W13 - Neuromuscular Junction Flashcards
Explain synaptic transmission at the neuromuscular junction
These stages can be influenced by drugs and toxins
1. Synthesis
- from precursor component parts w/ enzymes
2. Storage
i) protect from metabolic enzymes
ii) package quanta (1 vesicle) containing multiple neurotransmitters
3. Release
- Ca+ enters pre-synaptic voltage-gated channels -> binds to protein -> causes exocytosis of vesicles
4. Activation
- changes in excitation of post-synaptic
5. Inactivation
- specialised effectors recognise & re-uptake
- enzyme breaks down neurotransmitters into component parts
Explain how drugs can enhance synaptic transmission.
A) Direct stimulation
- the natural transmitter
- analogues
B) Indirect stimulation
- increased transmitter release
- inhibition of transmitter removal
Explain how drugs can inhibit synaptic transmission
A) Blocking synthesis, storage or release from pre-synaptic neurons
B) Blocking post-synaptic receptors
Define agonists
Drugs, hormones or transmitters that bind to specific receptors + initiate conformational change in receptors
- have both affinity and efficacy
- affinity: ability to bind
- efficacy: ability to initiate a biological response once bound
- the receptor only recognises the precise ligand/agonist, and activates them
- reversible
Define antagonists
Drugs, hormones or transmitters that bind to receptors but don’t activate them
- have affinity but lack efficacy
- blocks receptor activation (by agonist)
Explain competitive receptor antagonists
- competes with agonist for agonist binding site
- block can be reversed by increasing agonist concentration
Classify the two types of nicotinic acetylcholine receptors
- acetylcholine (ACh) is the neurotransmitter at the NMJ
- pre-synaptic neurons that synthesise and release ACh = cholinergic
- ACh acts on receptors called cholinoceptors
- Nicotinic cholinoceptors (nAChRs)
- gated-ion channel
- activated by ACh or nicotine (tobacco alkaloid)
- NOT muscarine
- Muscarinic cholinoceptors
- G-protein (GPCRs)
- activated by ACh or muscarine
- NOT nicotine
- slower than nicotinic cholinoceptors
Explain how fast synaptic transmission occurs with transmitter-gated ion channels
Eg. Nicotinic acetylcholine receptor nAChR
- Cation conducting channel (Na+ in, K+ out)
- Agonist binding -> rapid conformational change -> open channel
- selective for certain ions
- signalling is extremely rapid (millisec)
Explain how the patch-clamp technique can be used to record the functional properties of single receptors
- glass micropipette forming a tight gigaohm seal (suction) with the cell membrane
- contains a wire bathed in an electrolytic solution to conduct ions
- allows to report activity of single receptor, (a dip in graph = opening of channel)
- pA = 10^-12A (i.e. 1 million millionth of an Amp)
Explain the structure of neuromuscular junction
The motor axon is surrounded by α-bungarotoxin binding once identifying the nicotinic receptors
- α-bungarotoxin: protein that binds irreversibly
- shows up red on scans (?)
