Sarcolemmal Invaginations Research Articles

A fine-structural study of the termination of intrafusal muscle fibres in the Chinese hamster.

The termination of intrafusal muscle fibres was studied in muscle spindles of the tenuissimus muscle of the Chinese hamster. Most nuclear chain fibres terminated within the spindle capsule. The nuclear bag- and nuclear chain fibres that extended beyond the limit of the spindle capsule terminated in the endomysial connective tissue of the adjacent extrafusal fibres. Three types of intracapsular terminations were found: (1) termination on the neighbouring nuclear bag fibre, (2) termination in the periaxial space without any obvious attachment site, or (3) termination on the spindle capsule. The intrafusal fibres were tapered and longitudinal sarcolemmal invaginations were poorly developed or were completely lacking. The fibre ends were characterized by their folded basal lamina, which appeared to be mechanically reinforced by associated collagen fibrils and elastic fibres.

Ultrastructure and differentiation of ascidian muscle. II. Differentiation of the caudal muscle cells in the larva of Diplosoma macdonaldi.

The larval muscle cells of Diplosoma macdonaldi contain subcortical and medullary myofibrils which are invested by fenestrated sheets of the sarcoplasmic reticulum. Cisternae of the sarcoplasmic reticulum are coupled with tubular invaginations of the sarcolemma. To appreciate better such uncommon features of cellular organization, six embryonic stages were selected for an ultrastructural study of myogenesis. The proliferative, synthetic, and elaborative phases of myogenesis were represented by embryos ranging from neurulae to prehatching larvae. The contractile apparatus originates during the synthetic phase of myogenesis, when thick and thin myofilaments appear in the cortical sarcoplasm at the epidermal and notochordal poles of the cell. The myofilaments promptly aggregate into unstriated fascicles, and the fascicles unite in series to establish the rudimentary myofibrils. All major sarcomeric bands, except the Z-lines, are evident along the myofibrils. Cisternae of the sarcoplasmic reticulum form peripheral couplings with the overlying sarcolemma, and they also form interior couplings with sarcolemmal invaginations from the ends of the cell. The interior couplings localize over the I-bands of the myofibrils. In the elaborative phase of myogenesis, mitochondria invade the cortical sarcoplasm, and the contractile apparatus passively shifts to the subcortex and medulla of the cell. Relocation of the myofibrils coincides with the disappearance of all peripheral couplings. Cisternae of the sarcoplasmic reticulum anastomose around the myofibrils, creating the fenestrated sheets that extend between sarcomeres. As Z-lines begin to bisect the I-bands, the perifibrillar cisternae become confluent with the cisternae in the precocious interior couplings.

Effects of Ca2+ on the contractility and Sr2+ accumulation in the quail oviductal smooth muscle.

There occur strong spontaneous contractions in the avian oviduct, which are considered to effect the locus of the ovum during ovum transport. Previously it has been shown that stretch regulates these contractions by means of some unknown mechanisms in this tissue. When a transversal strip, cut from the proximal magnum of the oviduct, was first kept at the length (LO = length in situ) and then stretched to the length 200% LO the total integrated area of contractions increased 3.67 +/- 0.63 (6) times greater (p less than 0.01) than the contractility at the length LO. Ca2+ concentration of 5 mM increased the total integrated area of the contractions 1.42 +/- 0.22 (7) greater than the control while the increase at 8 mM was only 1.17 +/- 0.30 (7). 0.5 mM Ca2+ decreased the contractility to 0.72 +/- 0.15 (7). In a 50 microM EGTA containing solution, the contractility decreased in a few minutes. Sr2+, used as an indirect tracer of Ca2+, accumulated both in control and stretched strips in the sarcoplasmic reticulum, the mitochondria, and the sarcolemma. The electron-dense deposits were, however, most clearly seen in the sarcolemma. Compared to the Sr2+ accumulating sites in erythrocytes, deposits were on the outer side of the cell membrane, thus suggesting the glycocalyx as an important site for Ca2+ exchange in this smooth muscle. The sarcolemmal invaginations, caveolae, accumulated Sr2+ similarly to other parts of the sarcolemma. During ovum transport, stretch changes the Ca2+ accessibility to muscle cells, and at structural level, this takes place in the sarcolemma.

