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.     

Ultrastructurally different muscle cell types in Eisenia foetida (Annelida,Oligochaeta)

Muscles in the body wall, intestinal wall, and contractile hemolymphatic vessels (pseudohearts) of an oligochaete anelid (Eisenia foetida) were studied by electron microscopy. The muscle cells in all locations, except for the outer layer of the pseudohearts, are variants of obliquely striated muscle cells. Cells comprising the circular layer of the body wall possess single, peripherally located myofibrils that occupy most of the cytoplasm and surround other cytoplasmic organelles. The nuclei of the cells lie peripherally to the myofibrils. The sarcomeres consist of thin and thick myofilaments that are arranged in parallel arrays. In one plane of view, the filaments appear to be oriented obliquely to Z bands. Thin myofilaments measure 5–6 nm in diameter. Thick myofilaments are fusiform in shape and their width decreases from their centers (40–45 nm) to their tips (23–25 nm). The thin/thick filament ratio in the A bands is 10. The Z bands consist of Z bars alternating with tubules of the sarcoplasmic reticulum. Subsarcolemmal electron-dense plaques are found frequently. The cells forming the longitudinal layer of the body wall musculature are smaller than the cells in the circular layer and their thick filaments are smaller (31–33 nm centrally and 21–23 nm at the tips). Subsarcolemmal plaques are less numerous. The cells forming the heart wall inner layer, the large hemolymphatic vessels, and the intestinal wall are characterized by their large thick myofilaments (50–52 nm centrally and 27–28 nm at the tips) and abundance of mitochondria. The cells forming the outer muscular layer of the pseudohearts are smooth muscle cells. These cells are richer in thick filaments than vertebrate smooth muscle cells. They differ from obliquely striated muscle cells by possessing irregularly distributed electron-dense bodies for filament anchorage rather than sarcomeres and Z bands and by displaying tubules of smooth endoplasmic reticulum among the bundles of myofilaments. © 1995 Wiley-Liss, Inc.     

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.     

Observations on the ultrastructure of developing myocardium of rat embryos

Timed pregnancies were obtained in Sprague-Dawley rats and early ultrastructural differentiation of myocardium of embryos of 10, 11, 12, 13, and 14 days was investigated and compared with that of newborn. Ten-day myocardium is characterized by loosely packed cells; the cytoplasm is typified by a dearth of organelles. Both thick (myosin) and thin (actin) filaments become identifiable for the first time in the 10-day myocardium where the heart is pulsating but circulation is not established. These filaments are not visible in the embryos of 9-day-old myocardium. The formation of these filaments is observed to continue throughout the period covered in this investigation. Concomitant with the appearance of the myofilaments is the synthesis of Z band material. By the eleventh day of gestation and during the subsequent days there is a rapid proliferation and differentiation of most of the organelles. The myofilaments become organized into fully formed striated fibrils. Intercalated discs appear as. small wavy lines on the eleventh day and become plicated in later stages and serve as cell boundaries and points of attachment for myofilaments and fibrils. There is a perceptible change in the number and morphology of mitochondria from the tenth to eleventh day and later stages of development when the heart becomes functional. Similarly, there is a rapid proliferation and differentiation of granular endoplasmic reticulum and Golgi bodies. Large quantities of free ribosomes are dispersed in the cytoplasm of 10-day myocardium; however, in later stages there is a progressive reduction in the distribution of these particles. An intimate association of ribosomes and polysomes with the developing myofibrils is discernible. The T -system and sarcoplasmic reticulum begin to appear in II-day myocardium. The embryonic myocardium displays intense mitotic activity throughout its development and a unique feature of embryonic myocardial cells is the simultaneous occurrence of myofilament synthesis and mitotic activity within the same cells.     

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.     

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.     

