the surface membrane (20 µ). The thick myofilaments are organized in a hexagonal array with orbits of 10–13 thin myofilaments. The ratio of thick-to-thin filaments is 1:5. Transverse tubular system (TS) openings are located between lamellated myofibrils. In each sarcomere two TS's are found, one on each side of the H band. The TS is composed of a transverse tubule and tubular pockets (TP). The TP's form diadic contact with the terminal cisternae of the sarcoplasmic reticulum. The TS can be traced from the cell membrane down to the cell core. The surface area of the TS was calculated to be six times that of the outer surface membrane.     

The fine structure of fast and slow crustacean muscles

Known phasic and tonic muscle fibers of the crab Cancer magister were studied by electron microscopy. Phasic fibers have sarcomeres about 4.5 µ long, small polygonal myofibrils, and a well-developed sarcoplasmic reticulum. The thick myofilaments, disposed in hexagonal array, are each surrounded by six thin filaments. The tonic fibers have a sarcomere length of about 12 µ, larger myofibrils, a poorly developed sarcoplasmic reticulum, and a disorderly array of myofilaments. Each thick myofilament is surrounded by 10–12 thin filaments. The same morphological type of slow muscle has been found in the crustaceans, Macrocyclops albidus, Cypridopsis vidua, and Balanus cariosus, in each case in an anatomical location consistent with tonic action. A search of the literature indicates that this type of muscle is found in all classes of arthropods and is confined to visceral and postural muscles or specializations of these.     

THE RELATIONSHIP BETWEEN MYOFILAMENT PACKING DENSITY AND SARCOMERE LENGTH IN FROG STRIATED MUSCLE

In cross-sections of single fibers from the frog semitendinosus muscle the number of thick myofilaments per unit area (packing density) is a direct function of the sarcomere length. Our data, derived from electron microscopic studies, fit well with other data derived from in vivo, low-angle X-ray diffraction studies of whole semitendinosus muscles. The data are consistent with the assumption that the sarcomere of a fibril maintains a constant volume during changes in sarcomere length. The myofilament lattice, therefore, expands as the sarcomere shortens. Since the distance between adjacent myofilaments is an inverse square root function of sarcomere length, the interaction of the thick and the thin myofilaments during sarcomere shortening may occur over distances which increase 70 A or more. The "expanding-sarcomere, sliding-filament" model of sarcomere shortening is discussed in terms of the current concepts of muscle architecture and contraction.     

OBSERVATIONS ON THE FINE STRUCTURE OF PHARYNGEAL MUSCLE IN THE PLANARIAN DUGESIA TIGRINA

Pharyngeal muscle of the planarian Dugesia tigrina was studied by electron microscopy after osmium tetroxide fixation. The muscle cell was observed to contain one myofibril or bundle of myofilaments parallel to its longitudinal axis. The myofilaments were of two types, different in size and distribution. No Z lines or myofilament organization into cross or helical striations were seen. Dense bodies were seen as projections from an invagination of the plasma membrane and as dense lines parallel to the myofilaments. The muscle cells are surrounded by a plasma membrane which is structurally associated with dense body projections, with vesicles and cisternae of sarcoplasmic reticulum, and with synaptic nerve endings. The cell has sarcoplasmic projections perpendicular to its long axis; these projections are seen to contain the nucleus or mitochondria and granules. Mitochondria and granules are also seen in a sarcoplasm rim around the fibril. The dense bodies may serve as attachment for thin myofilaments and function in transmission of stimuli from plasma membrane to the interior of the fibril.     

A comparative study on sub-filament organization in primary myofilaments of basalar direct flight and tibial extensor muscles of the lepidopteran,Achalarus lyciades

Summary Primary myofilaments of direct flight muscle fibers are hexagonally arranged, are surrounded by six secondary myofilaments, and are composed of two sub-unit-fibrils, 90–120 Å in diameter, at mid-sarcomere levels. Primary myofilaments are resolvable into electron-densities 15–25 Å in diameter, which number 3–4 in each sub-fibril at mid-sarcomere levels, and number 6–8 elsewhere. Primary myofilaments of tibial extensor muscle are not found in hexagonal arrays, are surrounded by 10–12 secondary myofilaments, and are resolvable into electron densities which are 15–25 Å in diameter similar to primary myofilaments of basalar flight muscle. However, a binary sub-fibril structure at mid-sarcomere levels is lacking in tibial extensor muscle fibers.The functional significance of the two-sub-fibril organization of myofilaments at midsarcomere levels in basalar direct flight muscle is not known, but may be related to the high rate of excitation-contraction cycles in these muscles.Supported by U.S.P.H.S. No. NB-06285. — The author wishes to express his grateful appreciation for the technical assistance given by Mrs. Ann Florendo during the course of this investigation.     