Myofiber damage accompanying intramuscular parasitism by Sarcocystis muris

Myofiber degeneration which results from Sarcocystis infection exhibits a number of fine structural features suggestive of other myopathies and several welldefined fine structural features not characteristic of other myopathies. Some of these fine structural features are similar to those observed in intramuscular infections of Trichinella spiralis, another muscle parasite. Major alterations of the myofibrillar contractile apparatus occur at the periphery of the membrane bound parasitophorous vacuole which include splitting and fragmentation of the myofibrils at the longitudinal ends of the parasitophorous vacuole and Z line dissolution at the radial periphery. Membranous structural elements including mitochondria, sarcoplasmic reticulum and T system components become disarrayed as the myofibrils degenerate. Some minor hypertrophy of the sarcoplasmic reticulum occurs in conjunction with initial fragmentation of the myofibrils but no major dilation or hypertrophy has been observed. There is a distinctive membranous organization of the interface of the parasitophorous vacuole. The presence of pycnotic and fragmenting nuclei, sarcolemmal invaginations with accompanying fibrous connective tissue invasion and large areas of undifferentiated cytoplasm suggest the ultimate necrosis and destruction of infected myofibers. The similarity between morphological features of myofibrillar degeneration accompanying intramuscular Sarcocystis muris infections and those associated with a variety of myopathies resulting from other causes suggests that a common mechanism of muscle response to damage might result in the observed structural degeneration.

A quantitative estimation of components in crayfish muscle fibres by stereological methods.

The volume and surface densities of structural components in isolated long-sarcomere fibres of crayfish (Astacus fluviatilis) were estimated by stereological methods. The relative volumes were as follows (in % of the fibre volume): sarcolemmal invaginations (SI) 0.74; T-system 1.1; whole sarcoplasmic reticulum (SR) 4.7; junctional SR cisternae 0.41; mitochondria 1.1; nuclei 0.57; myofibrils 81.6; peripheral basal lamina 1.1; junctional gap 0.043; lysosomes 0.029. The values for surface densities were (in microns2/100 microns3): peripheral sarcolemma 1.27; SI 7.21; T-system 40.2; whole SR 146.4; junctional SR cisternae 14.88; junctional area of SR cisternae 7.05; mitochondria 8.98; nuclei 0.75; myofibrils 163. The invaginated membranes (SI and T-system) made up 97.4% of the total area of surface membranes and increased the effective area of peripheral sarcolemma 39 times. Using the values of the total electrical capacity from previous investigations on Astacus fibres, the specific capacity of surface membranes was calculated as 1.08 microF/cm2.

The plasma membrane of leading pacemaker cells in the rabbit sinus node. A qualitative and quantitative ultrastructural analysis.

We studied the fine structure of the plasma membrane of electrophysiologically identified leading pacemaker cells from the rabbit sinus node, using both ultrathin sections of fixed tissue and replicas of freeze-cleaved material. We found that differences exist between sinus node and working myocardial membranes, but these are only quantitative. The caveolae or sarcolemmal invaginations are present in very large numbers; they increase the surface area of the plasma membrane by about 100%. The small macular nexuses that are present represent 0.2% of the membrane surface area. Nexuses are therefore about 10 times less numerous in leading sinus node cells than in working myocardium cells. A single equivalent electrical representation of the sinus node shows, nevertheless, that an appreciable electrical coupling may be expected.

Invaginated membrane in crustacean tonic muscle fibers: estimates of membrane capacitance.

Surface and volume densities of the invaginated membrane components of tonic muscle fibers of crayfish (Orconectes rusticus) were determined by stereological methods. Surface densities of the sarcolemma, sarcolemmal invaginations, T tubules and Z tubules were 2.0, 5.9, 26.3, and 3.3 micron2/100 micron3 of fiber, respectively, and volume densities of the sarcolemmal invaginations, T tubules, and Z tubules were 0.28, 0.29, and 0.08 micron3/100 micron3. The values obtained for surface density show that 95% of the membrane exposed to the extracellular environment is composed of invaginated membrane. Specific membrane capacitance, determined from square-wave pulse analysis and referred to the apparent area of surface sarcolemma, was 30.3 muF/cm2. However, after correction for the extent of invaginated membrane, the specific membrane capacitance was 1.6 muF/cm2.