Myotube-like structures in cultures of rat liver cells

Branching and budding myotube-like structures developed in primary cultures of rat liver cells and in the JH1 cell line derived from them. Elongated uninuclear cells aligned in chains and fused into multinuclear tubes of varying length and thickness. The tubes contained thick and thin filaments running in all directions. The filaments were occasionally linked with M lines and formed incomplete hexagonal patterns resembling those of skeletal muscle myofilaments, but a regular arrangement of the filaments and organelles was lacking. Cross-striation and contractions were never observed. Both uninuclear cells and multinuclear tubes contained numerous lysosomes, myelin figures and lamellated bodies as well as electronlucent or content-filled vacuoles and cisternae of variable size, sometimes reminding the sarcoplasmic reticulum in early stages of its development. Endocytotic caveolae and vesicles were present in all elements. These features together with interdigitated cell processes and specialized cell contacts suggested a possible relationship of the cells to the reticuloendothelial system.     

A COMPARATIVE STUDY ON THE FINE STRUCTURE OF THE BASALAR MUSCLE OF THE WING AND THE TIBIAL EXTENSOR MUSCLE OF THE LEG OF THE LEPIDOPTERAN ACHALARUS LYCIADES

Basalar and tibial extensor muscle fibers of Achalarus lyciades were examined with light and electron microscopes. Basalar muscle fibers are 100–150 µ in diameter. T-system membranes and sarcoplasmic reticulum make triadic contacts midway between Z lines and the middle of each sarcomere. The sarcoplasmic reticulum is characterized by a transverse element situated among myofilaments halfway between Z lines in every sarcomere. The morphology of Z lines, hexagonal packing of thin and thick myofilaments, and thin/thick myofilament ratios are similar to those of fast-acting insect muscles. Tibial extensor muscle fibers are 50–100 µ in diameter. Except for a lack of the transverse element, the T system and sarcoplasmic reticulum are similar to those of basalar muscle. Wavy Z lines, lack of a hexagonal packing of myofilaments, and larger thin/thick myofilament ratios are similar to those of other postural muscles of insects. The morphology of basalar and tibial extensor muscle is compared to that of other insect muscle with known functions, and reference is made to the possible contribution of the transverse element of sarcoplasmic reticulum in basalar flight muscle to speed and synchrony in this muscle.     

The fine structure of muscle in a marine turbellarian

Summary The fine structure of the myofibers of Notoplana acticola as studied by electron microscopy indicates that they are composed of thick myofilaments about 200 Å wide with tapering ends and thin myofilaments about 50 Å wide, arranged alongside each other parallel to the long axis of the cell. There is no orderly transverse arrangement of filaments; instead they appear staggered in the fiber. In cross sections 6 to 10 thin filaments form an orbit around one thick filament with possible cross-linkage between the two types of filaments.Dense bodies are associated with the sarcolemma and with the sarcoplasmic reticulum, and appear to serve as attachments for the thin filaments. Dense bodies are compared to elements forming a fragmented Z-disc.Mitochondria, situated in the periphery or the center of fibers, are associated with granules interpreted as glycogen.The sarcoplasmic reticulum consists of: sacs or cisternae in close proximity to the sarcolemma, longitudinal tubular elements between and parallel to the myofilaments, and a tubular network around the filaments. There is no well-defined sarcolemmal-derived transverse tubule system as described in striated muscles. It is hypothesized that in these muscles, the functional equivalent of the T system may be the area of sarcolemma in contact with the cisternae of the sarcoplasmic reticulum.This work was supported by Grant No. GM 10292 from the U. S. Public Health Service to Professor Richard M. Eakin, Department of Zoology at the University of California, Berkeley, USA, where this investigation was conducted during the author's sabbatical leave of absence from the University of Illinois.I wish to thank Professor Eakin for valuable discussions and for his kind hospitality in extending the facilities of his laboratory and the use of the electron microscope to me, and the John Simon Guggenheim Memorial Foundation for the Fellowship which I held during 1964–65.     

