Neuromuscular Junction (Week 1--O'Dell)

Question 1

Anatomy of the NMJ

Answer 1

NMJ is called the end-plate, and is the site of contact between a motor neuron and a skeletal muscle fiber

Synaptic boutons at end of axons of motor fiber contain synaptic vesicles containing ACh

ACh released from active zones on presynaptic terminal (sites of NT release)

Postsynaptic muscle fiber has regularly spaced invaginations called junctional folds, which contain acetylcholine receptors (nicotinic) on the crests of the junctional folds

Synaptic basal lamina (basement membrane) within synaptic cleft which contains acetylcholinesterase (technically on postsynaptic membrane)

Question 2

Synaptic transmission at the NMJ

Answer 2

1) Depolarization of presynaptic terminal membrane by action potential activates voltage-sensitive Ca2+ channels at presynaptic terminal
2) Ca2+ flows into presynaptic terminal and triggers fusion of synaptic vesicles with presynaptic membrane
3) ACh in vesicles released into synaptic cleft and diffuses to postsynaptic membrane
4) ACh receptors in postsynaptic membrane are ligand-gated ion channels that undergo conformational change to open ion channel and let Na+ in (also permeable to K+) to depolarize (depolarization produced by activation of nicotinic receptors called end-plate potential, or EPP)
5) ACh cleared from synaptic cleft by being broken down by acetylcholinesterase

Question 3

Is an EPP an action potential?

Answer 3

NO!

EPP is just the depolarization that occurs when ACh binds nicotinic receptors at NMJ

AP is created if EPP strong enough (so, usually but not always). Then, voltage gated Na+ channels open to cause AP

Question 4

What does it mean that NT release is quantal?

Answer 4

NTs are released in small packets corresponding to individual synaptic vesicles

1 quanta = 1 synaptic vesicle

Question 5

Miniature end plate potentials (MEPPs)

Answer 5

Postsynaptic responses to ACh spontaneously released from presynaptic nerve terminal

Small, randomly occurring spontaneous potentials in postsynaptic cell occur about 1-5 per second

Identical to normal EPPs but much smaller

Question 6

What happens when extracellular [Ca2+] is decreased?

Answer 6

Normal EPP becomes smaller and smaller

Question 7

What are EPPs?

Answer 7

Near simultaneous release of multiple quanta (synaptic vesicles)

Size of EPP depends on number of quanta released (quantal content)

Question 8

Quantal content (m)

Answer 8

Number of quanta that make the EPP

Question 9

How does Ca2+ trigger exocytosis of ACh synaptic vesicles?

Answer 9

1) v-SNAREs on vesicle (synaptobrevin) bind t-SNAREs on presynaptic terminal membrane (syntaxin and SNAP-25) and vesicle “docks” on membrane
2) Complexin prevents vesicle from fusing and releasing ACh
3) Ca2+ comes into presynaptic terminal from outside and binds vesicle protein called synaptotagmin causing it to undergo a conformational change
4) Synaptotagmin completes fusion of vesicle to presynaptic membrane and ACh is released into synaptic cleft

Note: synaptobrevin is v-SNARE that binds SNAP-25 and syntaxin; synaptotagmin is the Ca2+ sensor

Question 10

How does Botulinum Toxin (BoTox) block synaptic transmission?

Answer 10

Cleaves synaptobrevin (v-SNARE) so that it can’t bind t-SNAREs to dock the synaptic vesicle to presynaptic membrane

Other components of BoTox also cleave syntaxin and SNAP-25

Basically, disrupt SNAREs so no synaptic transmission at NMJ

Muscle paralysis

Note: produced by anaerobic spore-forming bacteria and most often caused by eating improperly prepared/stored food

Question 11

Safety factor

Answer 11

For one presynaptic action potential, you will trigger a postsynaptic action potential

NMJ has high safety factor: only need 100 quanta to pass threshold and get postsynaptic AP, but instead release about 200 just to make sure you’ll get an AP

“synapses always work”

Question 12

For one presynaptic action potential, do we always get the same strength of postsynaptic EPP?

Answer 12

No, it is very variable!

For one presynaptic AP could get release of 200 quanta, but the next will be 170 quanta and the next 210 quanta, etc etc

Question 13

Facilitation

Answer 13

When multiple APs occur in rapid succession and EPPs evoked by each AP progressively increase in amplitude

This happens because transmitter release is sensitive to Ca2+ concentration and higher level of Ca2+ that builds up evokes release of larger number of synaptic vesicles which result in larger EPP

Usually, once AP over Ca2+ is removed from release sites, but if rapid succession of APs, there is residual Ca2+ that stays around which summates with new Ca2+ influx resulting in higher levels of Ca2+ in presynaptic terminal

Question 14

Depression

Answer 14

During prolonged burst of presynaptic action potentials, size of EPPs evoked by each AP eventually begins to decline

This happens because number of vesicles filled with NT and available for release declines (all used up!)

Usually this is not a problem, but people with myasthenia gravis (lower safety factor) could have decreased synaptic transmission because of depression

Question 15

Curare

Answer 15

Reversible, competitive inhibitor of ACh derived from bark of tropical tree

Used to partially block postsynaptic ACh receptors to reduce EPP to below threshold if you want to see just the EPP alone

(alpha-bungarotoxin does the same thing)

Question 16

Nicotinic ACh receptor

Answer 16

Ligand-gated ion channel

ACh binding causes conformational change that opens ion channel permeable to both Na+ and K+ (lets a little Ca2+ in also, but not much)

At normal resting potential, you’ll get influx of Na+ into cell

Pentameric protein with ACh binding site on alpha subunits; membrane-spanning

Question 17

When does the end plate current (EPC)/synaptic current terminate?

Answer 17

When there’s no more ACh around (because it’s been hydrolyzed by AChE)

Now the nicotinic ACh receptor is no longer open and membrane no longer permeable to Na+ and K+, so back to resting membrane potential

Question 18

Equation for current (I) through channel

Answer 18

I = g(V_m - E_rev)

g = membrane conductance

V_m - E_rev = driving force (moves ions through the channel)

Question 19

Equation for current through ACh receptor channels at any given membrane potential

Answer 19

I_epc = g_Na x (V_m - E_Na) + g_K x (V_m - E_K)

Sum of current carried by Na and current carried by K; end plate current

Question 20

Reversal, or equilibruim potential for nicotinic ACh ion channels

Answer 20

-15mV

Between E_Na (+40) and E_K (-90)

Question 21

Alpha-latrotoxin

Answer 21

From black widow spiders

Induces massive spontaneous release from nerve terminals via 2 mechanisms:

1) Activates presynaptic neurexin to directly cause NT release (even without Ca2+)
2) Forms Ca2+ permeable pores in plasma membrane and influx of Ca2+ triggers NT release