6.5 Muscle and Innervation Flashcards
What is the main function of muscle cells?
Movement - the tissues are made up of elongated contractile cells
What are contractile elements and where can they be found?
- Myosin
- Actin
They are found in nearly all cells (used to move, change shape or allow the intracellular movement of organelles)
What makes muscle cells unique?
- Permanently orientated contractile machinery
- Allows directional movements/contractions that are appropriate for the tissue
- Able to stretch beyond their resting length
- Return to their resting state (elasticity)
- Increase in size (hypertrophy) or number (hyperplasia) or both
What are the three types of muscle?
- Skeletal
- Cardiac
- Smooth
Briefly summarise the three types of muscle.
Skeletal: - Attached to bones crossing joints, controls the eye - Striated appearance - Multinucleate, peripherally orientated - Somatic/voluntary innervation Cardiac: - Found only in the heart - Striated appearance - Usually only uninucleate, centrally located - Autonomic/involuntary innervation Smooth: - Found in visceral organs, incl blood vessels and glands - Non-striated - Uninucleate, centrally located - Autonomic/involuntary innervatioin
What is the sarcolemma?
Muscle cell membranes
- Bordered externally by a basal lamina and collagen + reticular fibres
What is the sarcoplasm?
Muscle cell cytoplasm
What is the sarcoplasmic reticulum?
Muscle cell smooth endoplasmic reticulum (SER)
What are the sarcosomes?
Muscle cell mitochondria (only occasionally used)
What is a sarcomere?
A contractile unit within a muscle cell
How do muscles develop and from where are they derived?
- From the myotomes (derived from dermomyotomes) of the somites, therefore mesodermal tissue
- Process is called myogenesis
- Somite cell differentiation is induced by Shh from the notochord
- Dermomyotome expresses Pax3+ and Pax7+ (positive) cells, which are myoblasts that can then divide and migrate (Pax3/Pax7 positive cells are able to release muscle precursor cells during development, e.g. myoblasts and satellite cells)
- Myoblasts withdraw from the cell cycle and express transcription factors MyoD and Myf5
- > these activate genes for muscle cells
How are multinucleate cells of skeletal muscle formed?
- Through the fusion of myoblasts (multinucleate fibres produced by fusion of myoblasts are called myotubes)
- Myotubes then develop into mature tubular cells (multinucleate syncytium)
- In skeletal myocytes, striated structure is seen
What are all of the different terms for muscle cells?
- Myocytes
- Muscle fibres
- Muscle cells
- Myofibrils
What stain is used to recognise striated muscle?
Paraffin stains
How can skeletal muscle be identified?
Aka striated or voluntary muscle
- Cells are cylindrical
- Multinucleate, nuclei are elliptical and located peripherally within the cell
- Cytoplasm shows alternating light and dark patterning using paraffin stains in longitudinal sections
- > striations are the result of overlapping contractile elements
Where are the satellite cells located in skeletal muscle?
On the periphery/outside of cells, allow small amount of regeneration
What sort of blood supply is available to striated muscle cells?
- Rich capillary network formed
- Blood vessels penetrate muscles
- Vessels run parallel and between various muscle fibres
What are the gross structures of skeletal muscle in increasing size?
- Sarcomere
- Microfilaments
- Myofibrils
- Muscle fibre
- Myocyte
- Muscle tissue
What is the epimysium?
- Sheath of connective tissue that surrounds individual muscles
- Extends inwards to form the septa (dividing walls) of the perimysium
What is the perimysium?
- Sheaths of connective tissue that surround fascicles of muscle
- Linked to the epimysium
What are fascicles?
Bundles of tissue surrounded by perimysium that contains hundred/thousands of muscle fibres
What is the endomysium?
- Sheath of connective tissue around individual muscle fibres
- Contained within the fascicles
What do the surrounding connective tissue layers allow for?
- Entry and exit of arteries, nerves, lymphatics and veins
- Freedom of movement is allowed between fascicles and muscle fibres (connective tissue reduces friction)
- Continuity of connective tissue allows for all forces generated to be transmitted to the tendons
Where do skeletal muscles attach to?
- Bone via tendons (strong structures of dense regular connective tissue, continuous with the various sheaths surrounding muscles)
- Tongue and pharynx muscle attach to investments of connective tissue instead of bone, allowing their movements
- Smaller fascicles within long muscles may also end in tapering points attaching to the connective tissue sheaths within the muscle tissues
What does the striated pattern in myofibrils show?
- Repetitive contractile units/sarcomeres along the tissue
- Alternating light and dark bands caused by the presence of different tissues
- Only seen in longitudinal sections (transverse sections have a stippled appearance, showing the cut ends of myofibrils
Where are satellite cells located and what is their function?
