Lecture 1 - Skeletal

Skeletal muscle

Learning Objectives

• [ ] Name the structural components of skeletal muscle

• [ ] Explain the steps involved in excitation-contraction coupling

• [ ] Give a description of the cross-bridge cycle

• [ ] Explain four mechanisms of force regulation

• [ ] Compare different types of contraction (isometric, isotonic)

• [ ] Explain the classification into fibre types, give examples

• Can activate them voluntarily

• Controlled by brain

• Skeletal muscle cells (muscle fibres) have mechanisms of electrical activation and common molecular mechanism of contraction

1.1 Structure

Common structural elements of ALL skeletal muscles:

1. Muscle body = whole muscle

2. Fascicle = bundles of muscle fibres

3. Muscle fibre (cell) = fibres are cells

4. Myofibril = each fibre contains myofibril, which is chain-like protein threads. Each link (length 2-2.5 um) of the chain is called a sarcomere

5. Protein filaments = myofibrils contains myofilaments; thick and thin. Protein myosin is main constituent of thick filaments, actin for thin filaments and regulatory proteins. Both have filament-like (qauternary) structure

   • Thick filaments contain hundreds of myosin molecules; myosin assembles as dimers in which the monomer tails form a twisted tail which connects via a hinge region to the head domain (two heads, one per myosin monomer)

• Two filamentous F-actin molecules assemble in a double helix → forms backbone of thin filaments

Connective tissue muscle components

• Skeletal muscles contain collagen-rich connective tissue components = gives additional mechanical stability

• Muscle body enclosed by epimysium

• Fascicles by perimysium

• Fibres by endomysium

Characteristic features

• Skeletal muscle fibres are LARGE cells - Characterised by:

• Elongated, cylindrical shape

• A striation pattern → produced by sarcomeres

• Many nuclei (multinucleated cells), found at cell’s periphery, under cell membrane (during development, fibres are formed by the fusion of many ‘myotubes’ with one nucleus)

• Cell membrane of muscle cells = sarcolemma

Sarcomere

• Gives striation pattern to skeletal and cardiac

• Lighter and darker = actin and myosin

• A-band (anisotropic) = myosin

• Central darker line = M-line (midline)

• I-band (isotropic) = thin filaments, made of actin + other proteins

• Myosin-actin overlap region = A-band

• Boundaries of sarcomere = Z-discs (Z-lines), actin filaments attached to main Z-disc protein, alpha-actinin-1

• Myosin molecules attached to Z-discs, via titin (spring-like molecule, contr]ols elasticity of muscle fibres)

• Length = 2-2.5 µm

• During contraction, Z approach each other, moves to M, narrowing I-band. A does not change.

• When muscle fibre activated = all sarcomeres contract simultaneously

• Sarcomeres are connected in series = shortening velocities add (long contract faster than short fibres)

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1.2 Excitation-contraction coupling

• Skeletal muscle fibres are activated by nerve impulse

• For contraction, sequence of two action potentials (AP)

  1. Produced by motor nerve axon

  2. Second AP by muscle fibre

• Nerve AP evokes muscle fibre AP after chemical transmission at neuromuscular junction

The Neuromuscular Junction

• Contact point between nervous system (motor nerve axon) and skeletal muscle fibre, similar structure to synapse

• Nerve AP reaches axon terminal → transmitter acetylcholine ACh released and diffuses to membrane of muscle fibre (motor end plate)

• Motor end plate has ACh receptors (nicotinic AChR) → when ACh binds → ionotropic receptors function as non-specific cation channels and depolarise the muscle firbre membrane

• Muscle fibre makes AP when depolarised

• Sarcolemmal enzyme cholinesterase hydrolyses, inactivates ACh

• Muscle AP follows nerve AP with short latency → due to diffusion of ACh across cleft to motor end plate AND depolarisation of muscle membrane

• Muscle AP is longer than nerve AP

The t-tubule system:

• Skeletal muscle fibres contain t-tubules

• Muscle fibre AP travels along sarcolemma → AP is distributed to all parts of cell

• Effect: depolarisation is evenly spread throughout fibre, and contraction start uniformly

