M&R Session 5 Flashcards
What happens at the nerve terminal?
Depolarisation –> opens V-G calcium channels –> calcium influx –> release of neurotransmitter
Which channels are found in the nerve terminal?
V-G sodium
V-G potassium
V-G calcium
What is the difference between intracellular and extracellular calcium concentration?
10,000 fold
Describe the structure of a V-G calcium channel.
Very similar to V-G sodium channels
Four repeats each consisting of 6 transmembrane domains
Porphyrin alpha subunit
Only one needed for channel formation
What type of blockers act on L-type V-G calcium channels?
Dihydropyridines
Describe the subunit composition of calcium channels.
Porphyrin alpha subunit associated w/other subunits which fine tune properties and allow correct regulation of activity
How does calcium influx cause transmitter release?
Calcium influx –> binds to synaptotagmin –> brings vesicles close to plasma membrane –> snare complex makes fusion pore –> transmitter released through pore
What is synaptotagmin?
A protein which brings vesicles close to plasma membrane
How does increasing intracellular calcium affect transmitter release?
Increases
What affect does an increase in the number of action potentials to the nerve terminal have?
Increases pulses of transmitter released
How many subunits does a V-G calcium channel have?
5; incl. 2 alpha subunits
Where are nicotinic AChR found?
NMJ
Fast synaptic receptors
What feature of nAChR allows them to be used in fast synaptic receptors?
They allows fast depolarisation
How is an nAChR activated?
2 ACh molecules bind causing a conformational change that opens channels
What does the nACHR in an NMJ try to achieve with the membrane potential?
To reach -10 mV therefore causing depolarisation of the skeletal muscle fibre
How selective is the sodium-potassium channel?
Not very - equally permeable to both so ion movement is determined by equilibrium potential
What happens to the sodium-potassium channel at a membrane potential of -10 mV?
Reversal voltage is met
No net flow of charge
Current through channel reverses
What causes the end-plate potential?
ACh action on nAChR
How does decreasing extracellular calcium affect the end-plate potential?
Decreases amplitude
What are the sequence of events during neuromuscular transmission?
AP in neurone –> ACh release –> activate nAChR –> brief depolarisation (end-plate potential) –> adjacent sodium channels activated by local spread of charge –> excitation-contraction coupling
What are the two types of nAChR blockers?
Competitive
Depolarising
What is the mechanism of activation of tubocurarine?
Competitive blocker
Binds to alpha subunit but does not cause a conformational change
Decreases depolarisation without changing ACh amount
Can competitive nAChR blockers be overcome?
Yes - vastly increase ACh concentration
Which enzyme breaks down succinylcholine?
Plasma cholinesterase
Can depolarising nAChR blockers be overcome?
Nope
Describe the action of depolarising nAChR blockers.
Binds to alpha subunit –> receptor opens –> no AChE caused degradation –> receptor remains open –> sustained depolarisation –> sodium channels inactivated and receptors desensitised
What causes miniature end-plate potentials?
Spontaneous release of ACh vesicles w/out AP stimulus
What is the rate of ACh release in a miniature end-plate potential?
~1 quanta per second
What is the significance of the miniature end-plate potential?
It can be recorded (but does not reach anywhere near threshold potential)
What is myasthenia gravis?
Autoimmune targeting of nAChR
How are functional nAChR lost in myasthenia gravis?
Complement mediated lysis
Receptor degradation
What measurement can be used to determine myasthenia gravis and what does it show?
End-plate potentials - each quanta released gives a decreased response in comparison to unaffected muscle
S/S of myasthenia gravis?
Profound weakness
Muscle weakness increases w/exercise
Muscle fatigue
Where are muscarinic AChR found?
NMJ of parasympathetic branch of the ANS
Which give a faster response, nAChR or mAChR?
nAChR
Summarise the activation of an mAChR in one sentence.
ACh binds to the receptor causing the associated G-protein to split, allowing the alpha G-protein subunit to modify the activity of the target effector
Do both nAChR and mAChR use ligand binding?
Yes
Why are mAChR slower to initiate a response than nAChR?
They are coupled to G-proteins which trigger a cascade of events in the cell to cause a response. They do not have an ion channel like nAChR
What is calcium responsible for?
Bone and teeth integrity, fertilisation, proliferation, secretion, neurotransmission, metabolism, contraction, learning, memory, apoptosis, neurons etc.
What is the advantage of having a large inward calcium gradient?
Changes in calcium are rapid w/small amounts of calcium ion movement
What are the disadvantages of having a large inward calcium gradient?
Calcium overload is easy
Rapid loss of calcium regulation can lead to cell death
How is the calcium gradient set-up and maintained?
Relative impermeability of the plasma membrane
Cell ability to expel calcium
Calcium buffers
Intracellular calcium buffers
Which two transporter mechanisms are used to expel calcium across the CSM?
Calcium-ATPase
Sodium/calcium exchanger
What is calmodulin?
A high affinity, low capacity trigger protein used for calcium expulsion
Name three calcium binding proteins.
Synaptotagmin
Calmodulin
Troponin
What factors of calcium binding molecules affect the diffusion of calcium?
Concentration
Saturation
What two types of intracellular calcium stores are there?