Ultrastructure and differentiation of the larval esophageal muscle cells of the starfish Pisaster ochraceus

The muscle cells that cause constriction of the starfish larval esophagus (esophageal muscle cells) are one of the first cell types to express their differentiated morphological characteristics during development. Ultrastructurally these muscle cells resemble vertebrate and invertebrate smooth muscles. They contain a nucleus, a Golgi apparatus, contractile myofilaments, hemidesmosome-like structures, and what appears to be a simple sarcoplasmic reticulum. In asteroid embryos, this muscle layer originates during mouth formation when mesenchyme cells migrate from the tips of the coeloms to the esophagus. Once there, they elongate, forming processes. Over the next few days, the processes become filled with arrays of longitudinally arranged thick and thin myofilaments and thin sacs of smooth endoplasmic reticulum. The latter appear between the bundles of contractile filaments and the cell membranes. Contractile activity begins at approximately this time. The cisternae may represent a sarcoplasmic reticulum that is required for contraction. The majority of the esophageal muscle cell processes extend around the circumference of the developing esophagus, but occasional cells may be oriented in other directions. The latter cells are always farther away from the basal lamina and probably have little or no contact with it. Contact with basal lamina may serve to direct the migration of the cells and the orientation of the processes. J. Morphol. 237:1–18, 1998. © 1998 Wiley-Liss, Inc.     

THE SARCOPLASMIC RETICULUM AND ITS ASSOCIATION WITH THE T SYSTEM IN AN INSECT

The fine structure of the sarcoplasmic reticulum and the transverse tubular system of the femoral muscle of the cockroach, Leucophaea maderae, was studied after prefixation in glutaraldehyde, postfixation in osmium tetroxide, and embedding in Epon. The sarcoplasmic reticulum in this muscle reveals features not previously reported. The sarcoplasmic reticulum is abundant, consisting mainly of a fenestrated envelope which surrounds each myofibril at all levels in the sarcomere. This sarcoplasmic reticulum envelope is continuous transversally as well as longitudinally along the myofibrils. Dyadic junctions are formed by a single T system element which contacts the unfenestrated sarcoplasmic reticulum of adjacent myofibrils in an alternating manner at the ends of the A band. At the dyads, regularly spaced thickenings of the sarcoplasmic reticulum membranes bordering the dyadic spaces are noted. These thickenings, however, do not contact the T tubule membrane. Typical dyadic contacts also are seen between the cell surface membrane and sarcoplasmic reticulum. Z line-like material is seen in contact with the membranes of the cell surface and longitudinal branches of the T systems.     

An ultrastructural investigation of the striated subumbrellar musculature of the anthomedusan,Pennaria tiarella

The presence of striated subumbrellar musculature in hydromedusae can be related to the development in hydrozoans of a free-swimming life stage. The detailed ultrastructure of the striated subumbrellar musculature of the anthomedusan, Pennaria tiarella is presented. The striated musculature of Pennaria resembles vertebrate striated muscle in filament arrangement (L2 lattice pattern) and M line structure. The striation pattern, out-of-register myofilaments, filament structure as determined by rotational symmetry, Z line structure, types of intercellular junctions, and sarcoplasmic reticulum are more similar to structures found in other invertebrate striated muscle.     

Cytological differentiation of human fetal skeletal muscle.

The ultrastructural differentiation of several different muscles was investigated in human fetuses ranging in age from 13 weeks to neonatal. At approximately 16 weeks of gestation cell cluster containing both myotubes and satellite cells lie enclosed by a newly formed basal lamina and show evidence of fusion. The development of organelles is evident in myoblasts, proceeds as the cells transform into myofibers, and continues in the neonate. Filament synthesis occurs primarily in the cell periphery where thin filaments appear to align themselves in relations to parallel arrays of ribosome-studded thick filaments: Z line formation follows the appearance of thin filaments. Intermediate filaments, approximately 10-12 nm thick, were also consistently observed in perinuclear regions and distal to filament assembly. Although sarcoplasmic reticulum (SR) development is closely related to fibril formation, connections between Z lines and SR are not consistent, thus supporting the conclusion that SR does not evoke the formation of the Z line. Bristlecoated vesicles appear to be the precursors of elements of the SR, possibly the lateral sacs. Development of the transverse tubules, as invaginations of the sarcolemma, is closely associated with the formation of lateral sacs since the latter occur along the sarcolemma as soon as transverse tubules appear. Cytological differentiation is similar, though not identical, in several different muscles. During the last trimester muscle fibers show some evidence of diversity mainly of variation in Z line width. In gerneral the results suggest that the sequence and stages of human myogenesis are similar to those of other species.     

