Acetylcholine

Question 1

What is the criteria for neurotransmitters?

Answer 1

1. Presynaptic cell should contain the substance as well as a mechanism to synthesise it
2. Substance should be released when depolarizing stimulus is applied to the neuron
3. Receptors should be present on the postsynaptic cell
4. Known antagonists should block the effects
5. A system to inactivate the substance must exist
6. Exogenous application of the substance to the postsynaptic cell should produce the same response

Question 2

Describe neuromodulators

Answer 2

• Can be hormone-like (acting at a distance)
• Autocrine (self-acting)
• Juxtacrine (adjacent)
• Paracrine (near)
• Endocrine (far)
• Exocrine (outside)
• May act at locations other than the synapse
• Does not elicit a direct effect on the postsynaptic cell, but alters the action of a classical neurotransmitter
• Enhances, reduces, or prolongs NT action
• May function in anterograde or retrograde manner
• e.g. pre- ↔ post-synaptic
• May be released from glial cells

Question 3

Describe acetylcholine

Answer 3

• Acetylcholine was the first neurotransmitter identified, used in seminal experiments to determine the nature of chemical transmission at the synapse.
• Common neurotransmitter at neuromuscular junctions in the periphery.
• Neurons that use acetylcholine are termed cholinergic neurons.

Question 4

Describe the synthesis of acetylcholine

Answer 4

Acetyl-CoA binds to Choline using choline acetyl transferase (ChAT)
as a catalyst.

Question 5

Why is ChAT expression important?

Answer 5

• Immunofluorescence micrograph of cholinergic motor neurons in the ventral horn of the spinal cord
• ChAT is only expressed in the cytoplasm of neurons that use ACh as a neurotransmitter

Question 6

Describe choline

Answer 6

• Choline is derived primarily from hydrolysis of dietary lipids (phosphatidylcholines) by phospholipase D.
• Choline is water soluble and is only capable of crossing the BBB due to the presence of specific choline transporters.

Question 7

How is acetylcholine metabolized?

Answer 7

Acetylcholine is metabolized to choline and acetic acid/acetate by the enzyme acetylcholinesterase (AChE)

Question 8

Describe acetylcholine turnover

Answer 8

• ACh synthesis is tightly regulated

Product inhibition

• High levels of ACh in the cell inhibit the synthesis of further ACh by ChAT

Precursor availability (choline and acetyl-CoA)

• Increasing dietary choline has been explored as a treatment for conditions where the cholinergic system has been damaged (e.g. Alzheimer’s) – but not to any significant success

Neuron activity (increased during high activity)

• Activity-dependent synthesis ensures sufficient ACh is available during periods of high activity

Question 9

Describe ACH storage

Answer 9

• Stored in vesicles at the nerve terminal by vesicular ACh transporter
• Transport is inhibited by the drug vesamicol
• Leads to cytosolic accumulation of ACh, decreased release
• Blocking vesicle transport depletes the pool of release-ready vesicles

Question 10

Describe ACH release

Answer 10

• Released by classical vesicular exocytosis (voltage-dependent Ca2+-influx)
• Release triggered by latrotoxin (black widow venom, Latrodectus mactans) in periphery
• Muscle pain, tremors, nausea, excessive sweating

Question 11

Describe ACH re-uptake

Answer 11

• ACh is broken down in the synapse to attenuate signalling
• AChE is present primarily in the synaptic cleft and breaks ACh down to choline
• Breakdown product (choline) transported by choline transporter
• Majority of choline is recycled to ACh
• Re-uptake inhibited by the drug hemicholinium-3
• Leads to synaptic accumulation of choline and run-down of activity

Question 12

Describe Botox

Answer 12

• Clostridial neurotoxins (isolated from Clostridium botulinum) cause paralytic effects of botulism poisoning
• BoNT A/B are proteases that cleave VAMP or SNAP-25 (resp.) to prevent vesicle binding
• Local injection causes muscle paralysis by inhibiting ACh release
• Therapeutic use includes treatment of crossed eyes, eyelid spasms, facial spasms

Question 13

Describe insecticides and ACH

Answer 13

Manydrugsaffectingcholinergicsystemsare insecticidal

• ACh is a primary neurotransmitter of the insect CNS
  Plantalkyloidsaresecondarymetabolites commonly synthesized for defense against insect or animal predation

SeveralplantalkyloidsaffectingAChare exploited for commercial or therapeutic benefit

Question 14

Describe drugs affecting ACH breakdown

Answer 14

• AChE inhibition prolongs ACh signalling
• AChE inhibition decreases the rate of ACh breakdown in the synaptic cleft leaving more ACh to act at postsynaptic receptors
• Plant-derived toxin physostigmine is a BBB permeable inhibitor of AChE (found in Calabar beans) * Accidental poisoning leads to slurred speech, confusion, hallucinations, loss of reflexes, convulsions, coma, death
• SyntheticBBB-impermeableanaloguesneostigmine(Prostigmin)andpyridostigmine (Mestionon) are used to treat the autoimmune disease myasthenia gravis (MG)
• Physostigmine,pyridostigmine,andneostigminearereversibleinhibitorsofAChE

Question 15

What causes Myasthenia gravis?

Answer 15

Myasthenia gravis results from immune response to ACh receptors at neuromuscular junctions – attenuating response to ACh signals. Treatment with AChE inhibitors prolongs the ACh signal and compensates for the lost function.

Question 16

Describe irreversible ACH inhibitors

Answer 16

• Organophosphate pesticides (e.g. malathion) inhibit AChE irreversibly
• Insects are more sensitive than humans though human toxicity is possible
• Effectively banned in Canada (most recently in Winnipeg)
• Sarin and VX (nerve gas) are irreversible AChE inhibitors
• Sarin used in infamous Tokyo subway attacks (1995)
• Risks of exposure in Gulf and Iraq wars
• Pyridostigmine prescribed as preventative
  agent to nerve gas poisoning
• Pyridostigmine competes for the same site on AChE as Sarin/VX thus blocking the effects
• Pyridostigmine later found to have increased BBB permeability under stress conditions
• Use linked to one component of Gulf War Syndrome
• Confusion-ataxia syndrome
• Cognitiveimpairment,dizziness, balance and coordination problems