Explain how mepps at the NMJ are recorded + the mechanism
- i.e. small depolarisations, via a glass electrode (end plate region)
- release of ACh from vesicles
- 1 vesicle (multiple of neurotransmitter) = multiple nAChR
- exocytosis (coordinated to sharp increase in recording)
- activation of nAChR => cation channels open -> flux of Na+ ions into muscle fibres -> local depolarisation at endplate region
- 1 vesicle (multiple of neurotransmitter) = multiple nAChR
Explain how black widow spider venom influences spontaneous transmitter release
- Eg. Black Widow spider venom (α-LATROTOXIN –α-LTX) influences spontaneous transmitter release
- 10-15 mins: massive ACh release → muscle spasms
- 1 hr: ⇒ depletion of vesicles
- inhibition of exocytosis
- presynaptic neuron changes shape (expands); acts as if there is an extra membrane layered inbetween, (no exocytosis)
- distended terminal paralysis
- inhibition of exocytosis
Explain a simplified model of “fast” synaptic transmission
- presynaptic AP
- a synchronous localised Ca2+ influx via voltage-gated Ca2+ channels
- bind to sensor proteins on vesicles (approx. 0.1 ms)
- vesicles undergo exocytosis -> release of ACh
- activation of nAChR
- large depolarisation of the endplate region of the muscle cell (an epp; made up of multiple mepps)
- depolarisation is large enough => postsynaptic voltage-gated Na+ channels to initiate an action potential
- K+ exiting the cell bring depolarisation back down
Explain how Mg2+ and Ca2+ extracellular solution reduces
- Magnesium blocked the voltage-gated Ca2+ ion channel
- high magnesium and low calcium = reduced epp (not enough to cross the threshold)
- no muscle AP = no contraction
- high magnesium and low calcium = reduced epp (not enough to cross the threshold)
State how to calculate quantal content
- the amplitude of the epp is a multiple of the amplitude of mepp (smallest epp amplitude = mepp amplitude)
- quantal content
- mean EPP amplitude (mV)/mean MEPP amplitude (mV)
- i.e. no. of vesicles/stimulus
- release of a vesicle gives a “quanta” of transmitter
- each quanta -> mepp via activation of nAChR
- mepps are spontaneous (w/o nerve stimulation)
- each quanta -> mepp via activation of nAChR
- mean EPP amplitude (mV)/mean MEPP amplitude (mV)
Explain how cholinergic transmission can be blocked at the synthesis step
- From exocytosis, ACh not only activated channels, but also AChE (enzyme) on post synaptic neuron
- ACh is broken down into acetate and choline, and there is a reuptake of choline into the presynaptic neuron
- The choline acetyl transferase (CAT) synthesises ACh from choline and Acetyl Coenzyme A precursor from mitochondria
- the reuptake of choline is Na+-dependent
- can be blocked by hemicholinium 3 => less ACh in each vesicle
- amplitude of epp and mepp both decreased i.e. no change in quantal content
Explain how cholinergic transmission can be blocked at the storage step
- after ACh is synthesised, the transport into the vesicles can be blocked by inhibition of the ACh vesicular transporter
- by Vesamicol
- both epp and mepp decreased (no change in QC)
State the effects of tetrodotoxin on cholinergic transmission
- produced by bacteria; concentrated in organs of the puffer fish (e.g. liver)
- causes paralysis of the diaphragm => respiratory failure
- more potent than local lidocaine
- blocks Na+ channels (i.e. no AP, no release, no EPP)
State the effects of conotoxins on cholinergic transmission
- by cone snail
- voltage-gated Ca2+ channels blocked
- decrease in Ca2+ influx -> decrease in release
- EEP decrease, no mepp change, decrease in QC
State the effects of dendrotoxin in on cholinergic transmission
- from green mamba
- blocks voltage-gated K+ channels -> prolonged AP -> increase in Ca2+ influx -> more release
- increased epp amplitude, no change in mepp amplitude (QC increased)
- the synthetic drug 4-aminopyridine (A-AP) has a similar mechanism
State the effects of botulinum toxin in on cholinergic transmission
- most potent toxin from clostridium botulinum bacteria
- respiratory paralysis
- cosmetically used as botox, can also be used for chronic migraines
- blocks vesicle fusion by cleaving a vesicular protein required for exocytosis -> decreased release
- epp amplitude decreases; no change on mepp amplitude
- QC decrease
Describe the function of the organ bath
- Dissect the phrenic nerve hemi-diaphragm preparation & mount in an organ bath.
- Electrically stimulate the phrenic nerve 1/10 sec & record the resultant muscle twitch on a chart recorder or computer.
How does tubocurarine produce skeletal muscle relaxation at the NMJ
- muscle relaxant
- only in blood supply
- decreases twitch tension
- decreases depolarisation -> not sufficient for epp to pass threshold for muscle fibre AP generation
- competes with muscle nicotinic ACh receptors
- works post-synaptically, but not pre-synaptically
State the conditions of tubocurarine action on a synapse
