Cellular movement and Muscles. Muscles – general information Vertebrates and many invertebrates have three main classes of muscle Skeletal muscle connect

Cellular movement and Muscles

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 shrinkageFactors

• 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

• 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

• 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 1-2 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 potentialsChannels - 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. The high permeability to Cl- helps repolarize the membrane after an action potential

2) Voltage-gated Na+ channels .

3) Voltage-gated K+ channel - the delayed rectifier K+

4) Voltage gated Ca2+channels - high threshold Ca2+ channels

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



•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

• The link between depolarization and Ca2+ release or excitation-contraction 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.

• 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 close2. Ca2+ is recycled back into the SR through the Ca2+ ATPases3. Ca2+ binds to calsequesterin

General sequence of events

Ca2+ channels and its release

• 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

• 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 F-

actin 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


• Two processes

• Chemical

• Myosin binds to actin (Cross-bridge)

• Structural

• Myosin bends (Power stroke)

• Cross-bridge cycle

• Formation of cross-bridge, 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+ 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

**

**

*

Ca2+ Allows Myosin to Bind to Actin


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

• 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

• 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

• 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

• 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

Muscles – general information again

Vertebrates and many invertebrates have three main classes of muscle•Skeletal muscle•Smooth muscles surround internal organs such as the large and small intestines, the uterus, and large blood vessels•Cardiac muscle: Striated muscle of the heart.

Smooth Muscle - introduction

• Slow, regular contractions• Prolonged contractions• Contribute to many systems• Key differences from skeletal

muscle• Lack sarcomeres (no striations)• No t-tubules• Minimal SR• Gap junctions• Contract in all dimensions• More complex regulation

Smooth muscles - introduction

• Smooth muscle cells have multiple receptors and activation mechanisms

• Smooth muscle cells can be activated by neurotransmitters, hormones, neighbouring cells

• Important: The overall goal is always the same.... change levels of cytosolic Ca2+ to change the degree of contraction.

• Composed of elongated spindle-shaped cells, each with a single nucleus

• Packed with thick and thin filaments but these filaments are not organized into well-ordered sarcomeres and thus smooth muscle is not striated

Smooth muscles – more introduction

• The sarcoplasmic reticulum network is sparse

• Majority of the increase in cytosolic Ca2+ needed for muscle contraction enters the cell via the plasma-membrane Ca2+ channel

• Changes in the cytosolic Ca2+ level occur much more slowly in smooth muscle (seconds to minutes).

• Nerve innervation of smooth muscle cells is from the autonomic nervous system

• Many smooth muscle cells have the ability to spontaneously activate

• Contractions can occur over minutes rather than milliseconds as was seen with skeletal and hundreds of milliseconds as was seen with cardiac cells.

Smooth muscles – more introduction

• Filaments in smooth muscle are gathered into loose bundles, which are attached to dense bodies in the cytosol

• Dense bodies serve the same function as Z disks in skeletal muscle

• The other end of the thin filaments in many smooth muscle cells is connected to attachment plaques.

• Like a Z disk, an attachment plaque is rich in the actin-binding protein alpha-actinin; it also contains a second protein, vinculin, which binds to an integral membrane protein in the plaque and to alpha-actinin

Smooth muscles

Smooth muscle contraction

• Smooth muscle contraction is not controlled by the binding of Ca2+ to the troponin complex as it is in cardiac and skeletal muscles

• Ca2+ control of myosin attachment to the actin is through an intermediate step of Ca2+/calmodulin and it is this that controls contraction in smooth muscle cells

• Calmodulin = intracellular second messenger that binds Ca2+

• Troponin is not found in smooth muscle cells (tropomyosin is)• Caldesmon = regulatory protein on smooth muscle actin. Binds to actin

and prevents myosin from binding actin

Caldesmon and Ca2+/calmodulin

• The activation of smooth muscle myosin can be regulated by caldesmon (CD) which in low Ca2+ levels (10-6 M), binds to tropomyosin and actin and blocks myosin binding to actin

• As Ca2+ levels increase, Ca2+ activated calmodulin binds to caldesmon which releases caldesmon from the tropomyosin/actin complex

• Now myosin is free to bind and move along the thin filaments to contract the cell

• Phosphorylation by several kinases, including MAP kinase, and dephosphorylation by phosphatases also regulate caldesmon’s actin-binding activity

*

**

Myosin light chain kinase (MLCK) and Ca2+/calmodulin

• In vertebrate smooth muscle, phosphorylation of the myosin regulatory light chains on site X by Ca2+-dependent myosin LC kinase activates contraction