Ultrastructure of A cardiac rhadomyoma

Electron microscopic study of a cardiac rhabdomyoma removed at open heart operation revealed large rounded or polygonal cells that contained large amounts of monoparticulate glycogen. Myofibrils in these cells were few and located either subjacent to the plasma membranes or radiating from central areas. Leptofibrils and masses of anomalous Z band material were abundant. Shallow tubular sarcolemmal invaginations and elements of free and junctional sarcoplasmic reticulum were associated with the myofibrils. Desmosomes and nexuses were evident in intercellular junctions, which were extensive and randomly distributed throughout the cell surfaces. It is concluded that cardiac rhabdomyomas probably are hamartomas.

Ultrastructure of cardiac muscle of Trochosa terricola Thor., Pardosa amentata Clerck, P. pullata Clerck, and Pisaura mirabilis Clerck (Araneae: Lycosidae, Pisauridae).

The ultrastructure of the cardiac muscle in some araneae has been investigated. The sarcolemma invaginates at the Z band and may extend into the cells through several myofibrils. Numerous T-tubules are given off both from sarcolemmal invaginations and also directly from the cell surface. In passing through the Z band, the luminal diameter of T-tubules greatly increases. Dyadic and a few triadic couplings are found mainly at the A-I level. Peripheral couplings were not seen.

The Structure of the Terminal Filament, the Ovariole Sheath and the Oviduct Musculature of the Colorado Beetle (Leptinotarsa Decemlineata Say, 1824)

In the Colorado Beetle, the terminal filament cells resemble the epidermal cells which connect muscle cells with the cuticle, the so-called tendon cells. They contain many microtubules which have projections interconnecting them. The microtubules are also connected by these projections to desmosomes. The tunica propria is a basement membrane-like layer of low elasticity. It separates the terminal filament from the germarium. The outer ovariole sheath is a muscle sheath which is continuous with the muscle sheath around the oviducts. The whole muscle sheath consists of a monolayer network. Sarcolemmal invaginations at the I band and at the Z line probably form part of one T system. The cuticle of the common oviduct has scales with long caudally directed spines. The rostral attachment of the muscle sheath corresponds to the existing descriptions of tendon cells. The attachment of the muscle sheath to the cuticle of the common oviduct is different.

Electron microscopic investigations on leptomeric fibrils and leptomeric complexes in the hen and pigeon heart

Leptomeric fibrils are studied electron microscopically in Purkinje cells of the hen and pigeon heart and in ordinary myocardial cells of the hen. The leptomeric fibrils lie chiefly subjacent to the sarcolemma and are often related to sarcolemmal invaginations. They are frequently attached to normomeric myofibrils. Leptomeric fibrils are found preferentially in connective tissue embedded Purkinje cells. In the Purkinje cells of the hen heart a special leptomeric structure—the leptomeric complex—is observed. The leptomeric complexes are often crossed by normomeric myofibrils. Lysosomes and lamellated bodies occur in the neighbourhood and in the interior of leptomeric complexes. Possible functions of the leptomeric fibrils and leptomeric complexes as suggested in the literature are reviewed and discussed in the light of the present findings.

Localization and properties of the cholinesterase in crustacean muscle.