- Located between the sarcolemma and the basal lamina (occasionally)
- Have myogenetic potential i.e. are able to repair small amounts of damage to the tissue
- > may form new muscle fibres following injury
- > contribute nuclear DNA during hypertrophy
What is contained within the sarcoplasm?
- Myofibrils
- Golgi apparatus (near nucleus)
- Mitochondria (near nucleus)
- Nuclei (periphery)
- Sarcoplasmic reticulum (with T-tubules)
- Lipids
- Glycogen
What are T-tubules?
Transverse tubules, invaginations of the sarcolemma into the myocyte to extend past the myofibrils
- Found between myofibrils
- Adjacent to sarcoplasmic reticulum (triad structure, SR-TT-SR)
What are the two types of contractile unit in myofibrils?
- Thick filaments (myosin proteins)
- Thin filaments (actin proteins)
What are the Z lines?
- Delineate sarcomeres (a sarcomere is a z line to a z line)
- Discs to which actin filaments anchor (ross-linked by alpha actinin)
- Myosin filaments are linked to Z lines by titin
- In the I bands
What are A bands?
- Bands containing both thick and thin filaments (actin and myosin)
- Appear darker under the microscope
- Size remains constant during contraction
- Remember using dArk
What are I bands?
- Bands containing only thin filaments (actin)
- Appear lighter under the microscope
- Width decreases during contraction
- Remember using LIght
What are H bands?
- Lighter regions within the A band where only myosin filaments are present
- Will get shorter during contraction
- Found using electron microscopy
What are M lines?
- Where the myosin/thick filaments attach
- Found using electron microscopy
- Found in the H band
What is the sliding filament theory?
- First proposed by Huxley who viewed muscle contraction under a light microscope
- This is a proposed method of contraction where the interdigitating myosin and actin filaments are able to slide past each other and cause the muscle to contract
- > actin filaments slide along the myosin
- Sarcomeres in series allow the net effect of shortening the entire muscle
- The huge number of sarcomeres allows the net generation of a lot of power within the muscle
- Z lines/discs are brought closer together
- The filaments do not shorten
What happens to sarcomeres during muscle stretching?
- Sarcomeres extend in length
- But are immediately able to elastically recoil once tension is released
What is the arrangement of thick and thin filaments in sarcomeres?
One thick filament is surrounded by 6 equidistant thin filaments
What is the structure of thick filaments/myosin proteins?
- Myosin II = 6 polypeptide chains twisted to form a fibre helix
- Has globular heads
- > these have ATP activity and bind to actin
What is the structure and features of components making up the thin filaments?
Actin:
- G-actin, a globular protein that polymerises into polymeric (many repeated subunits) fibre
- Contains a myosin binding site
Tropomyosin:
- Fibre-like protein
- Wraps helically around the thin g-actin filament
- Blocks attachment site for myosin on actin in relaxed muscle, so prevents contraction
-> no myosin cross bridges are able to be formed
Troponin:
- Three forms, TnT, TnC and TnI
-> TnT binds to tropomyosin
-> TnC binds to calcium/is calcium-sensitive
-> TnI is inhibitory, binds to actin
- When calcium ions bind to TnC, conformational change is induced in the whole structure
-> displaces tropomyosin from the active sites of actin and so allows contraction
What is titin?
Aka connectin
- Links myosin to the Z discs, arranging them to be in the correct position relative to the actin filaments
- Molecular spring for passive elasticity of muscle
- Maintains sarcomere structure
- Largest known protein
- Stretches from Z disc to Z disc
What is alpha actinin?
- CapZ
- Anchors thin filaments to sarcolemma
What is nebulin?
- Actin-binding proteins localised to the thin filament
- > regulates thin filament length
How is rigor mortis initiated?
- Contraction is ATP dependant
- Once ATP isn’t available, the thick and thin filaments are unable to dissociate (calcium is also slowly released from the sarcoplasmic reticulum, enabling the filaments to bind)
- Myosin filaments remain attached to the actin filament until the muscle begins to decompose
- This permanent contraction is what results in rigor mortis/the stiffening of the body after death
- Time taken for process to occur is actually highly dependant upon conditions and location so this isn’t a reliable method of determining a time of death but can provide some indication
What are the two fibre types?
Type I: Slow twitch
- Aerobic
- High number of mitochondria
- Extensive blood supply
- High endurance
- High myoglobin (last-ditch O2 storage) concentrations
- Succinate dehydrogenase stains more intensely
Type II: Fast twitch
- Anaerobic
- Abundant glycogen
- Provides short bursts of energy
- Low endurance
- Both IIa and IIb forms, IIa is an intermediate between the two types, IIb is the classic ‘fast’ twitch muscle
- Low myoglobin concentration
- mATPase stains more intensely in type II
How does skeletal muscle contract in a coordinated fashion?