Rapidly and non-rapidly releasable stores
Where are rapidly releasable intracellular calcium stores found?
Sarcoplasmic reticulum
Endoplasmic reticulum
What pump is used to release calcium from rapidly releasable stores?
SERCA
What does SERCA require?
A low affinity, high capacity binding protein - e.g. Calsequestrin
Where are non-rapidly releasing calcium stored found?
Mitachondria
How does the calcium gradient across a mitochondria membrane compare to that across the cell membrane?
It is much greater
When will mitochondria take up calcium?
When the intracellular concentration is high
Why does mitochondrial uptake of calcium have to match energy demand and supply?
It requires ATPases
Describe the mechanism of uptake of calcium by mitochondria.
Uniporter uses driving force from respiratory chain hydrogen ion production
What are the properties of the uni porter used for calcium uptake in the mitochondria?
Low affinity
High capacity
How is the internal calcium concentration elevated and returned to basal levels?
Calcium influx across the plasma membrane
Calcium release from rapidly releasable intracellular stores
Calcium release from non-rapidly releasable intracellular stores
Which two channels are involved in influx of calcium across the plasma membrane?
V-G calcium channels
Receptor-operated calcium channels
Give some examples of receptors used for receptor-operated calcium channels.
NMDA
AMPA
Glutamate receptors (especially in the brain)
Some nAChR
What is the term given to describe receptor-operated calcium channels?
Inotropic channels
How do V-G calcium channels operate?
Depolarisation –> conformational change
How do receptor-operated calcium channels work?
Ligand/agonist binds –> opens aqueous pore –> calcium can move in
Why will mitochondria take up calcium?
They need a high concentration to stimulate mitochondrial metabolism
What is the role of mitochondria in calcium regulation?
Buffering Signalling Match energy demand and supply Apoptosis Altered redox potential
By which two methods is the release of calcium from rapidly releasable intracellular stores mediated?
G-protein coupled receptors
Calcium-induced calcium release
Describe the mechanism of GPCRs in calcium release from non-rapid stores.
Ligand/agonist binds to GPCR –> turns receptor on –> effector molecules activated –> cellular response
Which two effector molecules are stimulated by activation of GPCRs?
Plasma channels
Muscarinic receptors
How can the activation of GPCRs lead to different cellular responses in different cells?
Different muscarinic receptors are present in different cells - M1 to M5
What is the function of alpha-q once it has split from the 7-transmembrane domain of the GPCR?
It activates phospholipase C which activates IP3 and diacylglycerol
What is the action of IP3?
Ligand-gated ion channels found in the sarcoplasmic/endoplasmic reticulum which have a pore permeable to calcium, allowing its efflux from the S/ER to increase cellular calcium levels
What is the action of diacylglycerol?
Protein kinase C regulation
What molecule do alpha-s and alpha-I have opposite actions on?
Adenyl cyclase
What is the mechanism of adenylyl cyclase action?
Produces cAMP –> phosphorylates protein kinase A
Where are M3 receptors found?
Walls of the airways
What does activation of the M3 receptors cause?
Contraction of smooth muscle in the airways
Where is ryanodine derived from?
Plants
Where are ryanodine receptors found?
In the sarcoplasmic/endoplasmic reticulum
How are ryanodine receptors activated?
Conformational coupling with voltage operated calcium channels
Depolarisation
How does calcium exit through ryanodine receptors?
Acts as an agonist –> conformational change –> released
How does depolarisation of the T-tubule cause calcium release?
V-G sodium channels open
VOCC open
Why are the ryanodine receptors so important for calcium release?
Give explosive release of 85% of calcium
Located directly adjacent to contractile proteins
What happens in the cardiac myocyte when calcium levels decrease?
Ryanodine receptor stops
SERCA pumps calcium into stores
NCX reverses –> calcium into T-tubule for minor store
What restores normal function of the NCX following decreased calcium levels?
Sodium removal
What causes the plateau in the cardiac action potential before repolarisation occurs?
Calcium influx via VOCC
What is the role of calcium in muscle contraction?
Bind with tropomyosin and roll it out of the way
Exposes binding sites for cross-bridge recycling
How are microdomains formed?
Strategically placed calcium channels allow local variations in calcium levels
Why does repetitive signalling need restoration of basal calcium levels?
Too much calcium for too long is toxic
How do cells return intracellular calcium concentrations to basal levels?
Use of transient signals
What does return of calcium to basal levels require?
Termination of signal
Calcium ion removal
Calcium ion store refilling
How are calcium stores refilled?
Recycling of released cytosolic calcium
VOCC
Capacitative/store operated calcium channel
Describe the mechanism of recycling released calcium.
In cardiac myocytes effectively release and grab back cytosolic calcium
Describe the mechanism of channel led calcium store refilling.
Deleted signal to membrane allows more calcium in
Specific proteins interact to activate the channel
What is STIM-ER?
ER membrane located calcium sensor w/motif that binds to calcium which causes a conformational change
What is ORAI?
A plasma membrane channel which allows entry of calcium
Can channel led calcium store refilling regulate physiological processes?
Yes
Is the mechanism of calcium regulation always the same?
No, it may be dependent on cell type and circumstance