Cholinesterase (ChE) activity is present in crustacean muscle extracts. However, since acetylcholine (ACh) is not a neuromuscular transmitter in these animals, the role and exact localization of ChE was unknown. The histochemical localization of the enzyme was studied in whole muscle and in the sarcoplasmic reticulum fraction of the extract, 50-microm frozen sections of glutaraldehyde-fixed crayfish tail flexor muscle were incubated with acetylthiocholine (ATC) as substrate, and examined under the electron microscope. After some modifications in published techniques, dense deposits were found associated with the sarcolemma, sarcolemmal invaginations, and transverse tubules. No deposits were found in 10(-4) M eserine, or if butyrylthiocholine (BTC) was substituted for ATC. The vesicles in the sarcoplasmic reticulum fraction which demonstrate the activity must represent minced bits of these membranes. Using a spectrophotometric method, the kinetics of the crustacean muscle enzyme was compared to the acetylcholinesterase (AChE) on mammalian red blood cells and in the lobster ventral nerve cord. Surprisingly, and contrary to previous reports, the crustacean muscle enzyme did not demonstrate substrate inhibition. While a number of similarities to AChE were found, this lack of substrate inhibition makes questionable an unequivocal similarity with classical AChE.

Ultrastructural features of cardiac muscle cells in a tarantula spider.

AbstractThe 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.

The effect of potassium and chloride ions on the volume and membrane potential of single barnacle muscle cells.

1. In single barnacle skeletal muscle cells cell diameter has been measured as a function of external osmolality, and cell diameter and membrane potential have been measured during changes of external K and Cl concentrations ([K](o) and [Cl](o)) like those described in frog muscle by Hodgkin & Horowicz (1959). The diameter was monitored microscopically with a precision of 0.2-0.4% (S.D.). [K](o) was varied from 1 to 18 mM, a range of concentrations which does not cause contracture.2. At pH 8.0 the Cl permeability was so low that net KCl and water movements were absent. Such net movements were present at pH 4.5, corresponding to a change in the ratio (Cl conductance/K conductance) from approximately 1/12 at pH 8.0 to 1/2 at pH 4.5.3. Characteristically long time constants were observed for membrane potential responses to a change in [K](o) and/or [Cl](o), even at constant [K](o).[Cl](o). This phenomenon is attributed to a delayed equilibration by diffusion within the system of sarcolemmal invaginations and T-tubules. The delay in the response was increased by introducing polyvinylpyrrolidone (PVP) into this system, presumably because PVP raises intratubular viscosity.4. At pH 4.5 anomalous rectification for net movements of K was demonstrated by measurements of cell diameter and of membrane potential.

Ultrastructure of the neurogenic heart of Limulus polyphemus.

The ultrastructure of Limulus cardiac muscle was examined. The hearts were fixed in situ by perfusion with isotonic glutaraldehyde solution while in relaxed, contracted, or stretched states. The sarcomeres are relatively long, varying in length from about 2.5 to 6.6 μ. The average A-band length is 2.46 μ. M lines are absent, and H zones are poorly distinguished. Thick and thin filament diameters average about 200 A and 50 A, respectively; each thick filament is surrounded by 8–12 thin ones. Superficial invaginations of the sarcolemma occur, making contact with the Z lines of the outermost myofibrils. There is an extensive sarcoplasmic reticulum and transverse (T) tubules. Some T tubules run longitudinally and some open into deep sarcolemmal invaginations which extend into the fiber interior. The T tubules swell markedly in hypertonic solution. Single neurons and small bundles of neurons are observed in close apposition with myocardial cells. Intercalated disks are found in Limulus heart at regions of contact between contiguous myocardial cells lying end to end; semitight or gap junctions are essentially absent. Prominent differences in sarcomere lengths sometimes occur across the disk, thus indicating that the disks demarcate cells functionally. Hence, in addition to direct motoneuron activation, there may be some transfer of excitation across the intercalated disks in accord with our previous finding that propagating, overshooting action potentials can be induced in this heart.

Ultrastructure of stick insect and locust skeletal muscle in relation to excitation-contraction coupling

The subcellular structures involved in excitation-contraction coupling have been examined in locust metathoracic extensor tibialis muscle and stick insect prothoracic flexor tibialis muscle. In both muscles the sarcomeres are of similar resting length (4·5 μm) and the fibres are invaded by sarcolemmal invaginations and an extensive transverse tubular system is present. Locust muscle has a much more extensive sarcoplasmic reticulum and more frequent dyads per unit area than stick insect muscle. The myofilament array is fairly similar in both muscles; in locust, each myosin filament is surrounded by nine to eleven actin filaments, while in stick insect twelve actin filaments usually surround each myosin filament. The striking differences in speed of contraction and relaxation between these muscles can almost certainly be ascribed to differences in development of the sarcoplasmic reticulum and the dyads, which are areas of active calcium uptake and release during the excitation-contration coupling cycle.