- Coordination of the sarcoplasmic reticulum and the t-tubules
- T-tubules surround each myofibril
- Sarcoplasmic reticulum is associated with t-tubules (electromechanical coupling), terminal cisternae located near to the invaginations
- Triad structure formed (SR-TT-SR)
- > impulse is conveyed from T-tubules to cisternae
- > triggers calcium ion release
- > calcium binds to the troponin and allows the formation of myosin/actin cross-bridges
What is a neuromuscular junction (NMJ)?
- Where somatic nerve fibres terminate on skeletal muscle fibres
- Responsible for initiation of action potentials across the cell’s surface
What is the ratio of motor neurons:muscle fibres?
One motor neuron can supply multiple muscle fibres
But only there will only be one NMJ per muscle fibre/each fibre is only innervated by one neuron
What are the benefits of having multiple muscle fibres being innervated by the same nerve?
- Motor unit is formed
- Allows contraction in unison
- A lot faster and more reliable than having to innervate all of the fibres separately
What is the neurotransmitter at a neuromuscular junction?
Acetylcholine (ACh)
How does excitatory synaptic transmission occur at the NMJ?
- Action potential causes opening of voltage gated calcium channels at the presynaptic neuron
- Rapid influx of calcium ions activate SNARE proteins and triggers vesicles to release their neurotransmitters into the synaptic cleft/fuse with the membrane of the axon
- ACh binds to receptors on the motor end plate, opening channels which allow influx of ions (nicotinic ACh receptors, ligand gated)
- This triggers an end-plate potential at the motor end plate, which travels to the sarcolemma where the an action potential is triggered and propagated
What is the DGC?
Dystrophin-associated glycoprotein complex
- Connects cytoskeleton of muscle fibre to the surrounding extracellular matrix, through the cell membrane
- Allows contraction of muscle fibres to affect the actual cell and surrounding tissues
- Mutations in DGC-associated proteins results in muscular dystrophy
What is dystrophin?
- Rod-shaped cytoplasmic protein
- Vital part of the DGC (dystrophin-associated glycoprotein complex)
- Maintains mechanical integrity of the cell during contraction
- Anchors cytoskeletal elements
- Gene is the largest known human genome
- What is Duchenne’s muscular dystrophy?
- Fatal x-linked disorder
- Mutations in dystrophin gene result in a reading frame shift mutation
- Unstable form of dystrophin produced
- Absence of dystrophin results in impairment of the sarcolemma
- Symptoms include muscle weakness and wasting
- Potential treatments should be removing a section of the genome to encourage alternate splice sites, resulting in a less severe form of dystrophy (more manageable, not fatal - can also upregulate utrophin)
How can neuromuscular diseases effect muscle control?
- Can cause spasticity (constant contraction)
- Can cause paralysis (loss of function)
Leads to problems with movement and or motor coordination
Where is cardiac muscle derived from?
- Defined mass of splanchnic mesenchyme and myoepicardiaum mantel
- > these give rise to the epicardium and the myocardium
- Endocardium derives from vascular endothelial progenitors
What are the features of cardiac muscle?
- Striated, mononucleate/binucleate cardiomyocytes
- Specialised in branches
- Arranged in layers known as laminae
- Abundant mitochondria and extensive t-tubule network
- Intercalated discs between cells (complexes of cell connections that allow cardio cells to be coupled)
How are cardiomyocytes linked?
- Structurally and functionally
- Intercalated discs interface between adjacent cardiac muscle cells (supports synchronised contraction)
What are the three main junctional specialisations within intercalated discs?
- Fascia adherens (hemi Z bands), anchoring sites for actin filaments
- Macula adherens (desmosomes) binds cells together
- Gap junctions, provides continuity between adjacent cells, allows ions to pass through (communications)
- > electrochemical potentials directly conducted between cells through these
- Highly resistant to fatigue, many mitochondria and good blood supply
Allows cardiac myocytes to be an functional electrical syncytium
What is cardiac excitation contraction coupling?
- Follows membrane depolarisation
- Release of calcium ions from sarcoplasmic reticulum
- > these cause calcium induced myofilament activation
- Myocardial contraction activated by:
- > transient rise in cytosolic free calcium concentration
- > change is to about 1 mM from resting diastolic conc of 0.1uM
What are Purkinje fibres?
- Modified cardiac muscle cells
- Specialised myocardial fibres
- > located in inner ventricular walls
- > run beneath endocardium
- Larger than cardiomyocytes
- Carries impulse originating in SAN ventricle
- > heart’s own conduction system
- > enables synchronised conduction of ventricles
- > essential for maintaining consistent heart rhythm
- Pale-staining cytoplasm
- > rich in glycogen and mitochondria
- > contain limited contractile elements
- > no t-tubule system
- What is myocardial infarction?
