L05 Clinical Pharmacology Neuromuscular blockers

Overview

The lecture works from the normal signalling machinery of the neuromuscular junction to the drugs that interrupt it. It first sets out the transmitter (acetylcholine) and its receptor (the nicotinic acetylcholine receptor), then the acetylcholine cycle from synthesis through vesicular loading, calcium-triggered exocytosis, receptor binding and breakdown by acetylcholinesterase. Against that background it covers the two postsynaptic blocking mechanisms, non-depolarising (competitive antagonism) and depolarising (agonist-induced maintained depolarisation and desensitisation), and finishes with clinical use in surgery and rapid intubation, including reversal, side effects and a comparison of onset, duration and termination for the agents in current use.

Objectives of the lecture

1. The receptors and signalling molecules present at the neuromuscular junction: acetylcholine and the nicotinic receptor.
2. The mechanism of action of neuromuscular blockers: non-depolarising and depolarising.
3. Clinical applications of neuromuscular blockers: surgery and intubation.

Where the neuromuscular junction sits in the nervous system

• The nervous system divides into central and peripheral; the peripheral division splits into autonomic and somatic, and the autonomic further into sympathetic, parasympathetic and enteric.
• Transmitter and receptor arrangement from CNS to effector organs:
  • Somato-motor: CNS → ACh → nicotinic receptors → skeletal (striated) muscle.
  • Parasympathetic: CNS → ACh → nicotinic receptors at the ganglion → ACh → muscarinic receptors on effector organs (glands, eyes, heart, bronchi, vessels, G.I.).
  • Sympathetic: CNS → ACh → nicotinic receptors at the ganglion → noradrenaline → the same effector organs.
  • Sympathetic branch to the adrenal gland: CNS → ACh → nicotinic receptors → adrenal gland → release of adrenaline.

The nicotinic receptor as an ionotropic receptor

• Ionotropic receptors are involved mainly in fast synaptic transmission, and comprise a receptor together with an associated ion channel.
• The receptor contains 5 subunits (pentameric) with about 20 transmembrane segments arranged around an aqueous channel.
• ACh binds the nicotinic acetylcholine receptor (nACh) and causes an influx of Na${}^{+}$ ions into the cell.
• Channel architecture: the pentamer sits in the membrane with a synaptic (extracellular) face and a cytoplasmic face, subunits arranged around a central channel, ACh binding at the synaptic face.
• Subunit stoichiometry across subtypes:
  • Muscle type: γ, α1, α1, δ, β1, with two ACh-binding interfaces.
  • Neuronal heteromeric α4β2: α4, α4, β2, β2, β2, with two binding sites.
  • Neuronal homomeric α7: five α7 subunits, with five binding sites.
• Kinetics of a single signalling event:
  • ACh binds the receptor within 100 µsec of release.
  • The channel opens and allows 25 million Na${}^{+}$ ions per second to enter the cell.
  • The channel closes rapidly, within 1 ms.

Acetylcholine: structure, synthesis, storage and release

• Structure: a quaternary trimethylammonium group (N${}^{+}$ bonded to three CH${}_3$ groups) joined by a two-carbon ethylene bridge to an ester oxygen linked to a carbonyl and a terminal methyl, that is an acetate ester.
• Biosynthesis: acetyl-CoA + choline → acetylcholine, catalysed by choline acetyltransferase.
• Vesicular loading: after synthesis ACh is packaged into vesicles by active uptake through a specific transporter (an ACh carrier). Vesicles normally contain 10,000 to 50,000 ACh molecules, a concentration of 100 mmol/l.
• Release: when an action potential invades the presynaptic terminal, voltage-sensitive Ca${}^{2+}$ channels open, giving a large Ca${}^{2+}$ influx into the terminal. Ca${}^{2+}$ influx leads to vesicle mobilisation, fusion to the membrane and finally exocytosis.
• Termination by acetylcholinesterase: ACh is rapidly broken down at the NMJ by ACh esterase, which hydrolyses the ester bond to release choline (inactive) and acetate. Hydrolysis is very rapid, with all ACh convertible within 1 ms.

The acetylcholine cycle

The cycle spans the presynaptic terminal and the postsynaptic membrane in six steps:

1. Synthesis from SCoA by enzyme E1 in the presynaptic terminal.
2. Loading of ACh into a vesicle.
3. Transport of the vesicle to the presynaptic membrane and release of ACh.
4. ACh crosses the synapse and binds a receptor on the postsynaptic membrane.
5. The receptor-bound complex is processed at the postsynaptic membrane.
6. Product is returned and recycled into the presynaptic terminal, closing the cycle.

