Muscles – general information
Vertebrates and many invertebrates have three main classes of muscle
 Skeletal muscle connect bones are are used for complex coordianted
activities.
 Smooth muscles surround internal organs such as the large and small
intestines, the uterus, and large blood vessels
The contraction and relaxation of smooth muscles controls the
diameter of blood vessels and also propels food along the
gastrointestinal tract.
Compared with skeletal muscles, smooth muscle cells contract and
relax slowly, and they can create and maintain tension for long
periods of time.
 Cardiac muscle: Striated muscle of the heart.
Muscles - introduction
B. Skeletal muscle from the neck of a hamster
C. Heart muscle from a rat
D. Smooth muscle from the urinary bladder of a guinea pig
E. Myoepithelial cells in a secretory alveolus from a lactating rat mammary
gland
Microtubule Function
• Move subcellular components
• Use motor proteins kinesin and dynein
• e.g., Rapid change in skin color
Microtubules Show Dynamic Instability
Balance between growth and shrinkage
Factors
• Local concentrations of tubulin
• Dynamic instability
• Microtubule-associated proteins (MAPs)
• Temperature
Chemicals can disrupt the dynamics (e.g., plant poisons)
Movement Along Microtubules
Direction is determined by polarity and the type of motor protein
• Kinesin move in + direction
• Dynein moves in – direction
Fueled by ATP
Rate of movement is determined by the ATPase domain of the protein and
regulatory proteins
Dynein is larger than kinesin and moves 5-times faster
Cilia and Flagella
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Cilia – numerous, wavelike motion
Flagella – single or in pairs, whiplike movement
Composed of microtubules
Arranged into axoneme
Movement results from asymmetric activation of dynein
Microtubules and Physiology
Microfilaments
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Other type of cytoskeletal fiber
Polymers composed of the protein actin
Often use the motor protein myosin
Found in all eukaryotic cells
Movement arises from
• Actin polymerization
• Sliding filament model using myosin (more common)
Microfilament Structure and Growth
• Polymers of G-actin
called F-actin
• Spontaneous growth
(6-10X faster at +
end)
• Treadmilling when
length is constant
• Capping proteins
increase length by
stabilizing minus end
Microfilament Arrangement
Actin Polymerization
Amoeboid movement
Two types
• Filapodia are rodlike
extensions
• Neural connections
• Microvilli of digestive
epithelia
• Lamellapodia
resemble
pseudopodia
• Leukocytes
• Macrophages
Skeletal muscle (striated muscle)
• Skeletal muscle cells are one of the largest cells in the body
• Are multinucleate formed by the fusion of myoblasts
• Diameters range from 50 to 150 microns with lengths ranging
from mm to cm
• Muscle fibers contract in response to an electrical signal ie
depolarization
• The signal is generated at the synapse (the neuromuscular
junction) and propagated through an action potential via the
muscle fiber membrane
• The membrane of the cell has specialized invaginations called
Transverse-tubules (T-tubules) that enter into the cell (at every 12 microns)
• The action potential can be rapidly transmitted deep into the
interior of the cell resulting a delay of only 3-5 msec between the
depolarization at the synapse and the first muscle fiber tension.
• The T-tubule network is so extensive that 50-80% of the plasma
membrane is in the T-tubules.
Neuromuscular junction – things to remember
• Each muscle fiber is innervated by a motorneuron
• One motorneuron can innervate one or multiple fibers
• Each motorneuron plus its complement of muscle fibers is called a
motor unit as well all contract in concert.
• The synpase between the motorneuron and the muscle fiber is
called a neuromuscular junction(NMJ)
• Nerve terminal contains many mitochondria and vesicles which can
be seen lined up in double rows along side the voltage-gated Ca2+
channels attached to presynaptic membrane => active zone.
• The vesicles of the NMJ have very high concentrations of
neurotransmitter (2,000 to 10,000 molecules of ACH per vesicle)
• Excess of neurotransmitter is released to ensure that the resulting
post-synaptic depolarization is strong enough to generate an action
potential - "safety margin"
Neuromuscular junction – things to know
The nACHR - again
• The receptor is made up of 4 different transmembrane proteins one of
which (the alpha subunit) is repeated to give 5 subunits to create the ion
channel
• ACH binds to the alpha subunit and thus it takes two molecules of
acetylcholine to open the channel
• One nACHR opens and allow 1.5 x 104 Na+ ions/msec of open time
• The channel opens on average 1 msec.
