MUSCLE TISSUE Petr Vaňhara https://i.pinimg.com/originals/69/8d/e7/698de768ff8638068faea5c156a02034.jpg Epithelium Muscle Nerve Connective Based on morphology and function: Cytoskeleton → contraction Mesoderm – skeletal muscle, myocard, mesenchyme – smooth muscles Rarely ectoderm (eg. m. sphincter a m. dilatator pupillae) Neurons and neuroglia Reception and transmission of electric signals Ectoderm, rarely mesoderm (microglia) Dominant extracellular matrix Connective tissue, cartilage, bone… Mesenchyme Continual, avascular layers of cells with different functions, oriented to open space, with specific junctions and minimum of ECM and intercellular space. Derivates of all three germ layers CONTEMPORARY TISSUE CLASSIFICATION Hallmarks ▪ Unique cell architecture ▪ Excitability and contraction ▪ Mesodermal origin GENERAL CHARACTERISTIC OF MUSCLE TISSUE Striated skeletal Striated cardiac Smooth Classification according to cell and tissue structure MUSCLE TISSUE STRIATED SKELETAL MUSCLE TISSUE − Composition: muscle cells + connective tissue, blood vessels − Unique cell architecture: long multinuclear cells – muscle fibers (rhabdomyocytes) − Long axis of cells is oriented in parallel with direction of contraction − Specific terminology: • cell membrane = sarcolemma • cytoplasm = sarcoplasm • sER = sarcoplasmic reticulum • Muscle fiber – microscopic unit of skeletal muscle • Myofibril – LM unit – myofilaments – unit of muscle fibers • Myofilaments – filaments of actin and myosin (EM) TERMINOLOGY OF SKELETAL MUSCLE TISSUE TERMINOLOGY OF SKELETAL MUSCLE TISSUE SKELETAL MUSCLE TISSUE http://medcell.org/histology/muscle_lab/skeletal_muscle_longitudinal_section.php SKELETAL MUSCLE TISSUE http://medcell.org/histology/muscle_lab/skeletal_muscle_cross_section.php STRUCTURE OF SKELETAL MUSCLE KAPILÁRY KOLEM SVALOVÝCH VLÁKEN − morphological and functional unit: muscle fiber (rhabdomyocyte) – elongated, cylindrical-shaped, multinucleated cell (syncytium) − nuclei are located at the periphery (under sarcolemma) − myofibrils show cross striation − diameter of muscle fiber: 25-100 m − length: millimeters - centimeters (up to 15) STRUCTURE OF SKELETAL MUSCLE Muscle fiber = myofiber = syncitium = rhabdomyocyte Muscle fiber – morphological and functional unit of skeletal muscle [Ø 25 – 100 m] Myofibrils – compartment of fiber sarcoplasm [Ø 0.5 – 1.5 m] Sarcomere – the smallest contractile unit [2.5 m], serial arrangement in myofibrils Myofilaments – actin and myosin, are organized into sarcomeres [Ø 8 and 15 nm] ULTRASTRUCTURE OF RHABDOMYOCYTE • Myosin heavy chain (MHC) type I and II - distinct metabolic, contractile, and motor-unit properties - ATPase activity • Twitch type - Fast vs. slow • Fiber color - Red vs. white • Myoglobin content • Glycogen content • Energy metabolism • Endurance PHYSIOLOGICAL TYPES OF SKELETAL MUSCLES PHYSIOLOGICAL TYPES OF SKELETAL MUSCLES Properties Type I fibers Type IIA fibers Type IIX fibers Motor Unit Type Slow Oxidative (SO) Fast Oxidative/Glycolytic (FOG) Fast Glycolytic (FG) Twitch Speed Slow Fast Fast Twitch Force Small Medium Large Resistance to fatigue High High Low Glycogen Content Low High High Capillary Supply Rich Rich Poor Myoglobin High High Low Red Color Dark Dark Pale Mitochondrial density High High Low Capillary density High Intermediate Low Oxidative Enzyme Capacity High Intermediate-high Low Z-Line Width Intermediate