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ME.6200
Cholinergic Neuromodulation I
Prof. Gregor Rainer
Discovery of Acetylcholine - Otto Loewi 1921
He is almost as famous for the means by which the idea for his experiment came to him as he is for the experiment itself. On Easter
Saturday 1920, he dreamed of an experiment that would prove once and for all that transmission of nerve impulses was chemical, not
electrical. He woke up, scribbled the experiment onto a scrap of paper on his night-stand, and went back to sleep. The next morning he
arose very excited because he knew this dream had been very important. But he found, to his horror, that he couldn't read his midnight
scribbles. That day, he said, was the longest day of his life, as he could not remember his dream. That night, however, he had the same
dream. This time, he immediately went to his lab to perform the experiment.
Acetylcholine: Metabolism
Acetylcholine is hyrdolyzed by the membrane-associated
Acetylcholineesterase. AChE ranks as one of the most efficient enzymes, since
one molecule of the enzyme can hydrolyze 5000 molecules of acetylcholine per
second.
• Acetylcholine is formed by the combination of an Acetyl group
and Choline in the presence of the ChAT enzyme.
• Acetylcholine is metabolized by the membrane associated
enzyme AChE with very high efficiency.
• Choline is taken up into presynaptic neurons by a high affinity
carrier.
Take Home Message
Acetylcholine: Receptors
Reversal potential Na: ~+40mV
Reversal potential K: ~-75mV
Equilibrium potential typical Neuron: ~-70mV
The Nicotinic ACh Receptor
x5
There are 8 and 4 isoforms.
Note: The nAChR is permeable to Na and K and Ca ions.
The Muscarnic ACh Receptor
Phosphatidylinositoldiphosphate (PIP2)
Inositoltrisphosphate (IP3)
Diacylglycerol (DAG)
Protein Kinase C (PKC)
• There are two types of Acetylcholine receptors (AChR).
• The nicotinic receptor (nAChR) is the best studied of all
receptors. It is a ligand gated ion channel composed of five
subunits. There are many different nAChR varietes that are
specific to particular structures (7 receptor for Cortex and
Hippocampus; 12 for skeletal muscle).
• The muscarinic receptor (mAChR) is coupled to a G-protein
and activates a second messanger cascade. This cascade
triggers protein phosphorylation and Calcium release from
intracellular Calcium stores, and can lead to opening of
Potassium channels. There are five main types of mAChRs.
Take Home Message
Autonomic Nervous System I
Autonomic
Nervous System II
ACh ACh
NE
Adrenaline ACh
ACh ACh
nAChR mAChR
ACh NE, Adrenaline
nAChR adrenergicR
Sympathetic
Parasympathetic
‘rest and digest’
Stimulatory, activational
TARGET
TARGET
stimulates
constricts pupil
salivation
slows heartbeat
constricts lung
stimulates
erection
release NE: Norepinephrine
ACh: Acetylcholine
T, K, S: Thoracic,Lumbar and Sacral segments
• The autonomic nervous system is composed of a sympathetic
and a parasypmpathetic part. Both use (nictonic) Ach in
preganglionic neurotransmission, and the parasympathetic
system also uses (muscarinic) Ach transmission in
postganglionic neurotransmission.
• Sympathic modulation is activational and important for „fight
and flight“; parasympathetic modulation is important for body
maintenance in the resting state („rest and digest“).
• In parasympathetic modulation, ACh: constricts pupils,
stimulates tear glands, induces salivation, constricts lungs,
slows heartbeat, stimulates digestion, causes erections in males
(by vasodilation) and retention of urine.
• Generally, the activation of a particular target is determined by
the relative strength of parasympathetic and sympathetic
pathways.
Take Home Message
Neuromuscular Junction
• When an action potential reaches a neuromuscular synapse,
voltage-gated Ca channels open, and the inflow of Ca triggers
ACh release.
• ACh binds to nAChR on the motor endplate, Na+ moves into
the muscle cell (and less K+ moves out), leading to a local
depolarization of the motor end plate. This depolarization
spreads across the surface of the muscle fiber into transverse
tubules, eliciting the release of calcium and initiating muscle
contraction.
• Myasthenia gravis is an autoimmune neuromuscular disease
characterised by muscle weakness. It is caused by circulating
antibodies that block nAChR receptors at the neuromuscular
junction, and by a reduction in the number of nAChRs.