The pedal muscle of the land snail Megalobulimus oblongus (Gastropoda, Pulmonata): an ultrastructure approach

M. Cristina Faccioni-Heuser, Denise M. Zancan, Christiane Q. Lopes and Matilde Achaval. 1999. The pedal muscle of the land snail Megalobulimus oblongus (Gastropoda, Pulmonata): an ultrastructure approach. — Acta Zoologica (Stockholm) 80: 325–337
The ultrastructure of the pedal muscle of the Megalobulimus oblongus is described. This muscle consists of transverse, longitudinal and oblique bundles ensheathed in collagenous tissue. Each muscle cell is also ensheathed by collagen. The smooth muscle cells contain thin and thick filaments; the thin filaments are attached to dense bodies. These cells contain a simple system of sarcoplasmic reticulum, subsarcolemmal caveolae and mitochondria with dense granules in the matrix, and glycogen. Three types of muscle cells were identified. Type A cells exhibited densely packed myofilaments, abundant glycogen rosettes, numerous mitochondria and sarcoplasmic reticulum profiles. Type B cells exhibited scanty glycogen and mitochondria, few cisternae of sarcoplasmic reticulum and large intermyofibrillar spaces. Type C cells exhibited intermediate characteristics between type A and type B cells. Neither nexus nor desmosomes were observed between the muscle cell membranes. The muscle contains well developed connective tissue and blood vessels. These structures and the distribution of muscle cells are probably involved in the muscular-hydrostat system. The muscle is richly innervated, having neuromuscular junctions with clear and electron-dense synaptic vesicles. The clear vesicles probably contain acetylcholine because the axons to which they are connected arise from acetylcholinesterase positive neurones of the pedal ganglion. The other vesicles may secrete monoamines such as serotonin and/or neuropeptides such as substance P.     

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.     

Carbocyanine dyes stain the sarcoplasmic reticulum of beating heart cells

In search of a fluorescent dye suitable for monitoring membrane potentials of beating heart cells, we noticed that the carbocyanine dyes, CC₅ and CC₆, show a unique pattern of intracellular distribution in vital and glutaraldehyde-fixed cardiomyoblasts. This distribution is clearly different from that observed in fibroblasts. In heart cells, it parallels the localization of actin-myosin containing myofilaments as visualized by fluorescent antibody staining but it does not correspond to the localization of actin filaments or the microtubules. In fibroblasts these dyes stain only fine filaments and granules in the perinuclear space which correspond to the endoplasmic reticulum. This observation is evidence in support of the hypothesis that carbocyanine dyes accumulate selectively in the sarcoplasmic reticulum. It indicates that certain carbocyanine dyes may be useful tools to differentiate between muscle cells and connective tissue cells in cell cultures.     

The fine structure of differentiating muscle in the salamander tail

Summary Thin methacrylate sections of developing tails of Amblystoma opacum larvae were examined in the electron microscope and a series of stages in the differentiation of the myotome musculature was reconstructed from electron micrographs and earlier light microscopic studies of living muscle. The earliest muscle cell precursor that can be clearly identified is a round or oval cell with abundant cytoplasm containing scattered myofilaments and free ribonucleoprotein granules, but little endoplasmic reticulum. These cells sometimes form a syncytium and they may also be fused with adjacent formed muscle fibers by lateral processes. Nuclei are large and nucleoli are prominent. This cell, called a myoblast here, is distinctly different in its appearance from the adjacent mesenchymal cells which have abundant granular endoplasmic reticulum. The earliest myofilaments are of both the thick and thin varieties and are distributed in a disorganized fashion in the cytoplasm. These filaments are similar to the actin and myosin filaments described by Huxley and they are present in the cytoplasm at an earlier stage of differentiation than heretofore suspected from light microscopy studies. The first myofibrils are a heterogeneous combination of thick and thin filaments and dense Z bands and are not homogeneous as so many light microscopists have contended. As development progresses, cross striations become more orderly and definitive sarcomeres are formed. Thereafter, new myofilaments and Z bands seem to be added to the lateral surfaces and distal ends of existing myofibrils.Free ribonucleoprotein granules are a prominent part of the myoblast cytoplasm and are found in close association with the differentiating myofilaments in all stages of development. In early muscle fibers and some of the formed fibers, similar granules are often concentrated in the I bands. A theory of myofilament differentiation based on current concepts of the role of ribonucleoprotein in protein synthesis is presented in the discussion. Stages in myofibril formation and possible relationships of the filaments in developing muscle cells to other types of cytoplasmic filaments are also discussed.Supported by grant C-5196 from the United States Public Health Service.     

