Transcript Neuromuscular Transmission

Today’s Objective:
Neuromuscular Transmission
• Describe the sequence of electrical and chemical events at
the neuromuscular junction during transmission of an action
potential.
• Provide some of the evidence that acetylcholine (ACh) is
released, and that the release is quantal in nature.
• Describe the ionic mechanisms of the endplate potential
(EPP), including the types of ion channels involved.
• Consider the disorder Myasthenia Gravis, and the rationale
for treatment with inhibitors of acetylcholinesterase.
Review: Main Components of Neurons
•Soma, axon, myelin sheath, nodes of Ranvier and
terminal boutons.
Nodes of
Ranvier
Terminal Boutons:
Sites of transmitter
release
The Motor End Plate
•A myelinated nerve fiber
is seen reaching
the
surface of muscle cells
that have been teased
apart.
•The terminal region
branches extensively to
form the neuromuscular
junction.
Axon
Striated muscle
fibers
Motor End Plate
From R.A. Bergman, A.K.
Afifi and P.M. Heidger
Virtual Hospital: Atlas of
Microscopic Anatomy
University of Iowa
Sequence of Events in Neuromuscular
Transmission (1)
Presynaptic Events
1. Action potential (AP) is initiated in the presynaptic motor
neuron (MN) and invades the endplate region
2. Depolarization of MN terminal boutons, resulting in the
opening of voltage-dependent calcium channels
3. Influx of Ca2+, down its concentration gradient
4. Rise of intracellular free [Ca2+] initiates fusion of vesicles
containing acetylcholine (ACh) to the membrane of the
terminal boutons, resulting in exocytosis of ACh
5. Diffusion of ACh across synaptic cleft to the muscle cell
Sequence of Events in Neuromuscular
Transmission (2)
Postsynaptic Events
6. Binding of ACh to nicotinic ACh-receptors at endplate
7. Receptor binding causes opening of cation channels, leading
to influx of Na+ (ACh is then degraded by acetylcholinesterase
present in the synaptic cleft)
8. The resulting depolarization of muscle cell membrane at the
endplate is referred to as the endplate potential (EPP)
9. The local depolarization causes adjacent regions to be
depolarized, causing an AP in the muscle cell membrane
10. AP spreads out in all directions from the endplate, propagates
along muscle cell, initiating contraction
Motor End Plate
Electron micrograph of nerve terminal
Presynaptic
terminal, with
many small
vesicles containing
ACh
Postsynaptic
region of the
skeletal muscle,
with
mitochondria
and contractile
filaments
apparent in the
cytoplasm
500 nm
From Alberts, Bray,
Lewis, Raff, Roberts
and Watson, Molecular
Biology of the Cell,
2nd edition
Motor End Plate (continued)
Each vesicle
contains ~5000
ACh molecules.
1 vesicle=1 quanta
Each vesicle is
~50 nm diameter
Postsynaptic membrane:
Clusters of nicotinic ACh
receptors in the junctional folds
Synaptic cleft between
nerve and muscle cells
Size? A 50 to 100 nm gap.
Exocytosis at the Terminal Boutons
Resting nerve with
abundant vesicles
Stimulate nerve
and observe fusion
of vesicles with
membrane.
Vesicle exocytosis
releases ACh into
synaptic space
Synaptic cleft is 50
to 100 nm
Time for diffusion
of ACh is ~0.5 ms
End Plate Potentials
• From mini-EPP, to summation and EPP to Action Potential
Record
Membrane
potential
of muscle
Schematic
courtesy of
T. Stavraky
Stimulate nerve
Vm
0.5
mV
Miniature EPP
(spontaneous, 1 vesicle
aka 1 quantum)
Vm
5
mV
Stimulate
nerve
EPP
(evoked, ~200 quanta)
Vm
5
mV
Stimulate
nerve
EPP brings membrane
to threshold and initiates
action potential
Evidence that ACh is the neurotransmitter at the NMJ
• ACh is synthesized and released by the nerve terminal; synthesis of
ACh is blocked by hemicholinium (which inhibits choline uptake)
• Small amounts of ACh applied to the end plate cause depolarization
of the muscle. The ACh-induced depolarization and EPP have the
same reversal potential and permeability to Na+ and K+.
• The pharmacology of the ACh induced depolarization and EPP are the
same, including:
• (a) activation by ACh agonist (nicotine, carbachol, e.g.)
• (b) blocking by ACh receptor antagonist (curare)
• (c) enhancement and prolongation the EPP by anticholinesterase
(e.g., neostigmine, which prevents the hydrolysis of ACh)
Nicotinic ACh Receptor
• Receptor is a pentameric complex of 2 a, b, g and d subunits
• Binding of 2 ACh molecules causes opening of the channel,
which is a channel permeable to all cations, including Na+,
K+ and Ca2+.
• The evoked currents reverse direction close to 0 mV.
• Thus, at the resting potential, the primary effect is influx of
Na+, accounting for the depolarization
Summary of Neuromuscular Transmission
Action potential invades presynaptic terminal
Opening of Ca2+ channels leads to influx
of Ca2+ (extracellular Ca2+ is essential)
+ +
Vesicles fuse and release ACh into cleft
ACh diffuses across synaptic cleft
ACh-activated channel is permeable
to both Na+ and K+, so the reversal + +
potential is a mixture of the two
ACh activates cation channels to
cause depolarization of the endplate.
Summary of Neuromuscular Transmission
+ +
Depolarization of the end plate
initiates an action potential
that spreads over muscle cell
ACh is destroyed by
acetylcholinesterase enzymes
in the synaptic cleft
In healthy muscle, an AP in the
+ + motor neuron ALWAYS activates
an AP in the muscle, leading to
contraction (safety factor is ~2).
Pharmacology of the End Plate Potential
And now for something completely different
Ancient Chinese Proverb
I hear and I forget.
I see and I remember.
I DO and I understand.
In the doing is the learning!
Pharmacology of the End Plate Potential
Curare
Neostigmine
•Blocker of nicotinic receptors
•Plant extract used to induce paralysis
•Must be careful with patients, as they
may feel pain but cannot show it
Control EPP
Vm
5
mV
+ Curare
Stimulate
nerve
•Blocker of
cholinesterases
•Component of nerve
gases and insecticides
+ Neostigmine
Vm
5
mV
Control EPP
Stimulate
nerve
Clinical Case:
• A patient presents with muscle weakness, and fatigues easily
From Principles
of Neural
science, 3rd
edition, by E.
Kandel, J.
Schwartz and T.
Jessel.
Findings:
• No muscle atrophy
• Normal nerve conduction
• muscle response to repeated stimulation is decreased
• muscle response with repeated direct stimulation of the muscle is
normal
• When muscle biopsy is studied, small MEPPs and EPPs are observed.
• Muscle weakness improves with neostigmine (tensilon has briefer
duration, so is better in the clinical setting)
Tentative Diagnosis: Myasthenia Gravis
• Myasthenia Gravis is an autoimmune disease in which patients
develop antibodies against nicotinic ACh receptors.
• Thus, the amplitudes of MEPPs and EPPs are reduced (since there
will be less depolarization for the same amount of released ACh) and
the muscle membrane may not be depolarized sufficiently to fire an
action potential.
• Treatment is to give an acetylcholinesterase inhibitor to prolong and
increase the action of ACh at the available receptors and to restore the
muscle action potential.