- Heart attack
- Result of loss of blood supply to the heart/vessels supplying heart become blocked
- > causes cardiac tissue to die
- May be possible to repair scar tissue with stem cells in the future
- iPS cells can be used to differentiate into cardiomyocytes under certain conditions
- What is Dilated Cardiac Myopathy (DCM)?
- Heart enlarges and pumps efficiently
- Heritable or idiopathic form of heart failure
- Mutations in cytoskeletal proteins disrupt intercalated disc morphology
- Junctions between cells disrupted, also effecting contractile function
- Causes enlargement of heart chambers and thinning of ventricular walls
- Weakens heart pumping ability
- What is Hypertrophic Cardiac Myopathy (HCM)?
- Autosomal dominant familial disorder
- Sarcomeric proteins are mutated
- > causes defective contraction
- Abnormal growth and hypertrophy of cardiac muscles
- Leads to ventricular wall thickening
What are the features of smooth muscle?
- Elongated
- Spindle shaped (fusiform, elongated and taper at either end)
- Nonstriated
- Mononucleate cells
- Enclosed by the basal lamina and network of reticular fibres
- Connective tissue layers are thinner than seen in skeletal and therefore less easy to view
- Have gap junctions to allow communication between cells
How are filaments aligned in smooth muscle cells?
- Not aligned in regular arrays
- Actin and myosin form a lattice like network but contract via similar sliding filament methods as striated muscle
What are focal densities in smooth muscle cells?
- Attachment junctions
- Attach actin to the sarcolemma
What are dense bodies in smooth muscle cells?
- Attach intracellular actin filaments to each other
- Functionally analogous to/the same as Z-lines in striated muscle
- Maintain alignment of thin filaments
What are calveoli in smooth muscle cells?
- ‘Cave-like’, small dips in the membrane where proteins are concentrated to aid in cell signalling, increases SA
- Involved in transport
- Analogous to t-tubule system in skeletal muscle fibres
- Permanent structures involved in fluid and electrolyte transport (pinocytosis)
- Transmits depolarisation signal
In what ways do gap junctions couple adjacent smooth muscle cells?
- Chemically
- Electrically
Facilitate the movement of chemicals (e.g. calcium ions) or action potentials between smooth muscle cells
Where is smooth muscle found?
- Lining viscera in the gut, bladder, uterus, respiratory system and blood vessels
- Arranged loosely e.g. in the uterus or regularly e.g. in the bowel
How can smooth muscle cells be arranged?
- Single unit, unitary function is enabled by electric coupling via gap junctions, allows muscle to behave as a syncytium (seen in GI tract and bladder)
- Multi-unit, each cell is isolated and stimulated independently to allow greater control (seen in iris of the eye)
What is a syncytium?
- A single cell or cytoplasmic mass containing multiple nuclei
- Through connection of cells, they can behave as a syncytium and produce a highly coordinated response
- What are the two types of smooth muscle in the gut?
- Circular (runs around the gut)
- Longitudinal (runs along the gut)
Together, they cause peristalsis - Circular causes segmentation through wave-like contractions
- Longitudinal causes propulsion
What proteins are present in smooth muscle cells?
- Myosin
- Actin
- NO TROPONIN
- Tropomyosin (binds to and stabilises actin)
What regulates contractions of smooth muscle?
- Phosphorylation of the myosin light chains by myosin light chain kinases (MLCKs)
How does smooth muscle contraction occur?
- Extracellular calcium ions are released from the caveolae
- Calcium ions bind to calmodulin
- Calcium-calmodulin complex then activates theh myosin light chain kinases (MLCKs)
- MLCKs phosphorylate one of the myosin light chains known as the regulatory chain and increases ATPase action
- Phosphorylated light chain unmasks myosin’s active binding site
- Permits the interaction between actin and myosin that allows contraction
How are smooth muscles innervated?
- Autonomic nervous system doesn’t form NMJs
- Instead has swellings at terminal end of the axon known as varicosities
- These are loosely-formed motor units and packed with neurotransmitter-filled vesicles
- Neurotransmitters are released into the synaptic cleft at the varicosities
- Visceral pacesetter cells are also able to spontaneously trigger action potentials and muscle contraction
- What causes asthma?
- Prolonged and sustained contraction of bronchial smooth muscle
- Can be treated by salbutamol, beta-2 adrenergic receptor agonist causing smooth muscle relaxation
- What proliferative abnormalities can smooth muscle undergo and what can this cause?
- Hyperplasia (cell proliferation)
- Hypertrophy (cell enlargement)
- What causes hypertension?
- Contraction of smooth muscle
What regenerative ability does smooth muscle have?
- Greatest capacity to regenerate of all muscle types
- Retain the ability to divide
- > can increase their number in this way e.g. in pregnant uterus
- New cells can be produced by division of pericytes hat line some small blood vessels
- Smooth muscle can also undergo hypertrophy
What is the function of tonic skeletal muscles?
