Undergraduate – Graduate Muscle Histology Lecture Series Larry Johnson, Professor Veterinary Integrative Biosciences Texas A&M University College Station, TX 77843 Muscle – Introduction Contractivity is one of the fundamental properties of protoplasm and is exhibited in varying degree by nearly all cell types. In the cells of muscle, the ability to convert chemical energy into mechanical work has become highly developed. Locomotion of multicellular animals, beating of their hearts, and movement of their internal organs depends on muscles of different types. Skeletal muscle Cardiac muscle Smooth muscle Objectives Identify smooth, skeletal, and cardiac muscle on route histological preparations Explain the morphological basis for the different functions of these three types of muscle Distinguish between the modes of excitation of these three types of muscle Four basic types of tissues Epithelium (90% of tumors) Connective tissue Muscular tissue Nervous tissue Muscle Function: Generation of contractile force Distinguishing features: high concentration of contractile Smooth muscle proteins actin and myosin arranged either diffusely in the cytoplasm Striated muscles (smooth muscle) or in regular repeating units called sarcomeres (striated muscles, e.g., cardiac and skeletal muscles) Cardiac muscle Muscle - Histological identification Skeletal muscle – very long Dilator muscle of iris cylindrical striated muscle cells with multiple peripheral nuclei Myoepithelial cells Cardiac muscle – short branching striated muscle cells with centrally located nuclei Smooth muscle – closely packed spindle-shaped cells with a single centrally placed nucleus and cytoplasm that appears homogeneous by light microscopy Muscle Distribution: Skeletal – striated muscles mostly associated with the skeleton Muscle Distribution: Cardiac – striated muscles associated with the heart large artery of lung Muscle Distribution: Smooth – fusiform cells associated with the viscera, respiratory tract, blood vessels, uterus, etc. Smooth muscle Ureter Ductus deferens Types of muscle Skeletal muscle – Voluntary, large and multinucleated cells, striated Cardiac muscle – Involuntary, mononucleated and branched cells, striated Smooth muscle – Involuntary, mononucleated, non-striated Connective tissue layers of skeletal muscle Epimysium - coarse CT Perimysium - less coarse CT Endomysium - delicate CT Perimysium Epimysium Endomysium Tongue, monkey Skeletal muscle nuclei Fasciculi Endomysium Muscle cells skeletal muscle nuclei, Connective tissue of perimysium striations Connective Tissue connects cells (muscle fibers) of skeletal muscle Endomysium Connective Tissue Layers of Skeletal Muscle PERIMYSIUM ENDOMYSIUM Connective Tissue Layers of Skeletal Muscle Endomysium Individual cell Striated Muscle Skeletal Cardiac A I “A” Band = dark band Anisotropic = does alter polarized light A (Birefringent) I “I” Band = light band Isotropic = does not alter polarized light A I Polarized Light Micrograph Of Human… High-Res Stock Photography ...www.gettyimages.com Striated Muscle (Skeletal) Repeating A and I bands alone the cell’s length creates repeating sarcomeres A I A I A I A I Striated Muscle (skeletal) A I Sarcomeres are organized for rapid and highly controlled contraction Striated Muscle (Skeletal) Sarcomere = structural unit and functional unit of striated muscle Striated Muscle (Skeletal) Thin filament = actin + actin-associated proteins Actin-associated proteins dictate network or bundle creating the Z line Thick filament = myosin Striated Muscle Striated Muscle Note uniform spacing of troponin Striated Muscle Unexplained complexity in skeletal muscle 13 isoforms of myosin 128 isoforms of troponin • Footprints of evolution – fossils – comparative anatomy, morphology and physiology – biological macromolecules • nucleic acids & proteins • document evolutionary history • provide insights into evolution of form and function – life – biomolecules • e.g., cytochrome c in rice & tuna Slides adapted from Dr. Chris Collet Queensland University of Technology Australia Based on scientific research, what three characteristics do these mammals all have in common… 2. Mammary with these mammals? glands 1. Hair 3.Special inner ear bones Ear bones of mammals (including human) began as reptile jaws This 125-million year old fossil has inner-ear anatomy intermediate (still attached to the jaw) between reptiles and mammals. In the early embryonic stage of modern mammals, the middle ear was still attached to the jaw. • Footprints of evolution – fossils – comparative anatomy, morphology and physiology – biological macromolecules • nucleic acids & proteins • document evolutionary history • provide insights into evolution of form and function – life – biomolecules • e.g., cytochrome c in