CORRELATED MORPHOLOGICAL AND PHYSIOLOGICAL STUDIES ON ISOLATED SINGLE MUSCLE FIBERS

Living muscle fibers of crayfish become dark during efflux of Cl(-). This change in appearance is correlated with occurrence of vacuolation in the fixed fibers. The vacuoles begin at and are mainly confined to the terminals of the transverse tubular system (TTS) which are in diadic contact with the sarcoplasmic reticulum (SR). In electron micrographs swellings more than 1 micro in diameter may be seen connected to the sarcolemma or sarcolemmal invaginations by relatively unswollen tubules about 300-500 A wide. Darkening of the living fibers can be reversed by causing an influx of Cl(-). Vacuoles are then absent in the fixed preparations. These findings accord with the conclusion that the membrane of the TTS is anion permselective. Localization of the selectivity to the membrane of the terminals of the TTS strengthens the hypothesis that a channeling of current flow is responsible for initiation of excitation-contraction coupling. During the swelling, and upon its reversal, the area of the membrane of the terminals must change reversibly by about two to four orders of magnitude. The absence of changes in the dimensions of the unit membrane indicates that the expansion of the membrane and its subsequent shrinkage involve reversible incorporation of cytoplasmic material into the membrane phase.

Structure and Function of the Transverse Tubular System in Crustacean Muscle Fibers

The structure and function of the transverse tubular system (TTS) in two types of crustacean muscle fibers are examined. Giant fibers from the barnacle, Balanus nubilus , which are gradedly contracting, are compared with allor-none twitch fibers from the crab, Carcinus maenas . Both fiber types were found to have deep sarcolemmal invaginations which serve both to increase the fiber surface area and to kfeep the length of the tubules short enough for electrotonic propagation.The ultrastructure of the tubular system in both types of fiber is compared.The system is better developed in Carcinus than in Balanus , but the slow Balanus fibers do have a relatively well developed TTS and sarcoplasmic reticulum in contrast to slow vertebrate fibers. The apparent high, membrane-capacitance values of crustacean fibers are the result of investigators not taking into consideration the large increase in surface area due to the sarcolemmal infoldings.The tubular membranes in Carcinus fibers were found to be permselective to chloride ions, and could be made to swell (as confirmed by electron microscopy) by establishing an outward gradient for chloride across them. The capacitance of the tubular membrane relative to the plasma membrane was found to increase when the tubuleswere swollen. The implication of a fiber having two spatially separated, differentially permeable membranes on excitation-contraction coupling is discussed.

Pharmacology and fine structure of peripheral muscle innervation in the cockroach Periplaneta americana.

Drugs affecting a cholinergic system (i.e. acetylcholine, nicotine, paraoxon, eserine, atropine, succinylcholine) and drugs having no cholinergic effects (i.e. norepinephrine, harmine, gamma-aminobutyric acid) were perfused into the metathoracic leg of the cockroach Periplaneta americana , and the physiological results were observed. Nicotine produced a reversible tetanic response, and succinylcholine decreased the amplitude of the contraction. All the other drugs had no effect. Electron micrographs of the peripheral neuromuscular system revealed a typical insect junction: no junctional folds, synaptic vesicles, and a synaptic cleft of 200 Å. The sarcoplasmic reticulum of the cockroach muscle is extensive and consists of a series of tubules and cisternae often seen as a three-dimensional collar around the myofibrils. Sarcolemmal invaginations formed dyads with the sarcoplasmic reticulum supposedly providing the pathway for rapid transmission of excitation to the interior of the muscle fibre. The morphological evidence indicates that the junction is chemically mediated, but the physiological data demonstrate that the mediator is not acetylcholine.

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-micro 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 micro 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.