• 1 vesicle contains enough neurotransmitter to open ~3000 receptors
(wow!) and because two molecules of Ach is needed to open one receptor
there must be a minimum of ~6,000 molecules Ach per vesicle.
• Studies have shown that the amount of neurotransmitter contained in one
vesicle causes an post-synaptic potential of ~ 1 mV.
• If the average depolarization generated at a NMJ
of a muscle fiber is 40 mV then there must be at
least 40 vesicles released and in the order of
120,000 receptors activated at the NMJ. WOW!
Striated muscle channels and action potentials
Striated muscle channels and action potentials
Pumps and transporters
1) Na+/K+ ATPase pump - to establish the electrochemical gradients of
Na+ and K+
2) Ca2+ ATPase pump - uses energy from ATP to remove 2 Ca2+ from the
inside to the outside of the cell to ensure that internal Ca2+
concentrations remain low (10-7 mM internal)
3) Na+/Ca2+ cotransporter - to also remove Ca2+ from the inside of the cell
and uses the energy from the cotransport of 3 Na+ molecules to
export 1 Ca2+
4) Muscle Ca2+ ATPase pump - a different pump from number 2 above.
Found highly concentrated on the sarcoplasmic reticulum (SR)
(constitutes 80% of the protein found in the SR membranes).
The muscle Ca2+ ATPase pumps 2 Ca2+ into the SR to lower cytosolic
Ca 2+ and to concentrate Ca 2+ into internal stores.
Striated muscle channels and action potentials
Channels - muscle cells share many of the same ion channels as
neurons
1) leak channels - besides the leak K+ channel, skeletal muscle cells have a
high concentration of Cl- leak channels so much so that the resting
membrane potential is usually the same as the Nernst potential for Cl(around -80 mV in these cells).
The high permeability to Cl- helps repolarize the membrane after an
action potential
2) Voltage-gated Na+ channels - Voltage-gated Na+ channel (very much like
the neuronal voltage-gated Na+ channel)
Responsible for the production of the action potential. Remember the
regenerative cycle of these channels ie during an Action potential
3) Voltage-gated K+ channel - the delayed rectifier K+
Has a high threshold and needs a strong depolarization to open and
works to bring the membrane back to resting potential
4) Voltage gated Ca2+channels - high threshold Ca2+ channels
Needs a strong depolarization to open and very slowly
Concentrated in the T-tubules.
Skeletal muscle (striated muscle)
Terminology
• Muscle cell: Muscle fiber
• Myofibrils: Main intracellular structures in striated muscles. Are bundles of
contractile and elastic proteins
• Sarcolemma: Cell membrane of a muscle cell
• Cytoplasm: Sarcoplasm
• Sarcoplasmic reticulum: wraps around each myofibril like a piece of lace.
Release and sequester Ca2+ ions
Skeletal muscle (striated muscle)
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The sarcoplasmic reticulum (SR) regulates the cytosolic Ca2+ levels in
skeletal muscle
Myofibril: A long bundle of actin, myosin and associated proteins in
muscle cell.
Transverse (T) tubules: invaginations of the plasma membrane, enter
myofibers at the Z disks, where they come in close contact with the
terminal cisternae of the SR
Terminal cisternae: store Ca2+ ions and connect with the lacelike
network of SR tubules that overlie the A band.
T-Tubules and SRs
Transverse tubules
• Sarcolemmal invaginations
• Enhance action potential
penetration
• More developed in larger,
faster twitching muscles
• Sarcoplasmic reticulum (SR)
• Stores Ca2+
• Terminal cisternae -  storage
Triads and Sarcoplasmic reticulum
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The link between depolarization and Ca2+ release or excitationcontraction coupling occurs at the junctions between the T-tubule and
the sarcoplasmic reticulum
80% of the T-tubules membrane is associated with the sarcoplasmic
reticulum at triads
The voltage-gated Ca2+ channels are concentrated in the T-tubules in
the triads
The Ca2+ release channel found in the sarcoplasmic reticulum
membrane is associated with the voltage-gated Ca2+ channel at this
point
This close association allows for the rapid signaling from action
potential to Ca2+ release.