The skeletal muscle triad:

• T-tubules come close to sarcoplasmic reticulum (SR), where more Ca2+ is stored

• IMPORTANT: t-tubule is part of extracellular space, SR is intracellular compartment

• 1 t-tubule is associated with 2 ‘baggy’ SR endings (terminal cisternae), structure is called triad

• Functional interaction between t-tubule and SR occurs via: - Two coupled Ca2+ channel proteins

• Dihydropyridine receptor (DHPR, t-tubule membrane)

• Ryanodine receptor (RyR, SR membrane)

• SR is more extensive in skeletal than cardiac or smooth → fast-acting muscles need to release large amounts of calcium ions in short time

Intracellular Ca2+ release:

• Muscle fibre AP → depolarisation of sarcolemma, incl. T-tubules

• DHPR is voltage-sensitive and changes its conformation due to the depolarisation

• DHPR conformational change opens RYR Ca2+ channel in SR membrane. Both proteins are coupled by non-covalent protein-protein interaction (mech coupling)

• Opening of RyR Ca2+ channel → massive Ca2+ efflux out of SR into cytosol (large concentration gradient)

• Ca2+ activates motor proteins ⇒ contraction

• NOTE: DHPR can act as Ca2+ channel but not needed for skeletal. DHPT opens RyR

Ca2+ channel. Ca2+ ions needed for contraction stem from SR = skeletal can contact in Ca2+ free solution

VIDEO: Skeletal muscle excitation-contraction coupling

1.3 Relaxation

• Contraction then relaxation

• Muscle fibre AP which elicits contraction = few milliseconds

• When fibre contraction is at its peak (= shortest fibre length), membrane potential is at baseline

• Ca2+ could theoretically sustain a continuous contraction → Ca2+ concentration returns to baseline, but slower than membrane potential. If stayed at elevated levels then cell would suffer and die

Skeletal muscle relaxation mechanisms:

• Free Ca2+ ions bound to cytosolic Ca2+ buffers. Troponin C (TnC) = Ca2+ buffer protein → initiates contraction. Ca2+ and TnC dissociate → Ca2+ bound to protein parvalbumin, removing Ca2+ ions from cross-bridge cycle

• Slower time scale: Ca2+ ions are pumped back into sarcoplasmic reticulum by primary active transport (pump)

• ATP + low Ca2+ = muscle fibre relaxes due to elastic forces in proteins

• Degradation (hydrolysis) of ACh at motor endplate by enzyme cholinesterase ❌ transmitter ACh from eliciting muscle APs

1.4 The cross-bridge cycle

• Key interaction: b/w actin and myosin → controlled by regulatory proteins; tropomyosin and troponin complex (part of thin filament, w actin)

• Troponin complex = Ca2+ sensor

• When myosin-binding site on actin is revealed → molecular motor myosin can interact with actin. ATP keeps energy-consuming process running

• Low cytosolic Ca2+ (resting) → actin-myosin binding is PREVENTED by tropomyosin

• Thinner tropomyosin filament follows helical shape + blocks myosin binding sites on actin molecule

• At regular distances, tropomyosin =

  1. Troponin

  2. a trimeric complex of troponin I (inhibitory) = TnI

  3. Troponin T (tropomyosin-binding) = TnT

  4. Troponin C (calcium binding) = TnC

Initiation:

• When Ca2+ released from SR → TnC binds to Ca2+ ion. Reaction is FAST (large rate constant)

• Calcium-bound troponin CHANGES tropomyosin = tropomyosin displaced into ‘groove’ between two F-actin strands = revealing myosin binding sites of actin

• Actin-myosin now enabled, but needs ATP

Cycle:

1. Myosin head has ATPase activity, i.e. its an enzyme.

• When ATP binds to head → hydrolysed into ADP and Pi (inorganic phosphate) + energy released by hydrolysis is used to put head in high-energy state (cocked conformation)

1. Myosin-ADP-Pi binds to actin. Lowers stability of Pi bond

2. Pi is released + myosin head performs power stroke (change of angle b/w head and tail = small displacement)

3. ADP is released = myosin in a low-energy state. Actin-myosin complex remains rigor (firmly bound)

1.5 Force Regulation

1. Firing Rate of the Motor Nerve

• If muscle needs large force = nervous system (motor neurons) produce higher rate of

APs

• Single nerve AP evokes a single twitch of muscle fibre. Repeated = multiple twitches.

