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Fiber Types

Fiber Types. Three fiber typing schemes Metabolic/biochemical activity of fiber Protein/gene expression of fiber Functional performance of motor unit Explain the relationship between motor unit types and muscle fiber types Burke &al 1973 Schiaffino & Bottinelli, 2011.

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Fiber Types

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  1. Fiber Types • Three fiber typing schemes • Metabolic/biochemical activity of fiber • Protein/gene expression of fiber • Functional performance of motor unit • Explain the relationship between motor unit types and muscle fiber types • Burke &al 1973 • Schiaffino & Bottinelli, 2011

  2. Physiological fiber types Repeated activation Single activation • Fast Fatiguable • Fast force rise • High force • Low oxidative cap • Slow • Slow force rise • Low force • High oxidative capacity • Fast fatigue Resistant • High oxidative capacity

  3. “Fast” and “Slow” continuum • White vs red muscle (Ranvier, 1874) • Not just because of blood content • Some exceptions: masseter • Speed correlates with myosin ATPase kinetics • Heterogeneity within muscles • Mitochondria: density and activity • Sarcoplasmic reticulum volume • Glycogen and Myoglobin content • Z-disk morphology • Protein/gene expression

  4. Histochemistry • Staining: preferential dye binding • In situ chemical reactions • Structurally immobilized enzymes • Transmembrane • Sarcomeric • Reaction precipitate retained near enzyme • Soluble substrates • Often redox • NBTformazan • BCIPindigo Dephos oxidation BCIP Indigo

  5. Hematoxylin and Eosin • H: metal binding, nuclei • E: alkaline binding, protein

  6. Succinate dehydrogenase • TCA cycle, redox enzyme • Bound in mitochondria

  7. a-glycerophosphatedehydrogenase • Glycerol-3-phosphate dehydrogenase (GPDH) • Really looking at glycerol-3-phosphate shuttle • G-3-p participates in an undesired reaction Fructose 1-6 bisphosphate Dihydroxyacetone phosphate Glyceraldehyde-3-phosphate Glycerol-1-phosphate (not glycerol-3-phosphate) (not glygeraldehyde-3-phosphate) Dihydroxyacetonephosphate Glycerol-3-phosphate

  8. a-glycerolphosphatedehydrogenase • GPDH bound in Z-disks • More uniform in cross-section than mitochondria

  9. Myosin ATPase • Acid/Alkali-labile • i.e.: acid inactivates “fast” myosin; alkali, “slow” Myosin ATPase after pH 9.4 preinc

  10. Correlations in histochemistry • Mitochondrial enzymes • Succinate dehydrogenase • Citrate synthase • NAD diaphorase • Myosin • Acid-stable • Glycolytic pathway • GPDH • Lactate dehydrogenase • Glycogen • Myosin • Kinetic • Alkali-stable • Reactions tuned to give binary responses • Discrete combinations: metabolic fiber types • – Slow, oxidative (SO) • – Fast, glycolytic (FG) • – Fast, oxidative and glycolytic (FOG)

  11. Spatial distribution • Section through lower leg stained for SDH and acid-stable ATPase Lateral SO FOG FG Anterior Posterior Medial

  12. Enzyme content or activity? • Histochemistry measures total reaction • Enzyme content (moles) • Enzyme kinetics (Vmax, km) • Immunohistochemistry identifies proteins • Immunoglobulin antibodies • Extremely specific to 10-15 AA epitope • AntigenAbAb-enzyme (HRP) • AntigenAbAb-fluorophore (immunofluorescent histology) • Biochemistry, if you can purify activity

  13. Muscle proteins with multiple isoforms • Myosin • 11 heavy chains • 5 regulatory, 4 essential light chains • SERCA • Fast (1a) • Slow (2a) • Troponin • T: 4 fast, 2 slow • C, I: cardiac+slow; fast • a-actinin • Tropomyosin

  14. Myosin • 4 “conventional” isoforms: 1(o), 2a(*), 2x, 2b • Limited overlap (+) • Horseradish peroxidase/Diaminobenzidine staining A4.84 MHC1 SC-71 MHC2a My32 MHC2

  15. Myosin Slack test 2b 2x • Heavy chain • Solution ATPase rateP0 • ADP affinityVmax • F-actin velocity 2b>2x>2a>I • Light chains (essential/alkali) • Influence f-actin velocity • Probably not ATPase 2a 1 MLC3f/MLC2f Bottinelli et al., 1994

  16. SERCA • 2 dominant isoforms: 1a, 2a • Limited overlap • Fluorescein indirect fluorescence SERCA2 (in type 1 fibers) De Jonge et al., 2006 Ikezoe et al., 2003

  17. Calcium handling • SERCA • Faster calcium re-uptake with SERCA-1 • SERCA-2 inhibited by phospholamban • Parvalbumin • Only in slow fibers: quenches Ca2+ release Carroll et al., 1997

  18. Troponin • 3 subunits w/ fast/slow • C: calcium binding • T: tropomyosin binding • I: Inhibitory • Provide calcium-dependent activation • Ca2+-free form blocksMHC binding to actin Tobacman, 1996

  19. Troponin/Tropomyosin • Calcium sensitivity • Slow fibers: lower threshold & sensitivity • Fast fibers: higher sensitivity TnC-slow TM-b TnC-fast TM-b TnC-fast TM-a

  20. Expression fiber types • MHC-centric • Conventional: 1, 2a, 2x, 2b • Heart: cardiac-a, cardiac-b (MHC-I) • Developmental: embryonic, neonatal • Unconventional: Extra-ocular, Masseter (superfast) • Humans, no 2b • Co-expressed regulatory elements • miRNA • Antisense intergenic

  21. Functional correlations • Some muscles seem specialized • Soleus (cat, some rats): Type SO; high endurance • Ex Digitorum Longus (rodents): Type FG; low • Most muscles mixed • Opportunity for optimization • High use muscle benefits from oxidative metabolism • Low use muscle glycolysis minimizes oxygen delivery

  22. Fiber types • Motor unit • 1 motor neuron • 10-1000 muscle fibers • Burke et al., 1973 • Isolate MUs, test functionidentify fibers • Contractile response • # fibers • Calcium kinetics • Myosin kinetics • ATP turnover

  23. Three functional categories Repeated activation Single activation • Fast Fatiguable • Fast force rise • High force • Low oxidative cap • Fast fatigue Resistant • High oxidative capacity • Slow • Slow force rise • Low force • High oxidative capacity

  24. Physiological fiber types Characterized by Correlated with

  25. Rule-of-thumb Somewhat species & muscle-dependent Subject to activity-driven modification Staged ECCmetabolicmyosin Classification correlations

  26. Influence of FT on gross performance • Karatzaferi et al., 2001 • 10-25s ‘maximal’ cycle ergometer • ATP in type-identified fibers

  27. Influence of activity on FT • Chronic electrical stimulation • Increase oxidative metabolism • MHC 2b2x2a1 • Cross-reinnervation • MHC 12 • MHC 1masseter; MHC 2bEO • Activity pattern has some influence • Activity-independent nerve signals have some influence

  28. Summary • 3 fiber typing schemes • Protein isoforms (immunohistology) • Protein activity (histochemistry) • Function (force measurements) • Metabolic and biochemical activity correlates well with teleological function • Fiber type is dynamically regulated

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