They are antigravity/postural muscles
What is excitation-contraction coupling (ECC)?
How depolarisation/the arrival of a nervous impulse results in a muscle contraction
What type of ECC is seen in skeletal muscle?
Electromechanical coupling
- Lining the T-tubules are voltage sensitive proteins (voltage sensitive L type calcium ion channels, e.g. dihydropodine (DHP) receptors) that change conformationally when depolarised
- As the DHP receptors are physically touching the calcium release channels on the sarcoplasmic reticulum (e.g. Ryanodine (RyR1) receptors), the conformational change is also induced in the RyR1
- The RyR1s then allow the release of calcium from the sarcoplasmic reticulum which will then go on to activate contraction
- Release of calcium ions from SR is not dependant upon calcium influx, as it is in cardiac muscle/calcium induced calcium release (CICR) is not necessary
What is the cross bridge cycle?
- Myosin head forms cross-bridge with actin active site
- ATP binds to the myosin head causing the dissociation of the actin-myosin complex
- ATP is hydrolysed to form ADP + Pi, causing the myosin head to return to its resting position
- Cross bridge forms between myosin head and the next actin active site
- Pi is released and myosin heads change their conformation resulting in the power stroke, filaments slide past each other
- ADP is released so another ATP molecule can bind and the process can repeat
How is contraction terminated in skeletal muscle cells?
- Calcium ions are removed from the cytoplasm by various exchanged proteins and pumps
- SERCA is the most important, sarco-endoplasmic reticulum Ca-ATPase, antiporter, is primarily an ATPase but also exchanges calcium ions for protons (balances charge deficit, 1:2 ratio of Ca:H), causing the uptake of calcium into the SR where it can bind to proteins such as calreticulin and calsequestrin
- Cytoplasmic concentration is also decreased by PMCA (plasma membrane calcium ATPase, antiporter, 1:2 ratio of Ca:H) and Na-Ca exchanger (NCX), which will pump calcium ions out of the sarcoplasm and into the surrounding extracellular matrix
- The decrease in concentration causes calcium to dissociate from troponin-C, reverting the conformational change that allows cross bridges to be formed and therefore halting contraction, the fibres slide back to their original positions
How is contraction modulated in skeletal muscle?
- Recruitment is selective, greater or smaller number of motor units can be selected depending on necessity
- Frequency of stimulation can be changed, if increased will cause the summation of muscle twitch fibres
- The starting length of relaxed muscle can be changed (active length-tension relationship)
How does recruitment grade contraction in skeletal muscle?
- The more muscle fibres that are stimulated, the stronger the contraction will be
- Different fibres have different threshold voltages so stimulus intensity will dictate how many fibres are activated
How does summation grade contraction in skeletal muscle?
- Depends on the frequency of the stimulus
- The higher the frequency of stimulus, the higher the intracellular concentration of calcium (will be stimulated to be released from the SR more frequently)
- This means more actin filaments can be activated and more power generated resulting in stronger contraction
- Over-stimulation can result in fused tetanus however, with no breaks for relaxation of muscle as calcium levels are constantly above a certain threshold
What is one-to-one transmission in skeletal muscle?
- NMJ is highly specialised for this and amplifies response
- This means that each action potential will result in sufficient depolarisation for it to be propagated and contraction stimulated
- What is isometric contraction?
Where power is generated but the length of the muscle doesn’t change
- What is isotonic contraction?
Where the tension generated also results in a change of length (dependant on load)
- What is the ‘active’ length-tension relationship of skeletal muscle?
- The maximum force that a muscle fibre is able to produce is highly dependent on length
- Low force generation is seen at short and long lengths, with an optimum length in the middle producing the most force
- Highest force generation is produced when muscle fibres are at the correct length so that all of the myosin heads are engaged/able to attach to active sites of myosin
-> this is why hypertrophy of muscles causes an increase in strength, as the fibres are now more stretched
DRAW OUT DIAGRAM/GRAPH
What is systole?
When the heart muscles contract
What is diastole?
When the heart muscles relax
What is the purpose of the sinoatrial node (SAN)?
Acts as a pacemaker, has myogenic activity
What are the differences between the ventricular, atrial and SAN myocytes?
- Ventricular myocytes are large, mononucleate and striated
- Atrial myocytes are smaller, polynucleate and striated
- SAN cells have fewer contractile proteins and are very thin
What are the purpose of gap junctions and desmosomes in cardiomyocytes?
- Gap junctions provide electrical coupling and areas through which electricity can flow, allowing the cells to act as a syncytium
- Desmosomes allow mechanical coupling, very strong and span the plasma membranes of both myocytes
What is Starling’s law/Frank-Starling law of the heart?