Take Home Message
Cholinergic System Agonists
Direct receptor agonists
Have affinity for nAChR or mAChR, mimic ACh
Act mostly on mAChRs
Exception: Nicotine
AChE inhibitors
Inhibit AChEsterase, increasing Ach
Triggers activation of both nAChR and mAChR
Reversible AChE inhibitors for medical use; irreversible AChE
inhibitors are used as nerve gases and insecticides.
The extremely short half-life of ACh makes it
therapeutically useless.
Carbachol in not hydrolyzed by AChE. Activates
mAChR and nAChR. Does not pass blood brain
barrier.
Bethanecol is not hydrolyzed by AChE. Does not
activate nAChR. Does not pass blood brain
barrier.No cardiovascular effects.Treatment of
urinary retention.
Pilocarpine chief alkaloid in pilocarpus jabordani.
Treatment of glaucoma.
Bethanechol
Cholinergic receptor agonists
Systemic injection of Pilocarpine can
compromise the blood-brain barrier
allowing the pilocarpine to gain
access to the brain. This can lead to
chronic epilepsy.
• Exaggeration of all symptoms of muscarinic agonism
• Significance: High consumption of wild mushrooms (culinary)
• 30-60 minutes, salivation, lacrimation, excessive sweating, nausea, vomiting, diarrhea, bronchospasm, headache, visual disturbances, abdominal colic, bradychardia, hypotension, shock
• ALL SYMPTOMS REVERTED BY ATROPINE (mAChR receptor blocker)
Muscarine chief alkaloid in amanita muscaria.
No therapeutic application.
Cholinergic receptor agonists
• ACh is therapeutically useless due to its extremely efficient
metabolism. Direct ACh receptor agonists mimic ACh at both
nAChR and/or mAChR, but are not rapidly metabolized.
• Carbachol and Bethanecol are choline esters that are more
stable than ACh, and can be administered systemically to
support parasympathetic activation (parasympathomimetics).
• Pilocarpine is a plant alkaloid, used to treat glaucoma by
topical administration to the eye. It constricts iris muscle, and
also ciliary muscle, increasing the flow of acqueous humor from
the eye and reducing intraocular pressure.
• To counteract ACh agonist action, one may deliver an ACh
receptor antagonist such as Atropine.
• All therapeutic AChE inhibitors are reversible; irreversible
AChE inihibitors are used as insecticides and nerve gases.
Take Home Message
Reversible AChE Inhibitors
Physostigmine derived from Calabar bean
(West Africa). Passes blood brain barrier.
Used to treat Alzheimer disease. Can be
used to treat CNS effects of anticholinergic
drug overdoses.
Neostigmine synthesized first 1931.
Does not cross blood brain barrier.
Used to treat the autoimmune
neuromuscular disorder myasthenia gravis.
Edrophonium has short half life (about 5
minutes). Is used to diagnose myasthenia
gravis.
„Horny goat weed“ acts as AChE inhibitor
(active ingredient unknown).
Stimulation of vascular M3 receptors Nitric
oxide release vasodilation. Similar to
Viagra®.
• AChE inhibitors prolong the action of ACh by slowing down it‘s
metabolism, effectively increasing the extracellular
concentration of ACh.
• Physostigmine is a naturally derived AChE inhibitor that is
used to treat CNS disorders such as Alzheimer‘s disease or to
counteract CNS effects of ACh receptor blockers.
• Neostigmine and Edrophonium are synthetic AChE inhibitors,
used rescpectively to treat and diagnose Myasthenia gravis.
• The vasodilation action of AChE causes penile erections by
triggering Nitric Oxide release, and is used to counteract penile
dysfunction.
Take Home Message
Cholinergic System Antagonists
Muscarinic receptor blockers
Competitive antagonists of the mAChR receptor.
Widespread medical applications.
Nicotinic receptor blockers
Ganglion-specific blockers (no medical applications).
Neuromuscular blockers (muscle relaxants).
Atropine extracts from the Egyptian
henbane were used by Cleopatra in
the last century B.C. to dilate her
pupils, in the hope that she would
appear more alluring. In the
Renaissance, women used the juice
of the berries of Atropa belladonna
to enlarge the pupils of their eyes,
for cosmetic reasons
Atropine derived from atropa belladonna.