Wide Narrow Alkaline ATPase Activity Low High High Acidic ATPase Activity High Medium-high Low PHYSIOLOGICAL TYPES OF SKELETAL MUSCLES T-tubule terminal cisterna mitochondria myofibrils sarcolemma Sarcolemma + t-tubules, Sarcoplasm: Nuclei, Mitochondria, Golgi apparatus, Glycogen ( granules) Sarcoplasmic reticulum (smooth ER) – reservoir of Ca2+ Myofibrils (parallel to the length of the muscle fiber) tubules + cisternae of sER ULTRASTRUCTURE OF RHABDOMYOCYTE − elongated structures [Ø 0.5 – 1.5 ] in sarcoplasm of muscle fiber oriented in parallel to the length of the fiber, − Actin + myosin myofilaments − Sarcomere − Z-line − M-line and H-zone − I-band, A-band MYOFIBRILS MYOFIBRILS SARCOMERE A–band I–band½ I-band H-zone SARCOMERE SARCOMERE • Terminal cistern • T-tubule • Terminal cistern TRIAD • communicating intracellular cavities around myofibrils, separated from cytosol • terminal cisternae (“junction”) and longitudinal tubules (“L” system). • reservoir of CaII+ ions • T-tubules (“T” system ) are invaginations of sarcolemma and bring action potential to terminal cisternae change permeability of membrane for CaII+ ions SARCOPLASMIC RETICULUM SARCOPLASMIC RETICULUM SARCOPLASMIC RETICULUM SARCOPLASMIC RETICULUM http://medcell.org/histology/muscle_lab/skeletal_muscle_em.php ULTRASTRUCTURE OF RHABDOMYOCYTE MYOFILAMENTS • Fibrilar actin (F-actin), ( 7 nm, 1 m) • Tropomyosin – thin double helix in groove of actin double helix, spans 7 monomers of G-actin • Troponin – complex of 3 globular proteins • TnT (Troponin T) – binds tropomyosin • TnC (Troponin C) – binds calcium • TnI (Troponin I) inhibits interaction between thick and thin filaments THIN MYOFILAMENTS • Myosin II - Large polypeptide, golf stick shape, ( 15 nm, 1,5 m) - Bundles of myosin molecules form thick myofilament THICK MYOFILAMENTS • Nebulin - 600-900kDa - F-actin stabilization - length of sarcomere • Titin (konektin) - >MDa - myosin stabilization - elastic OTHER PROTEINS ASOCIATED WITH MYOFILAMENTS • α-actinin - Z-line - binds actin OTHER PROTEINS ASOCIATED WITH MYOFILAMENTS • Sarcomere is a complex structure. Defects caused by mutations in sarcomere proteins are linked to various myopathies. MYOFILAMENTS ASSEMBLE TO CONTRACTIVE STRUCTURES − Propagation of action potential (depolarization) via T-tubule (= invagination of sarcolemma) − Change of terminal cisternae permeability – releasing of Ca+ ions increases their concentration in sarcoplasm − Myosin binds actin - sarcomera then shortens by sliding movement – contraction − Relaxation: repolarization, decreasing of Ca2+ ions concentration, inactivation of binding sites of actin for myosin myosin actin MYOFILAMENTS ASSEMBLE TO CONTRACTIVE STRUCTURES CROSS BRIDGE CYCLE Huxley, H. E. & Hanson, J. Changes in the cross-striations of muscle during contraction and stretch and their structural interpretation. Nature 173, 973–976 (1954) doi:10.1038/173973a0. Huxley, A. F. & Niedergerke, R. Structural changes in muscle during contraction: Interference microscopy of living muscle fibres. Nature 173, 971–973 (1954) doi:10.1038/173971a0. 