- Starling’s law: the greater the filling of a cardiac chamber, the greater individual myocardial fibres are stretched (length-tension relationship) causing a greater force of contraction
- The Frank-Starling law: specific to the left ventricle, states that left ventricle stroke volume will increase with left ventricle stretch due to stretching of the muscle fibres resulting in a more forceful systolic contraction
- Both refer to contraction strength increasing, this is because double actin overlap will be changed/optimised at the optimal sarcomere length and the affinity of the TpnC (troponin complex) for calcium is dependent upon length
What is the function of the atrio-ventricular ring?
- Stops propagation of depolarisation, contains no gap junctions
- AVN, bundle of His and Purkinje fibres allow transfer of depolarisation to the ventricle tissue
How are the SA and AV nodes more adapted for myogenic activity?
The tissues are less negative than muscles
How does shape of action potential change during transmission?
- AP is delayed so repolarisation is delayed, width of action potential increases
- Different action potentials provide different functions:
- > pacemaker function in SAN and AVN, shallow and not as wide
- > fast and slow conduction of tissues allows synchronisation of contraction within the heart
- > long plateau to maintain refractoriness and avoid high frequency activation (limits heart rate, requirement for the heart which is a pulsatile pump)
What is the difference in action potential between the sub-epicardial tissues and sub-endocardial cells?
Sub-endocardial = Purkinje tissues/inner tissues, sub-epicardial = ventricular muscle/outer tissues
Sub-epicardial cells repolarise before the sub-endocardial, allowing the distribution of charge, depolarisation and positive deflection seen - even though outermost cells are stimulated last, their earlier repolarisation will result in the tissue depolarising at around the same time
What are the phases of action potential and the determining currents of the SAN?
- Phase 0, slow upstroke, only due to calcium current (open calcium channels)
- Phase 3, repolarisation, depends on potassium current (open potassium channels)
- Phase 4, electrical diastolic phase, in SA and AV nodes, changes in potassium, calcium and funny currents produce pacemaker activity (pacemaker potential) during phase 4. ‘Funny’ current is caused by HCN channel, Hyperpolarisation-activated Cyclic Nucleotide channel
What is the pacemaker current?
- Inwards calcium ion current
- De-activation of outward potassium current
- Inward cation current (funny current is activated at negative voltage)
- NCX current (sodium-calcium exchanged)
- Background inward sodium ion leak
Steepness of upwards current dictates rate of contraction, which is in turn dictated by stimuli to the SA node
What are the phases of action potential and the determining currents of the ventricular myocytes?
- Phase 0, fast upstroke, due to both calcium and sodium currents (blocked by TTX)
- Phase 1, rapid depolarisation, almost total inactivation of sodium or calcium current
- Phase 2, plateau is reached and prominent in ventricular muscle, continued entry of calcium or sodium ions through their major channels (channels from phase 1 not completely closed) and on the minor membrane current due to the sodium-calcium exchanger NCX1, 3 Na in for 1 Ca out
- Phase 3, repolarisation, depends on potassium current
- Phase 4, electrical diastolic, membrane voltage is termed the diastolic potential during phase 4
What type of ECC is seen in cardiac muscle?
- Electrochemical coupling
- Diads are seen
- Depolarisation of t-tubules causes a conformational change in the LTCC (L-type calcium channel), causing an influx of calcium ions into the sarcoplasm
- This causes calcium-induced calcium release, where the calcium ions bind to the ryanodine (RyR2) receptors on the sarcoplasmic reticulum and cause them to undergo a conformational change and release calcium ions from the SR
- This combined effect causes an increase in the intracellular concentration of calcium, which will bind to TnC and allow the formation of actin-myosin cross-bridges/for contraction to occur
- Contraction is dependant on this inflow of calcium, as shown by Springer in his experiments (attempts to create plasma for live hearts to be stored/transported in)
How does cardiac muscle relax?
- Only a little bit of action by SERCA pumps, which pumps calcium back into the SR with the aid of ATP hydrolysis and the transfer of protons to prevent a current across the membrane, is also associated with phospholamban (PLB)
- Calcium is pumped directly out of the cardiomyocytes by plasma membrane calcium ATPases, but only a small proportion
- There are also NCX proteins that exchange 3 sodium for every calcium ion, causing a net increase of charge, and the sodium ions are then pumped out again the the NKA (sodium-potassium ATPase) which exchanges 2 potassium for every 3 sodium. These pumps are electrogenic
- Decrease in intracellular concentration causes calcium ions to dissociate from the TnC molecules and inhibit cross-link formation again, allowing filaments to slide back past each other to their resting positions
What is phospholamban?
- Micropeptide protein
- Inhibits the action of the SERCA pump on the SR
- When phosphorylated, stops inhibitory action and allows function of SERCA pump to increase
- > phosphorylation is caused by specific kinases that are activated by beta adrenergic receptors
- > when phosphorylated, contractibility is increased allowing an increased heartrate to be achieved
- > general/unphosphorylated action is to slow or limit heartrate
What is the relation between systolic calcium concentration and active tension in cardiomyocytes?