Passes blood brain barrier. Deadly in high dose.
Uses: prevents salivation during anesthesia,
treatment for bradycardia, eye examinations,
visual neuroscience experiments.
Muscarinic receptor blockers
Muscarinic receptor blockers
Dose dependence of Atropine action
Scopolamine is main alkaloid in datura
stramonium. Passes blood brain barrier.
Used to treat motion sickness.
Toxin Detector Theory of Seasickness
Vestibular system acts as a toxin detector
Brain has evolved to recognize any changes in expected patterns of vestibular,
visual and kinesthetic information as evidence of CNS malfunction
initiation of vomiting by cholinergic brain vomiting center stimulation as a defense
against possible ingested neurotoxin
Women are more likely to get motion sick than men.
Children are more likely to get motion sick. Peak age around 12.
Prior history of motion sickness makes it more likely to get motion sick.
Bos et al Susceptibility to seasickness. Ergonomics Jun 2007
Muscarinic receptor blockers
Butyl-Scopolamine is a semi-synthetic
derivative of scopolamine that does not pass
the blood brain barrier. Used to treat
menstrual pain.
Glycopyrrolate is a synthetic muscarinic
antagonist that does not pass the blood brain
barrier.
Muscarinic receptor blockers
• Muscarnic antagonist action can be summarized as
“DRY AS BONE, RED AS A BEET, MAD AS HATTER.”
Dry is a consequence of decreased sweating,
salivation and lacrimation - Red is a result of impaired
peripheral vasodilation, problem to dissipate heat
(hyperthermia) - Mad is a result of the CNS effects of
muscarinic inhibition which can lead to sedation,
amnesia (hypersensitivity), or hallucination
• Atropine and Scopolamine are brain-blood-barrier permeable,
naturally derived mAChR blockers (parasympatholytics), with a
wide range of applications including inhibition of salivation
during anesthesia, pupil dilation for eye exams and
neuroscience experiments, treatment of motion sickness.
• Butyl-Scopolamine and Glycopyrrolate are synthetic non brain-
blood-barrier permeable mAChR antagonists with wide
applications. They are used to control for peripheral effects in
CNS studies of BBB-permeable mAChR antagonists.
Take Home Message
Nicotinic receptor blockers
Curare is a plant-derived arrow poison in
South America. Active ingredient is d-
Tubocurarine, it blocks the neuromuscular
nicotinic receptor.
Death occurs by respiratory paralysis.
Was used clinically as muscle relaxant
during surgery. No longer used because
blood pressure drops through histamine
release.
Pancuronium and related compounds are
synthetic neuromuscular nictonic receptor
blockers causing little histamine release, and
are now used for surgical muscle relaxation.
Tubocurarine by itself has no action, it gives neither contraction nor depolarization.
Tubocurarine inhibits muscular contraction induced by stimulation of the motor nerve or by the application of
acetylcholine. It does this by reducing amplitude and duration of the end-plate potential.
The application of an acetylcholine excess or an anticholinesterase agent reduces the effects of tubocurarine.
Tubocurarine has no effect when applied away from the neuromuscular junction, on the nerve or the muscle.
Botulinum toxin (BOTOX):
is produced by anaerobic bacteria called
Clostridium botulinum.
Temporarily blocks the nerve impulses
traveling from the nerve to the muscle
thereby relaxing the muscle.
Botox interferes with vesicle exocitosis, blocking ACh release
• D-tubocurarine, a nAChR blocker specific to the
neuromuscular junction, is the active ingredient in the South
American arrow poison Curare, which kills animals by
respiratory muscle paralysis. Pancuronium and related
compounds are synthetic neuromuscular nAChR
antagonists used for muscle paralysis during surgical
procedures.
• Botox blocks ACh release from presynaptic terminals,
leading to local muscle relaxation. It does this by disrupting
the formation of SNARE complexes that are necessary for
the fusion of vesicles with the cell membrane.
Take Home Message
Botox injection on forehead impairs touch discrimination on hand
• Botox injection on forehead silences muscle acticity for
periods lasting several weeks or months. This leads to
plastic reorganization in somatosensory (touch) cortex, in a
form of maladaptive plasticity that is similar to phantom limb
related cortical reorganization. Subjects’ judgement for
touch temporal order sensitivity is impaired following Botox
administration.
Take Home Message