1. Impulse along motor neuron axon 2. Depolarization of presynaptic membrane (Na+ influx) 3. Synaptic vesicles fuse with presynaptic membrane 4. Acetylcholine is exocyted to synaptic cleft 5. Acetylcholine diffuses over synaptic cleft 6. Acetylcholine binds to receptors in postsynaptic membrane 7. Depolarization of postsynaptic membrane and sarcolemma (Na+ influx) 8. T-tubules depolarization 9. Depolarization of terminal cisternae of sER 10. Depolarization of complete sER 11. Release of CaII+ from sER to sarcoplasm 12. CaII+ binds TnC 13. Troponin complex changes configuration 14. Tropomyosin allows binding of actin and myosin 15. Globular parts of myosin bind to actin 16. ATPase in globular parts of myosin activated 17. Energy generated from ATP→ADP + Pi enables cross-bridge cycling 18. Movement of globular parts of myosin 19. Actin myofilament drag to the center of sarcomere 20. Sarcomeres contract (H-zone, I-band shorten) 21. Myofibrils contract 22. Muscle fiber contract MECHANISM OF CONTRACTION Courtesy Dr. Pacherník, Faculty of Science MU CALCIUM FLOW FROM SARCOPLASMIC RETICULUM 1 2 3 4 Myelinated axons Neuromuscular junction Capillaries Muscle fiber nucleus NEUROMUSCULAR JUNCTION NEUROMUSKULÁRNÍ SPOJENÍ http://medcell.org/histology/muscle_lab/neuromuscular_junction.php NEUROMUSCULAR JUNCTION NEUROMUSCULAR JUNCTION NEUROMUSCULAR JUNCTION http://medcell.org/histology/muscle_lab/neuromuscular_junction_em.php MYASTHENIA GRAVIS NEUROMUSCULAR JUNCTION Curare Botulotoxin Clostridium botulinum NEUROMUSCULAR JUNCTION • Structural components linking myofibrils to sarcolemma • Circumferential alignment • dystrophin-associated glycoprotein (DAG) complex • links internal cytoskelet to ECM • Integrity of muscle fiber COSTAMERES COSTAMERES COSTAMERES AND DYSTROPHIN COSTAMERES AND DYSTROPHIN DUCHENNE MUSCULAR DYSTROPHY Lane 1: Becker dystrophy; Dystrophin has reduced abundance but normal size. Lane 2: Becker dystrophy; Dystrophin has reduced size and abundance. Lane 3: Normal; Dystrophin has normal size and amount. Lane 4: Duchenne dystrophy; Almost no protein is present. http://neuromuscular.wustl.edu/pathol/dmdpath.htm COSTAMERES AND DYSTROPHIN COSTAMERES AND DYSTROPHIN COSTAMERES - ATTACHMENT TO ECM − Endomysium – around each muscle cell (fiber) − Perimysium – around and among the primary bundles of muscle cells − Epimysium – dense irregular collagen c.t., continuous with tendons and fascia − Fascia – dense regular collagen c.t. − Containment − Limit of expansion of the muscle − Transmission of muscular forces CONNECTIVE TISSUE OF SKELETAL MUSCLE CONNECTIVE TISSUE OF SKELETAL MUSCLE Connective tissue around muscle bundles and muscle fibers Myotendinous junction CONNECTIVE TISSUE OF SKELETAL MUSCLE PROPRIORECEPTORS Golgiho tendon organs - myotendineous junction - senzory endings synapsed with inhibitory neurons - tension, stretch Muscle spindles - change in muscle elongation (stretch) - modified perimysium - thin muscle (intrafusal) fibers - sensory endings - reflexes, coordination of muscle groups CARDIAC MUSCLE TISSUE MUSCLE TISSUE • made up of long branched fiber (cells) – cardiomyocytes, • cardiomyocytes are cylindrical cells, branched on one or both ends (Y, X shaped cells), • sarcoplasm: single nucleus in the center of cell, striated myofibrils, numerous mitochondria, • cells are attached to one another by end-to-end junctions – intercalated discs. HISTOLOGY OF CARDIAC MUSCLE TISSUE chains of cardiomyocytes blood capillary with erythrocytes Intercalated disc HISTOLOGY OF CARDIAC MUSCLE TISSUE HISTOLOGY OF CARDIAC MUSCLE TISSUE ULTRASTRUCTURE OF CARDIOMYCYTE − no triads, but diads: 1 t-tubule + 1 cisterna − t-tubules around sarcomeres at Z lines