• At Ca2+ concentrations < 10-6 M, the myosin LC kinase is inactive

• A myosin LC phosphatase, which is not dependent on Ca2+ for activity, dephosphorylates the myosin LC, causing muscle relaxation

Contraction – simple

• Intracellular Ca2+ increase and Ca2+ is released from the SR

• Ca2+ binds to calmodulin (CaM)• Ca2+ - calmodulin complex activates

MLCK• MLCK phosphorylates light chains in

myosin heads and increases myosin ATPase activity

• Active myosin crossbridges slide along actin and create muscle

*

**

**

*

Relaxation - simple

• Free Ca2+ in cytosol decreases when Ca2+ is pumped out of the cell or back into the SR released from the SR

• Ca2+ unbinds from calmodulin • Myosin phosphatase removes

phosphate from myosin, which decreases myosin ATPase

• Less myosin ATPase activity results in decreased muscle tension

**

*

*

Regulation of smooth muscle contraction • The major means that control smooth muscle contraction is controlled

is through changes in resting membrane potential• Depolarization causes a greater increase in cytosolic Ca2+ and thus

greater contraction• Hyperpolarization causes a reduced amount of cytosolic Ca2+ and thus

relaxes the muscle cell• However it is important to note that release of Ca2+ from internal stores

may also lead to greater contraction through G protein mediated cascades that have nothing to do with changes in membrane depolarization.

Norepinephrine and epinephrine

• Depending on the type of receptor norepinephrine and epinephrine can have different results on the smooth muscle cell

• Epinephrine bound to beta-adrenergic receptors on smooth muscle cells of the intestine causes them to relax

• Epinephrine also binds to the alpha2-adrenergic receptor found on smooth muscle cells lining the blood vessels in the intestinal tract, skin, and kidneys

• Epinephrine bound to alpha2 receptors causes the arteries to contract (constrict), reducing circulation to these organs

Acetylcholine and Nitric Oxide

• ACH is released by autonomic nerves in the walls of a blood vessel, and it causes smooth muscle cells in the vessel wall to relax

• ACH acts indirectly by inducing the nearby endothelial cells to make and release NO, which then signals the underlying smooth muscle cells to relax.

• Regulation of contractility of arterial smooth muscle by NO and cGMP:

• NO synthesized in endothelial cells diffuses locally through tissue and activates guanylate cyclase in nearby smooth muscle cells

• The resulting rise in cGMP leads to the relaxation of the muscle and vasodilation.

• Cont…..

Acetylcholine and Nitric Oxide

• Schematic diagram of the structure of soluble guanylate cyclase

• Binding of NO to the heme group stimulates the enzyme’s catalytic activity, leading to formation of cGMP from GTP.

More about Nitric Oxide

• Catalyzed by the enzyme NO synthase from arginine

• Rapidly diffuses out of the cell into neighboring cells

• Very short half life (5-10 seconds) - so acts only locally

• In many target cells, NO binds to iron in the active site of the enzyme guanylyl cyclase, stimulating this enzyme to produce cyclic GMP.

• The effects of NO can occur within seconds, because the normal rate of turnover of cyclic GMP is high

• Increased cGMP activates a kinase that subsequently leads to the inhibition of calcium influx into the smooth muscle cell, and decreased calcium-calmodulin stimulation of myosin light chain kinase (MLCK).

• Decreases the phosphorylation of myosin light chains, thereby decreasing smooth muscle tension development and causing vasodilation.

Other regulators

Nitroglycerine

• Has been used for about 100 years to treat patients with angina (pain resulting from inadequate blood flow to the heart muscle)

• Converted to NO, which relaxes blood vessels

• This reduces the workload on the heart and reduces the oxygen levels needed by the heart muscle.

Viagra

• The drug sildenafil [Viagra] inhibits this cyclic GMP phosphodiesterase and increases the amount of time that cyclic GMP levels remain elevated.

• The cyclic GMP keeps blood vessels relaxed and in certain parts of the male anatomy blood pools and the resulting effect has sales of Viagra soaring. It is interesting to note however that Viagra is not specific to the penis it will affect cGMP levels throughout the body and can have some interesting side effects.