- Rise in calcium concentration in normal cardiomyocytes during a beat is only enough to produce a fraction of the tension available
- Strength of contraction can be increased through increments of calcium attained through inotropic (modulatory of contraction) measures
- The more calcium present, the stronger the contraction (S shaped graph, practise drawing it)
How can cardiac output be modulated?
- Change in force of contraction (inotropy, affects ventricular muscle)
- > change in cell length (Starling’s law/the Frank-Starling mechanism)
- > cardiac nerves and circulating hormones acting on cardiac cells
- Change in frequency (chronotropy, effects SAN)
What effect does sympathetic stimulation have on ventricular myocytes?
Inotropy increased/stronger contractions
- Stimulatory
- Uses (nor)adrenaline
- Binding of neurotransmitter to a GPCR causes conversion of ATP to cAMP
- This activates protein kinase A
- The kinase phosphorylates the LTCC, increasing intracellular influx of calcium and the calcium plateau
- Also phosphorylates phospholamban, boosting calcium reuptake into the SR
- Phosphorylation of TnI (troponin inhibitory component) reduces the calcium sensitivity of the thin filament, increasing rate of relaxation
What effect does sympathetic stimulation have on the SAN?
Chronotropy increased/more frequent contractions
- Stimulatory
- Uses (nor)adrenaline
- cAMP binds to funny current channels and increases open probability therefore accelerating rate of pacemaker potential decay
- Phosphorylation of LTCC also accelerates pacemaker potential decay
- Phosphorylation of delayed rectifier potassium channels activates them and increases outwards current, causing a shortening of the action potential as well as contributing to pacemaker potential decay
- Pacemaker potential decay is the slow, positive increase in potential across a pacemaker cell which will allow the cell to generate it’s own action potential/exceed the threshold value
What effect does parasympathetic stimulation of the heart have?
- Only chronotropic/only affects the SAN and the AVN
- Inhibitory
- Uses acetylcholine
- Neurotransmitter binds to a GPCR with inhibitory function (Gi proteins) that inhibit/reduce production of cAMP and therefore has the opposite effect to noradrenaline or adrenaline
- This limits change of pacemaker currents, phosphorylation of photolamban, phosphorylation of the LTCC and phosphorylation of TnI
- Results in decreased rate and decreased stroke strength of contraction
- ACh also binds to muscarinic (M2) receptors (blocked by atropine), activating beta and gamma proteins that mediate activation of muscarinic potassium channels which cause hyperpolarisation, reducing the pacemaker function of SAN and AVN cells
Are t-tubules present in smooth muscle?
No, as cells are far smaller and do not need t-tubules to depolarise the whole cell
What nerves innervate smooth muscle?
Autonomic nervous system (unmyelinated)
What is the messenger for smooth muscle contraction?
Calcium ions (Otto Loewi once said ‘yes, calcium, that is everything!’)
What source of calcium ions does smooth muscle rely on?
- Extracellular influx
- Intracellular stores are relatively important but contraction is reliant upon influx
What is the process of muscle contraction in smooth muscle?
Agonists can bind to GPCRs or ligand-gated calcium channels
- GPCR mechanism:
-> G-protein is activated, causing the production of second messenger IP3 (inositol triphosphate)
-> These molecules bind to IP3 receptors (IP3R) and RyR3 receptors (ryanodine) on the SR
-> Causes release of intracellularly stored calcium
- Ligand-gated calcium channel mechanism:
-> Agonist binds to calcium channel, activating it
-> The subsequent conformational change allows a rapid influx of calcium ions into the cell
(-> There are also voltage gated calcium channels which open upon depolarisation and allow even more influx of sodium into the cell)
Calcium ions then bind to calmodulin, a protein in the cytoplasm with 4 binding sites for calcium
- Calcium-calmodulin complex activates myosin light chain kinases (MLCKs)
- MLCKs phosphorylate serine residues on myosin light chains
- This allows cross bridges to form/stimulates contraction
Is troponin C present in smooth muscle?
No, caldesmon and calponin thin filament proteins instead inhibit the action of myosin ATPase and the myosin light chain kinase, both of which prevents contraction
- Their inhibition is removed by calcium-calmodulin complex
What is the ‘latch state’ in smooth muscle?
- Slow myosin-actin dissociation, conserves energy and allows contraction to be continued at a low energy rate for a long time
- This is suggested to be achieved through the fact that if MLCs are dephosphorylated whilst still attached to actin, they will still remain attached and maintain force
What is calcium sensitisation of smooth muscles?
- This is where, after a contraction, the contractile filaments show an increased sensitivity to calcium
- More force is generated for the same inward current of calcium
- This is due to rho-kinase enzymes, whose action results in the inhibition of MLCPhs (myosin light chain phosphatases) which would allow the muscle to relax
Draw out pathways for contraction and sensitisation in smooth muscle
DO IT!!