Fine Structure of Cardiac Sarcomeres in the Black Widow Spider Latrodectus mactans

Fine structural characteristics of the cardiac muscle and its sarcomere organization in the black widow spider, Latrodectus mactans were examined using transmission electron microscopy. The arrangement of cardiac muscle fibers was quite similar to that of skeletal muscle fibers, but they branched off at the ends and formed multiple connections with adjacent cells. Each cell contained multiple myofibrils and an extensive dyadic sarcotubular system consisting of sarcoplasmic reticulum and T‐tubules. Thin and thick myofilaments were highly organized in regular repetitive arrays and formed contractile sarcomeres. Each repeating band unit of the sarcomere had three apparent striations, but the H‐zone and M‐lines were not prominent. Myofilaments were arranged into distinct sarcomeres defined by adjacent Z‐lines with relatively short lengths of 2.0 μm to 3.3 μm. Cross sections of the A‐band showed hexagon‐like arrangement of thick filaments, but the orbit of thin filaments around each thick filament was different from that seen in other vertebrates. Although each thick filament was surrounded by 12 thin filaments, the filament ratio of thin and thick myofilaments varied from 3:1 to 5:1 because thin filaments were shared by adjacent thick filaments.     

ULTRASTRUCTURE OF BARNACLE GIANT MUSCLE FIBERS

Increasing use of barnacle giant muscle fibers for physiological research has prompted this investigation of their fine structure. The fibers are invaginated by a multibranched system of clefts connecting to the exterior and filled with material similar to that of the basement material of the sarcolemmal complex. Tubules originate from the surface plasma membrane at irregular sites, and also from the clefts They run transversely, spirally, and longitudinally, making many diadic and some triadic contacts with cisternal sacs of the longitudinal sarcoplasmic reticulum. The contacts are not confined to any particular region of the sarcomere. The tubules are wider and their walls are thicker at points of contact with Z material. Some linking of the Z regions occurs across spaces within the fiber which contain large numbers of glycogen particles. A-band lengths are extremely variable, in the range 2.2 µm–20.3 µm (average 5.2 µm) Individual thick filaments have thin (110 Å) hollow regions alternating with thick (340 Å) solid ones. Bridges between thick filaments occur at random points and are not concentrated into an M band The thin:thick filament ratio is variable in different parts of a fiber, from 3:1 to 6:1. Z bands are basically perforated, but the number of perforations may increase during contraction.     

CORRELATED MORPHOLOGICAL AND PHYSIOLOGICAL STUDIES ON ISOLATED SINGLE MUSCLE FIBERS : I. Fine Structure of the Crayfish Muscle Fiber

Single fibers isolated from walking leg muscles of crayfish have 8- to 10-µ sarcomeres which are divided into A, I, and Z bands. The H zone is poorly defined and no M band is distinguishable. Changes in the width of the I band, accompanied by change in the overlap between thick and thin myofilaments, occur when the length of the sarcomere is changed by stretching or by shortening the fiber. The thick myofilaments (ca. 200 A in diameter) are confined to the A band. The thin myofilaments (ca. 50 A in diameter) are difficult to resolve except in swollen fibers, when they clearly lie between the thick filaments and run to the Z disc. The sarcolemma invaginates at 50 to 200 sites in each sarcomere. The sarcolemmal invaginations (SI) form tubes about 0.2 µ in diameter which run radially into the fiber and have longitudinal side branches. Tubules about 150 A in diameter arise from the SI and from the sarcolemma. The invaginations and tubules are all derived from and are continuous with the plasma membrane, forming the transverse tubular system (TTS), which is analogous with the T system of vertebrate muscle. In the A band region each myofibril is enveloped by a fenestrated membranous covering of sarcoplasmic reticulum (SR). Sacculations of the SR extend over the A-I junctions of the myofibrils, where they make specialized contacts (diads) with the TTS. At the diads the opposing membranes of the TTS and SR are spaced 150 A apart, with a 35-A plate centrally located in the gap. It appears likely that the anion-permselective membrane of the TTS which was described previously is located at the diads, and that this property of the diadic structures therefore may function in excitation-contraction coupling.     