More about ACHTissue Effects of ACh

Vasculature (endothelial cells) Release of endothelium-derived relaxing factor (nitric oxide) and vasodilation

Eye iris (pupillae sphincter muscle) Contraction and miosis

Ciliary muscle Contraction and accommodation of lens to near vision

Salivary glands and lacrimal glands Secretion—thin and watery

Bronchi Constriction, increased secretions

HeartBradycardia, decreased conduction (atrioventricular block at high doses), small negative inotropic action

Gastrointestinal tractIncreased tone, increased gastrointestinal secretions, relaxation at sphincters

Urinary bladder Contraction of detrusor muscle, relaxation of the sphincter

Sweat glands Diaphoresis

Reproductive tract, male Erection

Uterus Variable, dependent on hormone influence

Cardiac muscle

Cardiac muscle – general info

• Many similar properties to skeletal muscles but there are some important differences

• Hearts of course vary greatly in size, shape and complexity from animal to animal - ranging from insects with a simple tube that pumps blood or hemolymph around an open circulatory system to our closed circulatory system and a four chambered heart

Cardiac muscle – general info

• The heart contains pace-maker cells that produce the depolarization and action potentials to drive cardiac cell contraction

• Heart contraction is not neuronally driven but self-driven or myogenically.

• Each muscle cell is a single cell not multinucleate like skeletal muscle

• Like skeletal muscle cells each cell contains multiple myofibrils and in the cases of higher vertebrates an extensive sarcoplasmic reticulum

• Cardiac muscle cells are linked to each other with gap junctions

Cardiac muscle – general info cont.

• There are different types of cardiac muscle cells ranging from the pacemaker cells in the sinoatrial node to the atrial and ventricular cells that produce the contraction of the heart chambers

• The action potential in cardiac cells is quite different from skeletal muscle and neuronal action potentials in that voltage-gated Ca2+ channels play a much larger role

• The mechanism of triggering the Ca2+ release channel in the sarcoplasmic reticulum is not the same as in vertebrate skeletal muscle cells

Pacemaker Cells

• Derived from cardiac muscle cells• Differences from most cardiac muscle

• Small with few myofibrils, mitochondria or other organelles• Do not contract• Have unstable resting membrane potential (pacemaker

potential) that slowly drifts upwards until it reaches a threshold and activates and action potential

Cardiac muscle channels and action potentials

Pumps and transporters • Na+/K+ ATPase pump - to establish the electrochemical gradients of

Na+ and K+

• Ca2+ ATPase pump - uses energy from ATP to remove 2 Ca2+ from the inside to the outside of the cell or into the sarcoplasmic reticulum to ensure that internal Ca2+ concentrations remain low (10-7 mM internal)

• Some cardiac cells (i.e. lower vertebrates, invertebrates) do not have an extensive sarcoplasmic reticulum and thus most of the Ca2+ that is used to trigger contraction is from extracellular sources

• 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+.

Excitation-contraction coupling

Cardiac muscle channels and action potentials

Channels• leak channels - leak K+channel

• voltage-gated Na+ channels

• voltage-gated K+ channel - the delayed rectifier K+ channel

• voltage gated Ca2+ channels - Note: In cardiac cells the Ca2+ channel plays a much greater role during the action potential- High threshold Ca2+ channels, called L channel or DHP (dihydropyridine channel)

Action potential in ventricular (and atrial) cardiac cells

• Resting potentials in these cells is set by a large K+ permeability due to a combination of the leak K+ channel and a voltage-gated K+ channel (called the inward rectifier K+ channel) that is open at rest

This means that rest is very close to EK+

• The rising phase of the action potential is set by the cardiac voltage-gated Na+ channel


• As the voltage-gated Na+ channels produce the rising phase and then start to inactivate two channels will now be opening, the delayed rectifier K+ channel and the voltage-gated Ca2+ channel (L or DHP channel)

• There are many Ca2+ channels in these cells and thus this channel dominates the membrane potential producing a long plateau of depolarization

• This plateau is a balance between the open Ca2+ channels and the open K+ channels

• Ca2+ channel only slowly inactivates and thus this plateau can persist for 100-200 msec.


• Voltage-gated Ca2+ channel inactivates and the voltage-gated K+ channels will now dominate and the membrane potential will repolarize to rest (EK+ in these cells).

• Then the voltage-gated K+ channels will close, the voltage-gated Na+ channels will switch from the inactive to the closed state and the membrane is set back at 4) ready to fire again.

• The long Ca2+ plateau allows Ca2+ inside the cell to elevate enough to generate contraction in the case of those cardiac cells that rely on external Ca2+ sources.

Action potential in sinoatrial cardiac cells

• Sinoatrical cells have the ability to spontaneously fire action potentials in a repeated fashion without any external influence.

• These cells are the pacemaker cells of the heart and once an action potential fires in these cells it is propagated via gap junctions to other regions of the heart first to the atrial cells and then eventually making it to the ventricular cells.

Action potential in sinoatrial cardiac cells

Very similar to the ventricular cardiac cells with a few major exceptions

• Do not have a stable rest.