rice & tuna Slides adapted from Dr. Chris Collet Queensland University of Technology Australia You can learn a lot about humans from studying animals • Footprints of evolution – fossils – comparative anatomy, morphology and physiology – biological macromolecules • nucleic acids & proteins • document evolutionary history • provide insights into evolution of form and function – life – biomolecules • e.g., cytochrome c in rice & tuna Slides adapted from Dr. Chris Collet Queensland University of Technology Australia Introduction: Pathways Of Protein Evolution Protein Evolutionary Trees Introduction: Pathways Of Protein Evolution • Point mutation – change of function to meet changing requirements • Duplication – simplest mechanism of evolving new proteins – functional divergence of duplicates to meet new requirements in biochemical pathways • Exon shuffling – creating novel proteins for new pathways of development • Alternate splicing – protein diversity from existing genes Exon Shuffling And Mosaic Proteins If structural = functional modules then – modules (domains) can be moved around genome – fulfill new functions – proteins show a mosaic history Exon Shuffling and Mosaic Proteins Many proteins are modular – units derived from many sources Alternate Pathways Of Transcript Splicing • Different exons may be joined to produce a related set of mRNAs encoding a small family of related proteins – protein isoforms • Splicing patterns often tissue-specific • Related proteins may perform similar, not necessarily identical, functions in different types of cells • Splicing is the norm in elks as a means of producing diversity Unexplained complexity in skeletal muscle 13 isoforms of myosin 128 isoforms of troponin Cell Structure of Skeletal Muscle Myofiber = multinucleated cell Myofibrils Sarcomere – Z Line (α-actinin) – I Band (actin, tropomyosin, H troponins) – A Band (myosin, overlaps actin) – H Band (myosin with no overlap of actin) Cell Structure of Skeletal Muscle Individual cell Individual cells Cell Structure of Skeletal Muscle Cell Structure of Skeletal Muscle Skeletal Muscle Wall Paper Skeletal Muscle Sarcomeres shorten to create contraction Skeletal Muscle Remember the Intermediate Filaments on Epithelium Structural support of epithelial desmosomes and hemidesmosomes Intermediate Filaments – Function in Muscle Cells Myofibril organization – Muscle cells Cell = Contraction of the Sarcomere Thin Filament Actin (F-actin) Tropomyosin Troponin T - attaches to tropomyosin C - binds calcium ions I - inhibits actin-myosin interaction Thick Filament (myosin) Sliding filament theory of contraction Sliding filament theory of contraction of the sarcomere Contraction (know five steps) 1. Troponin-C binds calcium 2. Troponin changes shape causing conformational change in tropomyosin exposing actin binding site 3. Myosin binds actin and released inorganic phosphate inducing 4. Movement of myosin head (motor, power stroke ) and sliding of actin filament in relation to the myosin filament 5. ATP ADP and inorganic phosphate binds to myosin head cocking it Contraction of the sarcomere Contraction of the sarcomere http://www.youtube.com/watch?v=gJ309LfHQ3M https://www.youtube.com/watch?v=0kFmbrRJq4w Calcium Regulation Transverse (T) tubule (invagination of sarcolemma) transmit depolarization of membrane deep into the cell Sarcoplasmic reticulum (SER of cell) release Ca++ for contraction – then recovers Ca++ after contraction Triad = (T tubule and two ends of SER) Calcium Regulation TRANVERSE (T) TUBULE TRIAD = (T TUBULE + TWO ENDS OF SER) Calcium Regulation Transverse Tubule Stimulation of Muscle Cells Innervation of Skeletal Muscle Motor end-plate: Synaptic cleft Acetylcholine and receptor Junctional Folds Innervation of Muscle Slide HISTO007 skeletal muscle cells Nerve – muscle interface at the motor end plates Note the motor end plates in several skeletal muscle cells Innervation of Muscle Innervation of Muscle Innervation of Muscle Innervation of Muscle Innervation of Muscle Innervation of Muscle Sensory Innervation of Muscle Muscle Fiber / Cell Muscle Spindle Innervation of Muscle Muscle Spindle Muscle Spindle Intracapsular fibers Tongue Muscle spindle Muscle spindles Intrafusal fibers inside the capsule capillaries nerve fibroblasts Types of Fibers in Skeletal Muscle Red (Slow, Oxidative) – High Myoglobin – High Cytochromes/ Mitochondria – Posture, flight muscle in birds Types of Fibers in Skeletal Muscle White (Fast, Glycolytic) – Low Myoglobin – Fewer Mitochondria Types of Fibers in Skeletal Muscle Intermediate (Fast, Oxidative, and Glycolytic) http://www.youtube.com/watch?v=pbTah5NVOtU&feature=r elated Cardiac Muscle Cardiac Muscle is Striated Muscle Differences From Skeletal Muscle – Mononucleated vs. Multinucleated – Central