粘虫蛾飞行肌超微结构的研究

应用电子显微镜对粘虫雌蛾Mythimna separata(Walker)飞行(背纵)肌的研究结果表明,其肌原纤维由500-700根肌球蛋白丝(粗丝)组成,每根粗丝由6根肌动蛋白丝(细丝)环绕排列成六角形,每根细丝精确地位于两根粗丝间1/2处,从而使粗丝和细丝的比为1：3。肌节较短,长度约2.2-2.6μm。肌原纤维之间充满着线粒体和横管。每个肌节约有线粒体三个,横管二根。线粒体约占肌纤维体积的40%,而横管为7%。每根横管准确地位于肌节的1/4、3/4处,或Z线和中膈的中央,并与肌质网交接形成二位体(dyads)或三位体(triads)。肌质网相当不发达,约占肌纤维体积的2.5%。但其分布很有特色,即除了紧贴于肌原纤维周围的由单层液泡组成的肌质网以外,在中膈处还有一层横穿于肌原纤维的肌质网。和其它同步飞行肌的结构和功能分析比较的结果还表明,粘虫蛾飞行肌具有较善于飞行的结构。     

Ultrastructural features of cardiac muscle cells in a tarantula spider

The ultrastructural features of cardiac muscle cells and their innervation were examined in the tarantula spider Eurypelma marxi Simon. The cells are transversely striated and have an A band length of about three microns. H zones are indistinct and M lines are absent. Thick and thin myofilament diameters are approximately 200 and 70 Å respectively. Eight to 12 thin filaments usually surround each thick one. Accumulations of thick and thin myofilaments occur perpendicular to the bulk of the myofilaments in some cells. The Z line is discontinuous and thick filaments may pass through the spaces in the Z line. Extensive systems of sarcoplasmic reticulum and transverse tubules are present; these form numerous dyadic junctions in both A and I band regions. Sarcolemmal invaginations form Z line tubules; lateral extensions of the plasma membrane portion of these invaginations form dyads. Nerve branches of the cardiac ganglion make multiple neuromuscular synapses with at least some of the cardiac muscle cells. Both large granular and small agranular vesicles are present in the presynaptic terminals. Intercalated discs similar to those present in other arthropod hearts occur between the ends of adjacent cardiac muscle cells.     

STRUCTURE OF THE LONGITUDINAL BODY MUSCLES OF AMPHIOXUS

The structure of the longitudinal body muscles of Branchiostoma caribaeum has been studied by light and electron microscopy. These muscles are shown to be composed of fibers in the form of flat lamellae about 0.8µ in thickness, more than 100 µ wide, and reaching in length from one intermuscular septum to the next, a distance of about 0.6 mm. Each flat fiber is covered by a plasma membrane and contains a single myofibril consisting of myofilaments packed in the interdigitating hexagonal array characteristic of vertebrate striated muscle. Little or no sarcoplasmic reticulum is present. Mitochondria are found infrequently and have a tubular internal structure. These morphological observations are discussed in relation to a proposed hypothesis of excitation-contraction coupling. It is pointed out that the maximum distance from surface to myofilament in these muscles is about 0.5 µ and that diffusion of an "activating" substance over this distance would essentially be complete in less than 0.5 msec. after its release from the plasma membrane. It is concluded that the flat form of amphioxus muscle substitutes for the specialized mechanisms of excitation-contraction coupling thought possibly to involve the sarcoplasmic reticulum in higher vertebrate muscles.     