General sequence of events
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Resting [Ca2+]i = 0.1 μM
AP propagation along Sarcolemma and into T- tubules
Depolarization opens the voltage-gated Ca2+ channels at triad junctions
This results in a release of Ca2+ through the Ca2+ release channels from
the SR
Cytosolic [Ca2+]i reaches 1-10 μM
Diffusion and binding of Ca2+ to TnC
Contraction events
[Ca2+] to resting levels:
1. After the action potential is passed and the voltage-gated Ca2+
channels close, the Ca2+ release channels close
2. Ca2+ is recycled back into the SR through the Ca2+ ATPases
3. Ca2+ binds to calsequesterin
Ca2+ channels and its release
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Release of Ca2+ stores mediated by ryanodine receptors (RYRs) in
skeletal muscle
Voltage sensing dihydropyridine (DHP) receptors in the plasma
membrane contact ryanodine receptors located in the membrane of the
SR
In response to a change in voltage, the dihydropyridine receptors
undergo a conformational change
This produces a conformational change in the associated RYRs, opening
them so that Ca2+ ions can exit into the cytosol.
The voltage-gated Ca2+ channel is either closely localized to or makes a
physical connection to the Ca2+ release channels in the SR
Cont……..
Ca2+ channels and its release
• Not all Ca2+ release channels are associated with voltage-gated Ca2+
channels
• These non-associated channels are thought to be opened solely by Ca2
influx into the cytosol from the voltage-gated Ca2+ channels.
• The Ca2+ release channel in the SR of most muscle cells (smooth, cardiac,
skeletal) is a Ca2+ activated Ca2+ channel
• The Ca2+ release channel is stimulated to open at low concentrations of
Ca2+ ( up to 0.1 mM) in the cytosol but inhibited by high concentrations of
Ca2+ in the cytosol (0.5 mM and higher for cardiac cells)
• So as Ca2+ is released from the SR it starts to inhibit the Ca2+ release
channel.
Depolarization induced Ca2+ release
Ca2+ induced Ca2+ release
Experiment
Sarcomeres
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Skeletal muscle is made up of bundles of multinucleate muscle cells
(myofibers)
Each cell contains myofibrils that are composed of repeated units of
actin and myosin called sacromeres
Thick and thin filaments arranged into sarcomeres
Repeated in parallel and
in series
Features
• Z-disk
• A-band
• I-band
• M-lines
Sarcomeres
• Electron micrograph of a longitudinal section through a skeletal muscle
cell of a rabbit
• Schematic diagram of a single sarcomere
• Z discs: At each end of the sarcomere
• Attachment sites for the plus ends of actin filaments (thin filaments)
• M line: Midline.
• Location of proteins that link adjacent myosin II filaments (thick filaments)
to one another
• Dark bands: mark the location of the thick filaments = A bands
• Light bands: which contain only thin filaments and therefore have a lower
density of protein = I bands.
Sarcomeres
Myofibril
• A single continuous stretch of interconnected sarcomeres
• Runs the length of the muscle cell
• More myofibrils in parallel  more force
Striated Muscle Cell Structure
Composed of thick and thin
filaments
• Thick: myosin
• 300 myosin II hexamers
• Thin: actin
• Capped by tropomodulin
(-) and CapZ (+) to
stabilize
• Decorated by troponin
and tropomyosin
• Globular protein (G-actin)
• Form long chains called
F-actin
• In skeletal muscle 2 Factin polymers twist
together
Actin and Myosin Function
Myosin
• Motor protein used by actin
• Sliding filament model
• Most common type of movement
• Myosin is an ATPase
• Converts E released
• from ATP to mechanical E
• 17 classes of myosin with
• multiple isoforms
• Similar structure
• Head, tail, and neck
Sliding Filament Model
Analogous to pulling yourself along a rope
• Actin: the rope
• Myosin: your arm
Sliding Filament Model
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Two processes
• Chemical
• Myosin binds to actin
(Cross-bridge)
• Structural
• Myosin bends
(Power stroke)
Cross-bridge cycle
• Formation of crossbridge, power stroke,
and release
Need ATP to attach and
release
• No ATP  rigor mortis
Sliding filament model
Sliding Filament Model, Cont.