• The action potential is driven by the voltage-gated Ca2+ channel in most SA cells

• The rising phase is due the opening of the voltage-gated Ca2+ channel

• As the Ca2+ channel inactivates the membrane is repolarized by the delayed rectifier K+channel as in other excitable cells

• Spontaneously depolarize once the delayed K+ channel has closed

• Due to presence of an ion channel that is activated by hyperpolarization – the funny channel.

The funny channel

• Activated by hyperpolarization• As the membrane repolarizes after the action potential the threshold for

opening of the funny channel is reached at about -50 mV• The channel opens and allows Na+ to preferentially flow into the cell• Also called the HCN channel or hyperpolarization, cyclic nucleotide

gated ion channel• cAMP can have dramatic influences on this channel and shift its

threshold of activation from -50 mV to -40 mV.• The funny channel actually looks very much like a voltage-gated K+

channel but has differences in its pore to allow Na+ influx and in the voltage sensing/opening mechanism.

Putting it all together……

• Located in the right atrium at the superior vena cava is the sinus node (sinoatrial or SA node) which consists of specialized muscle cells

• The SA nodal cells are self-excitatory, pacemaker cells• They generate an action potential at the rate of about 70 per minute in

humans (your heart beat)• From the sinus node, activation propagates throughout the atria, but

cannot propagate directly across the boundary between atria and ventricles

• This boundary serves to ensure a delay between the activation of the atria and the ventricles

• The atrioventricular node (AV node) is located at the boundary between the atria and ventricles

• In a normal heart, the AV node provides the only conducting path from the atria to the ventricles

Putting it all together……

• Propagation from the AV node to the ventricles is provided by a specialized muscle cells called the bundle of His conduct the signal system

• Further down the bundle separates into two bundle branches which travel along each side of the septum, constituting the right and left bundle branches.

• Even more distally the bundles split into Purkinje fibers that branch and contact the inner sides of the ventricular walls.

• From the inner side of the ventricular wall, these activation sites cause the formation of a wave of depolarization which propagates through gap junctions between the ventricular cells toward the outer wall

• After each ventricular muscle region has depolarized, repolarization occurs.

Electrocardiogram - ECG

• P wave - an impulse is generated at the sinoatrial node and spreads across both atria, causing them to contract

• Delay: The Fibro-fatty atrioventricular groove insulates the ventricles from the atrial impulse

• The AV node is the only normal gateway of conduction to the ventricles

• QRS wave - The impulse travels down the AV bundle and it's branches and reaches the Purkinje fibers

• The ventricles are stimulated to contract

• T wave - correlates with repolarization of the ventricles.



Increasing the heart rate

Epinephrine and norepinephrine• Released from the sympathetic nervous system• Epinephrine and norepinephrine are synthesized and released

into the blood by the adrenal medulla, an endocrine organ• Epinephrine and the related norepinephrine are all synthesized

from tyrosine and contain the catechol moiety; hence they are referred to as catecholamines

• Nerves that synthesize and use epinephrine or norepinephrine are termed adrenergic

• Adrenergic receptors: bind epinephrine and norepinephrine. Because different receptors are linked to different G proteins, the activation leads to different signal transduction cascades

• More Na+ and Ca2+ channels open• Rate of depolarization and action potentials increase

Increasing the heart rate cont.

Epinephrine and norepinephrine cont….

• In sinoatrial cells: norepinephrine binds to the b-adrenergic receptor which is a G protein associated membrane receptor

• This triggers a signal transduction cascade outlined below that activates the G protein (Gs - stimulates) that activates adenylate cyclase to produce cAMP.

• Beta-blockers: Drugs which are used to slow heart contractions in the treatment of cardiac arrhythmia and angina, are beta1-adrenergic receptor antagonists

• They bind the beta1-adrenergic receptor to block the receptor and thus slow heart contraction

• Cardiac muscle cells possess beta1 adenergic receptors

Decreasing the heart rate

Acetylcholine: released from parasympathetic nervous system

• Muscarinic acetylcholine receptor: a G protein associated receptor. The G protein activated in this case is a Gi subunit that inhibits adenylate cyclase

• More K+ channels open• Pacemaker cells hyperpolarize• Time for depolarization takes longer

Modulating the funny channel

• Through G protein coupled receptors various hormones/neurotransmitters or drugs can increase or decrease the heart rate by simply increasing or decreasing the ability of the funny channel to open.