ULTRASTRUCTURAL ORGANIZATION OF OBLIQUELY STRIATED MUSCLE FIBERS IN ASCARIS LUMBRICOIDES

The somatic musculature of the nematode, Ascaris, is currently thought to consist of smooth muscle fibers, which contain intracellular supporting fibrils arranged in a regular pattern. Electron microscopic examination shows that the muscle fibers are, in fact, comparable to the striated muscles of vertebrates in that they contain interdigitating arrays of thick and thin myofilaments which form H, A, and I bands. In the A bands each thick filament is surrounded by about 10 to 12 thin filaments. The earlier confusion about the classification of this muscle probably arose from the fact that in one longitudinal plane the myofilaments are markedly staggered and, as a result, the striations in that plane of section are not transverse but oblique, forming an angle of only about 6° with the filament axis. The apparent direction of the striations changes with the plane of the section and may vary all the way from radial to longitudinal. A three-dimensional model is proposed which accounts for the appearance of this muscle in various planes. Z lines as such are absent but are replaced by smaller, less orderly, counterpart "Z bundles" to which thin filaments attach. These bundles are closely associated with fibrillar dense bodies and with deep infoldings of the plasma membrane. The invaginations of the plasma membrane together with intracellular, flattened, membranous cisternae form dyads and triads. It is suggested that these complexes, which also occur at the cell surface, may constitute strategically located, low-impedance patches through which local currents are channeled selectively.     

STUDIES ON THE STRUCTURE OF MUSCLE : III. PHASE CONTRAST AND ELECTRON MICROSCOPY OF DIPTERAN FLIGHT MUSCLE

1. The flight muscles of blowflies are easily dispersed in appropriate media to form suspensions of myofibrils which are highly suitable for phase contrast observation of the band changes associated with ATP-induced contraction. 2. Fresh myofibrils show a simple band pattern in which the A substance is uniformly distributed throughout the sarcomere, while the pattern characteristic of glycerinated material is identical with that generally regarded as typical of relaxed vertebrate myofibrils (A, I, H, Z, and M bands present). 3. Unrestrained myofibrils of both fresh and glycerinated muscle shorten by not more than about 20 per cent on exposure to ATP. In both cases the A substance migrates during contraction and accumulates in dense bands in the Z region, while material also accumulates in the M region. It is proposed that these dense contraction bands be designated the C_z, and C_m bands respectively. In restrained myofibrils, the I band does not disappear, but the C_z and C_m bands still appear in the presence of ATP. 4. The birefringence of the myofibrils decreases somewhat during contraction, but the shift of A substance does not result in an increase of birefringence in the C_z and C_m bands. It seems therefore that the A substance, if it is oriented parallel with the fibre axis in the relaxed myofibril, must exist in a coiled or folded configuration in the C hands of contracted myofibrils. 5. The fine structure of the flight muscle has been determined from electron microscopic examination of ultrathin sections. The myofibrils are of roughly hexagonal cross-section and consist of a regular single hexagonal array of compound myofilaments the cores of which extend continuously throughout all bands of the sarcomere in all states of contraction or relaxation so far investigated. 6. Each myofilament is joined laterally with its six nearest neighbours by thin filamentous bridges which repeat at regular intervals along the fibre axis and are present in the A, I, and Z, but not in the H or M bands. When stained with PTA, the myofilaments display a compound structure. In the A band, a lightly staining medullary region about 40 A in diameter is surrounded by a densely staining cortex, the over-all diameter of the myofilament being about 120 A. This thick cortex is absent in the I and H bands, but a thinner cortex is often visible. 7. It is suggested that the basic structure is a longitudinally continuous framework of F actin filaments, which are linked periodically by the lateral bridges (possibly tropomyosin). The A substance is free under certain conditions to migrate to the Z bands to form the C_z bands. The material forming the C_m bands possibly represents another component of the A substance. The results do not clearly indicate whether myosin is confined to the A bands or distributed throughout the sarcomere.     