Ultrastructural features of supraspinal muscles in rabbits after long-term transcutaneous lateral electrical surface stimulation (LESS)

Lateral electrical surface stimulation is one of methods used in the therapy of the progressive form of idiopathic scoliosis (IS) in children and youth. However, there are data suggesting that this method may lead to serious adverse side effects, when used for a too long period of time per day. To clarify this issue, the present study was aimed at disclosing possible changes in the ultrastructural appearance of rabbit supraspinal muscles undergoing long-term stimulation (9 h per day, 3 months), an animal model successfully used to mimic the situation in humans. In comparison to the control animals, muscles of "overstimulated" rabbits exhibited clear signs of microscopical lesions, including depletion and disintegration of myofilaments, proliferation, dilatation and, sometimes, swelling of sarcoplasmic reticulum and/or mitochondria, as well as signs of destruction of the Z line. The above-mentioned abnormalities, especially the signs of degenerative processes associated with the Z line and the observed microlesions strongly suggest that the failure of the long-term LESS therapy of the IS may be attributable to these ultrastructural lesions.     

Fine structure of the homologous tergo-coxal muscles of flying and flightless beetles

The flight-related tergo-coxal muscles of flying and flightless beetles are compared. In the flying beetle, Pachynoda sinuata, the myofibrils and cylindrical and the myofilaments packed in double hexagonal arrays. The sarcomeres are short (2.8 micrometer) and wide with many large, closely packed adjacent mitochondria but the sarcoplasmic reticulum is poorly developed in this fibrillar (asynchronous) muscle. Sarcoplasmic glycogen in rosette form is abundant. In the flightless beetle, Anthia thoracica, the myofibrils are lamellar-like with sarcomeres of 5.3 micrometer. The myosin filaments form a single hexagonal array each thick filament having an orbital of 11 to 12 thin filaments. The width of the Z-line (120 nm) of A. thoracia muscle was twice that of the Z-line of P. sinuata muscle. The sarcoplasmic reticulum and T-system are well-developed in this afibrillar (synchronous) muscle. Few glycogen granules are present. Triangular projections of the sarcolemma occur regularly opposite the Z-lines in A. thoracica and they appear to extend into the Z-lines. Membranous connections joint adjacent Z-lines in A. thoracica and occasionally in P. sinuata.     

Morphometric in vitro analysis of the control of the activity of the neurosecretory dark green cells in the freshwater snail Lymnaea stagnalis (L.)

Summary The intracellular localization of calcium by means of cytochemical techniques was studied in smooth muscle cells of mouse intestine. When the lead acetate method according to Carasso and Favard (1966) was used calcium was found in mitochondria and sarcoplasmic reticulum and occasionally between the myofilaments. The active ATP-dependent accumulation of calcium into cell structures was investigated by the oxalate method (Heumann and Zebe, 1967). After appropriate treatment the only structures of smooth muscle cells which contained calcium oxalate (identified by microprobe analysis) were elements of the sarcoplasmic reticulum.The results are discussed in relation to the role of calcium in the control of muscle activity during the contraction-relaxation cycle.The electron probe microanalysis was carried out at SIEMENS (Berlin) in collaboration with Dr. von Muschwitz. I thank Miss M. Schlatter for her skillful assistance. The investigation was supported by the Deutsche Forschungsgemeinschaft.