How can calcium be visualised?
- Use indicator dies such as FURA-2/aequorin, fluorescent indicator protein isolated from jellyfish
- Waves of contraction can be viewed in the uterus through fluorescence caused as calcium is released, waves are mediated primarily by IP3 receptors
- Sparks can be seen in the ureter in a more spontaneous manner, reflects release of calcium through RyR3 channels
- What drugs are used to treat hypertension?
- L-type VGCC antagonists e.g. nifedipine
- > blocks calcium entry
- Alpha 1 adrenoreceptor antagonists e.g. doxasozin
- > blocks noradrenaline action
- Results in the vasodilation of arterioles, reducing vascular resistance to blood flow
- What drugs are used to treat asthma?
- Beta2 adrenoreceptor agonists e.g. salbutamol
- > in the lungs sympathetic nervous system optimises gas exchange so sympathetic innervation results in dilation
- ACh muscarinic receptor antagonists e.g. ipratropium
- > parasympathetic nervous system opposes sympathetic, so ACh will cause constriction, an antagonist will block this effect
- Relaxes bronchial smooth muscle to open airways
How is blood pressure determined?
- Blood pressure = cardiac output x total peripheral resistance (R ~ 1/r^4)
- Increasing diameter decreases resistance
- Decreasing diameter increases resistance
- As smooth muscle lines the blood vessels, it can have a huge effect on blood pressure
What are the three classes of calcium-channel blocking drugs (with examples)?
- Dihydropyridines (e.g. nifedipine, amlodipine)
- Phenylalkylamines (e.g. verapamil)
- Benzothiazepines (e.g. diltiazem)
All bind to alpha-1 subunits of LTCCs, but to distinct regions, causing the prevention of calcium entry into smooth muscle cells
How do the differing binding sites of calcium channel blockers (CCBs) result in different pharmacological effects?
- Some use use-dependant binding (e.g. diltiazem and verapamil), targeting cardiac cells, meaning the channels have to be activated before the drugs can have the effect and bind to specific regions
- Some use voltage-dependant binding (e.g. nifedipine), targeting smooth muscle cells, meaning that a specific voltage is required before the drugs are able to take effect
What are L-type voltage-gated calcium channels?
- ‘L-type’ is a reference to the long-lasting nature of the calcium channels
- Once activated, they will be open for as long as the depolarisation of the membrane is maintained
What is neurogenic activity?
This is activity that is controlled by the nervous system
What is myogenic activity?
This is where contraction of a muscle cell is controlled by the cell itself rather than through innervation
How do smooth muscle cells show both myogenic and neurogenic activity?
- Neurogenic, innervated by autonomic nervous system, receive neurotransmitters from varicosities which can stimulate or inhibit contraction (these are far less specific than seen in skeletal muscle, neurotransmitters are even able to diffuse to adjacent cells)
- Myogenic, seen in smooth muscle lining the blood vessels (in this case stretch-mediated, has been shown that stretching the fibres causes them to contract)
What effect does an action potential have in smooth muscle (when present)?
- Not always necessary
- Can initiate contraction or relaxation (not just contraction as seen at skeletal muscle)
- No defined motor end plate
- Both sympathetic and parasympathetic nervous systems innervate smooth muscle
- Both release different neurotransmitters, with opposite effects/which bind to different receptors and start different cascade reactions or inhibit the action of different molecules
- Depolarisation of the membrane will also result in an influx of calcium/activation due to the L-type calcium channels being voltage gated as well
How other than by an action potential can contraction or relaxation be stimulated in a smooth muscle cell?
- Hormones
- NO (can be synthesised in endothelial cells - found in vascular structures - lipid soluble, causes smooth muscle to relax)
How can smooth muscle be relaxed?
- As dependant upon calcium as in skeletal and cardiac muscles, intracellular concentration of calcium needs to be decreased
- > Mg/Ca ATPases exchange two calcium ions for one magnesium ion stored within the SR using the hydrolysis of ATP (similar to SERCA), these pumps also exist on the cell membrane
- > NCX proteins exist along the sarcolemma, exchanging extracellular sodium for calcium
- Also requires the dephosphorylating action of myosin light chain phosphatases, deactivating the myosin chains
- ACh binds to GPCRs, triggering the release of nitric oxide, which in turn activates guanylyl cyclase, converting GTP into cyclic GMP (cGMP), which then activates to protein kinase G
- > PKG results in a cascade that causes the uptake of calcium
- > upregulates MLCPhs, downregulates MLCKs
- Same process is seen in the activation of beta1 adrenergic receptors in vascular smooth muscle, apart from cAMP is the secondary messenger and the action of protein kinase A is encouraged, but relaxation still occurs