Warning

The acetylcholine cycle figure sits over a faint background of unrelated body text (the page appears reused or scanned from a textbook figure) that is illegible and not part of the diagram’s own labelling. Individual step labels beyond the numbered boxes and generic markers (E1, ACh, SCoA, vesicle icon) could not be read with confidence.

Action at the neuromuscular junction

• Na${}^{+}$ flux entering the cell results in depolarisation.
• In skeletal muscle this depolarisation is termed the end-plate potential.
• If the end-plate potential reaches a sufficient threshold, the muscle fibre contracts.

Where drugs can interrupt transmission

The junctional pathway and the agents that block each step:

1. Nerve action potential — blocked by tetrodotoxin, batrachotoxin and local anaesthetics.
2. Vesicular ACh release — blocked by hemicholinium, botulinus toxin, procaine, Mg${}^{2+}$, 4-aminopyridine and lack of Ca${}^{2+}$; excess Ca${}^{2+}$ promotes release.
3. Depolarisation (end-plate potential, increased permeability to Na${}^{+}$ and K${}^{+}$) — blocked by curare alkaloids and snake α-toxins; succinylcholine and decamethonium also act at this step.
4. Hydrolysis of ACh by AChE — blocked by cholinesterase inhibitors.
5. Muscle action potential — promoted by Ca${}^{2+}$ and veratridine, blocked by quinine and tetrodotoxin.
6. Spread of excitation in muscle, then muscle contraction — contraction blocked by metabolic poisons, lack of Ca${}^{2+}$, procaine and dantrolene.

Summarised as three points of attack, neuromuscular block can be achieved by:

• Inhibiting ACh release presynaptically.
• Inhibiting ACh esterase.
• Blocking the action of ACh postsynaptically.

A postsynaptic neuromuscular block can be achieved by either:

• Blocking ACh receptors — non-depolarising block, produced by a competitive antagonist such as tubocurarine (a bis-quaternary ammonium alkaloid with two aromatic ring systems joined by ether and methylene bridges, carrying OH and OCH${}_3$ substituents).
• Over-activating ACh receptors — depolarising block, produced by an agonist such as decamethonium, a simple linear bis-quaternary ammonium compound, (CH${}_3$)${}_3$N${}^{+}$–(CH${}_2$)${}_{10}$–N${}^{+}$(CH${}_3$)${}_3$.

Non-depolarising blocks can be overcome with ACh esterase inhibitors.

Clinical effects of neuromuscular block

Important

Neuromuscular blocks cause motor paralysis. They DO NOT affect consciousness.

• Symptoms progress gradually in the order: eye muscles → facial muscles → limbs → respiratory system.
• The respiratory muscles are the last to be affected, and therefore the first to recover.

Non-depolarising blockers

• Origin: curare is found in a variety of South American plants and was added to the tips of poisonous arrows and spears by South American Indians, causing paralysis. Mechanistic studies showed that curare blocks the NMJ rather than changing muscle contractility.
• Tubocurarine was later isolated from the plant extracts and found to be the most potent of the plant alkaloids.
• It has now been replaced with synthetic, structurally similar drugs: pancuronium, vecuronium and atracurium. These are all quaternary ammonium compounds, are not absorbed orally and will not cross the placenta, and are therefore safe to use in hunting.
• Mechanism: all clinically used non-depolarising NMBs are competitive antagonists of ACh at the motor end plate.
• Safety factor of transmission: when a nerve impulse releases ACh at the NMJ, the amount released greatly exceeds what is needed. Consequently 70-80% of postsynaptic receptors must be blocked before transmission fails.
• Applications: the main application is surgery. The blocker is given by IV injection, and must be accompanied by artificial ventilation because of the effects on the lung.
• Maintaining a reversible block for surgery: a low dose of anaesthetic for unconsciousness, an analgesic for pain relief, and a neuromuscular blocker to prevent reflex movement. Following surgery the block is reversed by adding an acetylcholinesterase inhibitor, for example neostigmine.

Depolarising blockers

• In initial studies with the paralysis-inducing drug decamethonium, muscle spasms were noted before paralysis, in contrast to tubocurarine, which causes flaccid paralysis.
• Decamethonium was later found to cause paralysis by maintained depolarisation of the muscle.
• Because it acts as an agonist, the block cannot be reversed by acetylcholinesterase inhibitors.