rather than at zone of overlap − sarcoplasmic reticulum via its tubules contact sarcolemma as well as the t-tubules − cardiac muscle cells are totally dependent on aerobic metabolism to obtain the energy − large numbers of mitochondria in sarcoplasm and abundant reserves of myoglobin (to store oxygen) − abundant glycogen and lipid inclusions CARDIAC MUSCLE COMPARED TO SKELETAL − fasciae adherentes (adhesion of cells) − nexus (quick intercellular communication – transport of ions, electric impulses, information) − „scalariform“ shape of cell ends INTERCALATED DISC INTERCALATED DISC nexus fascia adherensIntercalated disc: INTERCALATED DISC • Actin + myosin myofilaments • Sarcomere • Z-line • M-line and H-zone • I-band, A-band • T-tubule + 1 cistern = diad (around Z-line) MYOFIBRILS IN CARDIOMYOCYTE MYOFIBRILS IN CARDIOMYOCYTE PURKINJE FIBERS − are located in the inner layer of heart ventricle wall − are specialized cells fibers that conduct electrical stimuli or impulses that enables the heart to contract in a coordinated fashion − numerous sodium ion channels and mitochondria, fewer myofibrils SPECIALIZED CARDIOMYOCYTES PURKINJE FIBERS http://medcell.org/histology/muscle_lab/purkinje_fibers.php ATRIAL CARDIOMYOCYTES • Natriuretic peptide A (ANP) • atrial cardiomyocytes • vasodilatation, diuresis SPECIALIZED CARDIOMYOCYTES SMOOTH MUSCLE TISSUE MUSCLE TISSUE • Cells – leiomyocytes - form layers - eg. in walls of hollow organs Crosssection Longitudinal SMOOTH MUSCLE TISSUE • spindle shaped cells (leiomyocytes) with myofilaments not arranged into myofibrils (no striation), 1 nucleus in the centre of the cell • myofilaments form bands throughout the cell • actin filaments attach to the sarcolemma by focal adhesions or to the dense bodies substituting Z-lines in sarcoplasm • sarcoplasmic reticulum forms only tubules, CaII+ ions are transported to the cell via caveolae and pinocytic vesicles • zonulae occludentes and nexuses connect cells • calmodulin SMOOTH MUSCLE TISSUE • caveolae are equivalent to t-tubules • transmembrane ion channels CAVEOLAE CAVEOLAE + nexuses CONTRACTION OF LEIOMYCYTES CONTRACTION OF LEIOMYCYTES SMOOTH MUSCLE TISSUE Hallmark Skeletal muscle Cardiac muscle Smooth muscle Cells Thick, long, cylindrical, non-branched Branched, cylindrical Small, spindle-shaped Nuclei Abundant, peripherally 1-2, centrally 1, centrally Filaments ratio (thin:thick) 6:1 6:1 12:1 sER and myofibrils Regular sER around myofibrils Less regular sER, myofibrils less apparent Less regular sER, myofibrils not developed T tubules Between A-I band, triads Z lines, diads Not developed Motor end plate Present Not present Not present Motor regulation Voluntary control No voluntary control No voluntary control Other Large multinucleated cells in bundles, c.t. Intercalated discs, working and specialized cardiomyocytes Caveolae, overlapping cells in layers SUMMARY HISTOGENESIS OF SKELETAL MUSCLE TISSUE HISTOGENESIS OF SKELETAL MUSCLE TISSUE Embryonic progenitors Satellite cells Myoblasts Myocytes Myotubes & myofibers SATELLITE CELLS ARE REQUIRED FOR REGENERATION Satellite cells are equivalent to embryonic precursors of muscle fibers DIFERENTIATION IN VITRO TISSUE ENGINEERING https://www.nature.com/news/artificial- jellyfish-built-from-rat-cells-1.11046 https://www.nature.com/articles/nbt.2269 THANK YOU FOR ATTENTION pvanhara@med.muni.cz http://www.histology.med.muni.cz/