• Binding of hormone (e.g., epinephrine, glucagon) to a Gs protein coupled receptor

• Gs protein relays the hormone signal to the effector protein, ie adenylyl cyclase

• Gs cycles between an inactive form with bound GDP and an active form with bound GTP

• Dissociation of the active form yields the Gsalpha · GTP complex, which directly activates adenylyl cyclase

• The increase in cAMP physically binds to the funny channel and makes the channel open more easily

• Funny channel will open sooner during the repolarization stage of the sinoatrial action potential and a second action potential will be triggered sooner

• Increase heart rate.

Modulation of Ca2+ channel

Epinephrine:

• Causes an increase in cAMP that stimulates PKA (protein kinase A) which in turn phosphorylates the voltage-gated Ca2+ channel (L channel)

• Results in a protein conformational change that enhances the channels activity

• This new conformation of Ca2+ channel opens more readily (i.e. less time between action potentials) and opens for longer (i.e. more Ca2+ flow into the cell = greater [Ca2+] intracellular = greater contraction).

• Stimulates glycogen breakdown in skeletal muscles

Caffeine (mmmmhhh):

• Blocks the activty of phosphodiesterases. Phosphodiesterases break down cyclic nucleotides. cAMP levels remain elevated and thus the funny channel continues to open more readily.

• Affects the Ca2+ release channel or ryanodine receptor such that more Ca2+ is released through the channel. Therefore heart contractions are stronger in the presence of caffeine as well


• Acetylcholine works to block any rise in cAMP and reduces cAMP levels in the cell

• Therefore the funny channel will now not open so readily and the slow depolarziation of the membrane will occur later thus resulting in a longer time to generate a second action potential.

Modulating the voltage-gated K+ channels

• Acetylcholine-induced opening of K+ channels in the heart muscle plasma membrane

• Binding of ACH by muscarinic ACH receptors triggers activation of a transducing G protein by catalyzing exchange of GTP for GDP on the alpha subunit

• The released beta/gamma subunit then binds to and opens a K+ channel• The increase in K+ permeability hyperpolarizes the membrane, which

reduces the frequency of heart muscle contraction• Activation is terminated when the GTP bound alpha subunit is

hydrolyzed to GDP and Galpha · GDP recombines with Gbeta/gamma.

Modulating the voltage-gated K+ channels

Application of acetylcholine (or muscarine) to frog heart muscle produces, after a lag period of about 40 ms (not visible in graph), a hyperpolarization of 2 3 mV, which lasts several seconds

From receptor to control of muscle cell contraction


• The cardiovascular system is highly regulated so that there is always an adequate supply of oxygenated blood to the body tissues under a wide range of circumstances

• There are receptors that respond to the degree of blood pressure and provide mechanical (barosensory) information about pressure in the arteries system

• There are receptors that provide information about the level of oxygen and carbon dioxide in the blood

• These sensory systems provide input to the respiratory control centers of the brain which in turn control the parasympathetic and sympathetic nerves that will control the heart, blood vessels and diaphragm muscles for breathing.


• We will concentrate only on the chemoreceptors which are located primarily in the aortic and carotid bodies

• These are small, specialized organs located at the bifurcation of the common carotid arteries (some chemosensory tissue is also found in the aorta)

• The chemoreceptors in the carotid bodies and aorta provide information about the partial pressure of oxygen (PO2) and carbon dioxide (PCO2) in the blood

• This information is relayed by second order neurons to the hypothalamus and other regions in the brainstem

• This information about blood gas levels works in a reflex to modulate the autonomic nervous system to control smooth and cardiac muscles


• The carotid chemosensory cells detect levels of PO2 in the blood by simply depolarizing in response to decreased levels of oxygen

• The mechanism appears to be an O2 sensitive K+ channel, that in the presence of normal levels of PO2 is open

• Therefore the Vm is close to EK+

• As oxygen levels drop the K+ channel closes and Vm depolarizes allowing the voltage-gated Ca2+ channel to open and to trigger vesicle fusion and neurotransmitter release


• PO2 levels can have a direct effect on smooth muscles around blood vessels

• Many of these cells have K+ channel that is inhibited by ATP

• As PO2 drops so does respiration and ATP production

• This reduction in ATP results in the opening of K + channels and the inhibition of smooth muscle contraction

• This results in the relaxation of the smooth muscles the relaxation of the blood vessels and the increase blood flow into the tissue that is experiencing reduced PO2

• Conversely an increase in PO2 results in greater inhibition of the ATP

sensitive K + channels and thus a greater degree of depolarization

• More Ca 2+channels are open and thus there is greater cytosolic Ca2+ levels, greater degree of smooth muscle contraction

• This causes the blood vessel to constrict (vasoconstriction) and less PO2 transfer to the surrounding tissues.


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