Phase I depolarisation block

• Repeated activation of the receptor causes loss of membrane potential (depolarisation) in the postsynaptic cell.
• The muscle fibre no longer responds to ACh stimulation.
• On the membrane potential trace: baseline −70 mV with an AP threshold line, several repeated spikes reaching threshold, then a sustained plateau at 0 mV labelled Phase I block, followed by a gradual decline toward baseline.

Phase II depolarisation block

• Persistent activation of the receptor causes receptor desensitisation, a biochemical adaptation to a prolonged stimulus.
• The receptor is “turned off” by β arrestin binding, and β arrestin then acts as a target for receptor internalisation.
• Receptor activity only returns when the receptor is recycled or re-synthesised.
• On the same style of trace, Phase II block corresponds to the declining, return-to-baseline portion of the curve.

Suxamethonium

• The major depolarising NMB in clinical use, used for rapid intubation, administered by IV injection.
• Fasciculations within 1 min, followed by relaxation.
• Structure: a symmetric bis-quaternary ammonium diester, two trimethylammonium-ethyl groups each esterified to one end of a four-carbon succinate chain (succinylcholine).
• Common side effects:
  • Bradycardia at low doses (parasympathetic activation).
  • Tachycardia at high doses (sympathetic action at ganglia).
  • Muscle pain due to excessive contraction.
  • Hyperkalaemia due to excessive muscle contraction.
  • Intracranial tension (neck muscle contraction).

Comparison of clinical neuromuscular blockers

Drug	Onset (min)	Duration (min)	Termination	Mechanism
Suxamethonium	1	5-10	Plasma cholinesterases	Depolarising
Vecuronium	2-3	25-50	Renal and hepatic clearance	Competitive
Rocuronium	0.5-2	35-75	Hepatic clearance	Competitive
Pancuronium	3-4	85-100	Renal and hepatic clearance	Competitive

Rocuronium is highlighted for emphasis in the table.

Self-test

1. Define an ionotropic receptor and describe the subunit architecture of the nicotinic acetylcholine receptor.
2. State the subunit stoichiometry of the muscle-type nicotinic receptor and the number of ACh-binding sites it carries, and contrast this with the homomeric α7 neuronal receptor.
3. Give the three kinetic figures the lecture quotes for a single ACh signalling event at the receptor.
4. Describe the steps of acetylcholine synthesis, vesicular loading and release at the presynaptic terminal, naming the enzyme and the trigger for exocytosis.
5. What is the ACh content of a single vesicle, and what concentration does this correspond to?
6. Explain how neurotransmission at the NMJ is terminated, naming the products and the time course.
7. List the six steps of the acetylcholine cycle in order.
8. Define the end-plate potential and explain what determines whether the muscle fibre contracts.
9. List the three general ways neuromuscular block can be achieved.
10. Distinguish a non-depolarising block from a depolarising block in terms of drug action at the receptor, the example agent given for each, and whether an AChE inhibitor reverses it.
11. Describe the order in which muscle groups are affected during neuromuscular blockade, and explain which group recovers first and why.
12. What is the “safety factor of transmission”, and what proportion of postsynaptic receptors must be blocked before transmission fails?
13. Name the synthetic replacements for tubocurarine and give two pharmacokinetic properties they share.
14. Describe how a reversible block is maintained and then reversed for surgery, naming the three drug classes given and the reversal agent.
15. Distinguish Phase I from Phase II depolarisation block by mechanism.
16. Predict what happens if neostigmine is given to reverse a suxamethonium block, and explain why.
17. List the common side effects of suxamethonium, with the mechanism given for each.
18. A patient needs rapid intubation. Using the comparison table, state which agent gives the fastest onset with the shortest duration, its onset and duration, and how its action is terminated.
19. Distinguish vecuronium from rocuronium using onset, duration and route of termination.
20. Name the agents that block vesicular ACh release, and the ion whose excess promotes it.
21. Integrative: trace an impulse from nerve action potential to muscle contraction, naming at each step one class of agent that blocks it, and state at which step tubocurarine and suxamethonium act.