CARDIAC MUSCLE OF THE HORSESHOE CRAB, LIMULUS POLYPHEMUS : I. Ultrastructure

The fine structure of the cardiac muscle of the horseshoe crab, Limulus polyphemus, has been studied with respect to the organization of its contractile material, and the structure of its organelles and the cell junctions. Longitudinal sections show long sarcomeres (5.37 µ at L_max), wide A bands (2.7 µ), irregular Z lines, no M line, and no apparent H zone. Transverse sections through the S zone of the A band show that each thick filament is ca. 180 A in diameter, is circular in profile with a center of low density, and is surrounded by an orbit of 9–12 thin filaments, each 60 A in diameter. Thick filaments are confined to the A band: thin filaments originate at the Z band, extend through the I band, and pass into the A band between the thick filaments. The sarcolemmal surface area is increased significantly by intercellular clefts. Extending into the fiber from these clefts and from the sarcolemma, T tubules pass into the fiber at the A-I level. Each fibril is enveloped by a profuse membranous covering of sarcoplasmic reticulum (SR). Sacculations of the SR occur at the A-I boundary where they make diadic contact with longitudinal branches of the T system. These branches also extend toward the Z, enlarge at the Z line, and pass into the next sarcomere. Infrequently noted were intercalated discs possessing terminal insertion and desmosome modifications, but lacking close junctions (fasciae occludentes). These structural details are compared with those of mammalian cardiac and invertebrate muscles.     

THE MYOFILAMENT ARRANGEMENT IN THE FEMORAL MUSCLE OF THE COCKROACH, LEUCOPHAEA MADERAE FABRICIUS

The structure of the femoral muscle of the cockroach, Leucophaea maderae, was investigated by light and electron microscopy. The several hundred fibers of either the extensor or flexor muscle are 20 to 40 µ in diameter in transverse sections and are subdivided into closely packed myofibrils. In glutaraldehyde-fixed and epoxy resin-embedded material of stretched fibers, the A band is about 4.5 µ long, the thin filaments are about 2.3 µ in length, the H zone and I band vary with the amount of stretch, and the M band is absent. The transverse sections of the filaments reveal in the area of a single overlap of thick and thin filaments an array of 10 to 12 thin filaments encircling each thick filament; whereas, in the area of double overlap in which the thin filaments interdigitate from opposite ends of the A band, the thin filaments show a twofold increase in number. The thick filament is approximately 205 to 185 A in diameter along most of its length, but at about 0.2 µ from the end it tapers to a point. Furthermore, some well oriented, very thin transverse sections show these filaments to have electron-transparent cores. The diameter of the thin filament is about 70 A. Transverse sections exhibit the sarcolemma invaginating clearly at regular intervals into the lateral regions of the A band. Three distinct types of mitochondria are associated with the muscle: an oval, an elongate, and a type with three processes. It is evident, in this muscle, that the sliding filament hypothesis is valid, and that perhaps the function of the extra thin filaments is to increase the tensile strength of the fiber and to create additional reactive sites between the thick and thin filaments. These sites are probably required for the functioning of the long sarcomeres.     

A COMPARISON OF THE FINE STRUCTURES OF FROG SLOW AND TWITCH MUSCLE FIBRES

The organisation of the myofibrils and the sarcoplasmic reticulum in frog slow muscle fibres has been compared with that in twitch fibres. It has been found that the filaments have the same length in the two types of fibre, but that there are differences in their packing: (a) in contrast to the regular arrangement of the I filaments near the Z line in twitch fibres, those in slow fibres are irregularly packed right up to their insertion into the Z line; (b) the Z line itself shows no ordered structure in slow fibres; (c) the fine cross-links seen between the A filaments at the M line level in twitch fibres are not present in slow fibres. The sarcoplasmic reticulum in slow fibres consists of two separate networks of tubules. One set of tubules (diameter about 500 to 800 A) is oriented mainly in a longitudinal direction. The tubules of the other network (diameter about 300 A) are oriented either transversely at approximately Z line level or longitudinally, connecting the transverse tubules. Triads are very rarely found, occurring at only every 5th or 6th Z line of each fibril. The central element of these triads is continuous with the thin tubules. Slow fibres from muscles soaked in ferritin-containing solutions contain ferritin particles in the network of thin tubules, the rest of the sarcoplasm remaining free of ferritin.     