When stimuli is sent at higher rate:

• Muscle does not relax completely before next twitch (tension ≠ zero)

• OR peach force becomes larger

(3) average tension reaches peak level, but partial relaxation occurs

(4) no relaxation between stimuli, muscle produces maximum force output

• Mechanism: increasing frequency = muscle fibre CANNOT reduce cytosolic Ca2+ concentration to resting levels. Elevated Ca2+ concentration recruits more proteins and fibre produces more tension

2. Motor Unit Recruitment

• Each muscle fibre controlled by ONE spinal cord motor neuron (alpha motor neuron, ɑ-MN), but each ɑ-MN can control MANY muscle fibres

• Small motor units are composed of a single ɑ-MN and only few fibres controlled by ɑ-MN

• Large motor units contains single ɑ-MN and hundred of muscle fibres

• Single AP fired by motor neuron will activate all fibres in motor unit = different motor neurons produce different force outputs

• Small motor unit = less force

| Size principle (Hennemann, 1957) | During contractions of increasing force, small motor units are recruited first (light contraction) and larger MU are recruited later (max force)         -     Small increases at low force, larger steps at high total force levls | | --- | --- |

3. Fibre/sarcomere length

• Initial fibre length determines force

• At shorter and longer sarcomere lengths ⇒ force decreases rapidly

• At peak (c) → optimum overlap of actin and myosin → all myosin heads can contribute to force production

• When sarcomere is stretched beyond 2.5 um → less overlap + fewer heads produce force

• At short lengths → myosin filament crympling, folding and collision with Z-discs is LIKELY to COUNTERACT contraction

4. Fibre diameter

• Change in fibre diameter = long-term mechanism to adapt muscle force

• g. exercise → increases rate of protein synthesis in muscle fibres → proteins assemble as sarcomere → larger amount of myofibrils → when hypertrophic fibre is activated → larger output

1.6 Types of contraction

Muscle contraction types:

• Isotonic contractions

• Concentric contractions

• Eccentric contractions

• Isometric contractions

Isotonic contraction:

• Muscle tension remains constant, length changes

• During initial phase, tension changes from zero to constant plateau

Concentric contraction

-     Isotonic contraction where muscle SHORTENS

Eccentric

• Isotonic contraction but muscle LENGTHENS even when cross-bridge is running

• g. supporting heavy weight in your hand and extend flexed elbow → biceps is active and lengthening

Isometric

• Muscle length stays constant while muscle produces tension

• No net shortening or lengthening

• Myosin heads perform power strokes but do not slide along actin, due to counteracting force pointing in opposite direction

1.7 Fibre Types

3 Fibre Types

1. Slow oxidative fibres (type I)

2. Fast oxidative fibres (type IIA)

3. Fast glycolytic fibres (type IIX)

• Different rate of force prod\uction = kinetics of myosin-ATPase fibre types contain

• Rate constant of ATP hydrolysis is different → fibres contain different myosin molecules = slower ATP hydrolysis rates + slower shortening velocities

Slow Oxidative Fibres

• Produce low forces

• Contract slowly but sustains for longer (fatigue resistant)

• Generates ATP for long time cia oxidative metabolic pathways (fatty acid oxidation, oxidative phosphorylation

• Need many mitochondria + high density of capillaries

• High content of oxygen-binding molecule myoglobin = red

Fast glycolytic fibres

• Produce large forces

• Higher speed of shortening, cannot contract for long (non fatigue-resistant)

• Generates ATP via anaerobic pathways (glycolysis), produces lactate

• Less mitochondria and few capillaries

• Low myoglobin = white

Same muscle, different fibres

• In EDL (extensor digitorum longus) = shin

• Superficial part: activated in fast, high-energy movements (walking, running, jumping) = more gast glycolytic (type IIX) fibres

• Deeper: postural stability (standing) = slow oxidative fibres (type I)