Answers

Reveal answers

1. An ionotropic receptor is involved mainly in fast synaptic transmission and comprises a receptor with an associated ion channel. The nicotinic ACh receptor is pentameric, with 5 subunits and about 20 transmembrane segments arranged around an aqueous channel; ACh binds at the synaptic (extracellular) face and causes Na${}^{+}$ influx.
2. Muscle type is γ, α1, α1, δ, β1 with two ACh-binding sites. The neuronal α7 receptor is homomeric, five α7 subunits, with five binding sites.
3. ACh binds the receptor within 100 µsec of release; the open channel passes 25 million Na${}^{+}$ ions per second; the channel closes within 1 ms.
4. Acetyl-CoA and choline are combined by choline acetyltransferase to form ACh. ACh is then actively taken up into vesicles by a specific transporter (ACh carrier). When an action potential invades the presynaptic terminal, voltage-sensitive Ca${}^{2+}$ channels open, and the large Ca${}^{2+}$ influx causes vesicle mobilisation, fusion with the membrane and exocytosis.
5. Between 10,000 and 50,000 ACh molecules per vesicle, corresponding to 100 mmol/l.
6. ACh esterase hydrolyses the ester bond of ACh at the NMJ, releasing choline (inactive) and acetate. It is very rapid: all ACh can be converted within 1 ms.
7. (1) Synthesis from SCoA by enzyme E1 in the presynaptic terminal; (2) loading of ACh into a vesicle; (3) vesicle transported to the presynaptic membrane and ACh released; (4) ACh crosses and binds a postsynaptic receptor; (5) the receptor-bound complex is processed at the postsynaptic membrane; (6) product returned and recycled into the presynaptic terminal.
8. Na${}^{+}$ entry depolarises the postsynaptic cell, and in skeletal muscle this depolarisation is called the end-plate potential. If it reaches a sufficient threshold the muscle fibre contracts.
9. Inhibiting ACh release presynaptically; inhibiting ACh esterase; blocking the action of ACh postsynaptically.
10. Non-depolarising block is competitive antagonism at the ACh receptor, example tubocurarine, and it can be overcome with an ACh esterase inhibitor. Depolarising block is over-activation of the receptor by an agonist, example decamethonium (or suxamethonium), and because the drug is an agonist the block cannot be reversed by acetylcholinesterase inhibitors.
11. Eye muscles, then facial muscles, then limbs, then the respiratory system. The respiratory muscles are affected last and are therefore the first to recover.
12. The ACh released by a nerve impulse greatly exceeds the amount needed for transmission. Because of this reserve, 70-80% of postsynaptic receptors must be blocked before transmission fails.
13. Pancuronium, vecuronium and atracurium. They are all quaternary ammonium compounds, are not absorbed orally, and do not cross the placenta.
14. A low dose of anaesthetic is given to cause unconsciousness, an analgesic for pain relief, and a neuromuscular blocker to prevent reflex movement. After surgery the block is reversed by adding an acetylcholinesterase inhibitor such as neostigmine.
15. Phase I: repeated receptor activation causes loss of membrane potential (sustained depolarisation) so the muscle fibre no longer responds to ACh. Phase II: persistent activation causes receptor desensitisation, the receptor being turned off by β arrestin binding, with β arrestin targeting the receptor for internalisation; activity returns only when the receptor is recycled or re-synthesised.
16. It would not reverse the block. Suxamethonium is a depolarising agonist, and an agonist-induced block cannot be reversed by acetylcholinesterase inhibitors; raising ACh would add to receptor activation rather than opposing it.
17. Bradycardia at low doses from parasympathetic activation; tachycardia at high doses from sympathetic action at ganglia; muscle pain from excessive contraction; hyperkalaemia from excessive muscle contraction; intracranial tension from neck muscle contraction.
18. Suxamethonium, onset 1 min, duration 5-10 min, terminated by plasma cholinesterases.
19. Vecuronium: onset 2-3 min, duration 25-50 min, terminated by renal and hepatic clearance. Rocuronium: onset 0.5-2 min, duration 35-75 min, terminated by hepatic clearance alone. Both are competitive blockers.
20. Hemicholinium, botulinus toxin, procaine, Mg${}^{2+}$, 4-aminopyridine and lack of Ca${}^{2+}$ block vesicular ACh release; excess Ca${}^{2+}$ promotes it.
21. Nerve action potential (blocked by tetrodotoxin, batrachotoxin, local anaesthetics) → vesicular ACh release (blocked by hemicholinium, botulinus toxin, procaine, Mg${}^{2+}$, 4-aminopyridine, lack of Ca${}^{2+}$) → depolarisation/end-plate potential with increased permeability to Na${}^{+}$ and K${}^{+}$ (blocked by curare alkaloids and snake α-toxins) → hydrolysis of ACh by AChE (blocked by cholinesterase inhibitors) → muscle action potential (blocked by quinine and tetrodotoxin, promoted by Ca${}^{2+}$ and veratridine) → spread of excitation in muscle → muscle contraction (blocked by metabolic poisons, lack of Ca${}^{2+}$, procaine, dantrolene). Tubocurarine (a curare alkaloid) and suxamethonium both act at the depolarisation step.