THE SARCOPLASMIC RETICULUM IN MUSCLE CELLS OF AMBLYSTOMA LARVAE

Electron microscopy of thin sections of muscle fibers in myotomes of Amblystoma larvae has revealed the presence of a complex, membrane-limited system of canaliculi and vesicles which form a lace-like reticulum around and among the myofibrils. This seems to correspond to the sarcoplasmic reticulum of the earlier light microscopists and the endoplasmic reticulum of other cell types. The elements constituting the reticulum are disposed in a pattern which bears a constant relation to the bands of the adjacent myofibrils and is therefore repeated in each sarcomere. At the H band the system is transversely continuous but not so at other levels. Longitudinally continuity is interrupted at the Z bands where large vesicles belonging to adjacent sarcomere segments of the system face off on opposite sides of the band. The opposing faces of these vesicles are flat and separated by a space of more or less constant width, in which are located small, finger-shaped vesicles. In view of these and other close structural relationships with the myofibrils it seems appropriate to assign to the system a role in the conduction of the excitatory impulse.     

SARCOPLASMIC RETICULUM OF AN UNUSUALLY FAST-ACTING CRUSTACEAN MUSCLE

The fast-acting, synchronous "remotor" muscle of the lobster second antenna was examined by light and electron microscopy and was found to have a more profuse sarcoplasmic reticulum (SR) than any other muscle known. Myofibrils are widely separated from one another and occupy only about one-fourth of the volume of the muscle; most of the remaining volume is taken up by the SR, which resembles the smooth-surfaced reticulum of steroid-secreting cells. Dense granules (0.03–0.1 µ in diameter) are scattered through the reticulum. T-tubules penetrate into the fibers and form dyads along the A bands of myofibrils; however, ferritin-labeling experiments show that the volume of the T-system is very small compared with that of the SR. Myofibrils are ~0.5 µ x 1.0 µ in cross section and consist of thick filaments, which appear tubular except at the M region, and thin filaments, which are situated midway between neighboring thick filaments. The ratio of thin to thick filaments is 3:1. The extreme development of the SR in this muscle is discussed in relation to the exceedingly short duration of the contraction-relaxation cycle.     

A STRAND OF CARDIAC MUSCLE : Its Ultrastructure and the Electrophysiological Implications of Its Geometry

The structure of a small strand of rabbit heart muscle fibers (trabecula carnea), 30–80 µ in diameter, has been examined with light and electron microscopy. By establishing a correlation between the appearance of regions of close fiber contact in light and electron microscopy, the extent and distribution of regions of close apposition of fibers has been evaluated in approximately 200 µ length of a strand. The distribution of possible regions of resistive coupling between fibers has been approximated by a model system of cables. The theoretical linear electrical properties of such a system have been analyzed and the implications of the results of this analysis are discussed. Since this preparation is to be used for correlated studies of the electrical, mechanical, and cytochemical properties of cardiac muscle, a comprehensive study of the morphology of this preparation has been made. The muscle fibers in it are distinguished from those of the rabbit papillary muscle, in that they have no triads and have a kind of mitochondrion not found in papillary muscle. No evidence of a transverse tubular system was found, but junctions of cisternae of the sarcoplasmic reticulum and the sarcolemma, peripheral couplings, were present. The electrophysiological implications of the absence of transverse tubules are discussed. The cisternae of the couplings showed periodic tubular extensions toward the sarcolemma. A regularly spaced array of Z line-like material was observed, suggesting a possible mechanism for sarcomere growth.     

STRUCTURE OF LIMULUS STRIATED MUSCLE : The Contractile Apparatus at Various Sarcomere Lengths

The musculature of the telson of Limulus polyphemus L. consists of three dorsal muscles: the medial and lateral telson levators and the telson abductor, and one large ventral muscle; the telson depressor, which has three major divisions: the dorsal, medioventral, and lateroventral heads. The telson muscles are composed of one type of striated muscle fiber, which has irregularly shaped myofibrils. The sarcomeres are long, with discrete A and I and discontinuous Z bands. M lines are not present. H zones can be identified easily, only in thick (1.0 µm) longitudinal sections or thin cross sections. In lengthened fibers, the Z bands are irregular and the A bands appear very long due to misalignment of constituent thick filaments. As the sarcomeres shorten, the Z lines straighten somewhat and the thick filaments become more aligned within the A band, leading to apparent decrease in A band length. Further A band shortening, seen at sarcomere lengths below 7.4 µm may be a function of conformational changes of the thick filaments, possibly brought about by alterations in the ordering of their paramyosin cores.