• Myosin is bound to actin in the
absence of ATP and this is the
"rigor" state i.e. gives rigidity to
the muscle
• ATP binds to the myosin causing
the head domain to dissociate
from actin
• ATP is then hydrolyzed causing
a conformational change in the
mysoin head to move it to a new
position and bind to actin
• Pi is released causing the
myosin head to change
conformation again and it is this
movement that moves the actin
• ADP is released
Ca2+ Allows Myosin to Bind to Actin
• Ca2+ binds to TnC
• Reorganization of troponin-tropomyosin
• Expose myosin-binding site on actin
Ca2+ Allows Myosin to Bind to Actin
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Ca2+ levels increase in cytosol
Ca2+ binds to troponin C
Troponin-Ca2+ complex pulls tropomyosin away form
G-actin binging site
Myosin binds to actin and completes power stroke
Actin filament moves
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Sliding Filament Model
Contractile Force
• Depends on sarcomere length: distance
between the Z-disks
• Number of myofibrils
• Number of cells (recruitment)
Isotonic and isometric contraction
Regulation of Contraction
• Excitation-contraction coupling
• Depolarization of the muscle plasma
membrane (sarcolemma)
• Elevation of intracellular Ca2+
• Contraction
• Relaxation when the sarcolemma
repolarizes and Ca2+ returns to resting
levels
Excitation – contraction coupling
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ACH released at the NMJ
Net entry of Na+ initiates a
muscle action potential
AP in T- tubule alters
conformation of DHP receptor
DHP receptor opens Ca2+ release
channels in SR and Ca2+ enters
the cytoplasm
Ca2+ binds to troponin C
Troponin-Ca2+ complex pulls
tropomyosin away form G-actin
binging site
Myosin binds to actin and
completes power stroke
Actin filament moves
Time Course of Depolarization
Time Course of Depolarization
Cause of Depolarization
Myogenic
• Spontaneous
• e.g., Vertebrate heart
• Pacemaker
• Cells that depolarize fastest
• Unstable resting membrane
potential
Neurogenic
• Excited by
neurotransmitters
• e.g., Vertebrate skeletal
muscle
• Can have multiple (tonic)
or single (twitch)
innervation sites
Relaxation
• Repolarization
• Reestablish Ca2+ gradients
• Extracellular
• Ca2+ ATPase
• Na+/Ca2+ exchanger (NaCaX) in reverse
• Intracellular
• Ca2+ ATPase (SERCA)
• Parvalbumin – cytosolic Ca2+ buffer
The Z disk
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The Z disk is complex of proteins
Responsible for anchoring the actin filaments to ensure that the
sacromere will shorten during contraction
Actin filaments are capped at both ends to ensure that the actin will not
depolymerize
Titin-nebulin filament system stabilizes the alignment of thick and thin
filaments in skeletal muscle
Thick filaments are connected at both ends to Z disks through titin
Nebulin is associated with a thin filament from its (+) end at the Z disk to
its other end
Accessory proteins in a sarcomere
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Each giant titin molecule extends from the Z disc to the M line
Part of each titin molecule is closely associated with a myosin thick
filament
The rest of the titin molecule is elastic and changes length as the
sarcomere contracts and relaxes
Each nebulin molecule is exactly the length of a thin filament
The actin filaments are also coated with tropomyosin and troponin and are
capped at both ends. Tropomodulin caps the minus end of the actin
filaments, and CapZ anchors the plus end at the Z disc, which also
contains a-actinin.
Skeletal muscle metabolism
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Muscles require a large source of ATP to allow for contraction and for
transport of Ca2+
ATP requirements are normally met by glycolysis or respiration
Skeletal muscles contain large glycogen stores
Skeletal muscles contain creatine phosphate that generates ATP:
creatine phosphate + ADP = creatine + ATP
Muscles have lots of mitochondria which extend through out the
myofibrils and are red coloured due to a large blood flow and myoglobin
(which stores oxygen)
The breakdown of glycogen stores in muscles can be stimulated by both
Ca2+ and epinephrine
Asynchronous Insect Flight Muscles
• Wing beats: 250 to 1000 Hz
• Fastest vertebrate
contraction: 100 Hz
(toadfish sonic muscle)
• Asynchronous because nervous
stimulation is not synchronized
to contraction
• Due to stretch-activation
• Contracted: Ca2+
insensitive
• Stretched: Ca2+ sensitive
Trans-Differentiation
• Cells with novel properties
• Heater organs
• Electric organs
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