Lecture 5: Nicotinic Receptors

Question 1

What are the features of the Neuromuscular Junction (NMJ)?

Answer 1

• Muscle fibers: The target tissue for motor neuron innervation.
• Axon bifurcation: Motor neurons branch into motor end plates or NMJs.
• Termina glutons: Subdivisions of the motor neuron endings at the NMJ.
• Fluorescent dyes: Used to label individual constituents for visualization.
• Increased surface area: Allows for the packing of more receptors on muscle fibers.
• Acetylcholinesterase: Enzyme that metabolizes acetylcholine, terminating its action.
• Voltage-gated calcium channels: High density for calcium influx, essential for vesicle fusion with the membrane.
• Nicotinic receptors: Found at high density specifically at the NMJ.

Question 2

Why is the high density of voltage-gated calcium channels important at the NMJ?

Answer 2

Facilitating calcium influx, which catalyzes vesicle fusion with the membrane, leading to neurotransmitter release.

Question 3

What role does acetylcholinesterase play at the NMJ?

Answer 3

Enzyme that metabolizes acetylcholine, terminating its action and allowing for precise control over neurotransmission at the NMJ.

Question 4

How is acetylcholine ACh synthesized at the NMJ?

Answer 4

• Acetylcholine synthesis requires acetyl coenzyme A (acetyl-CoA) and choline
• Choline is produced from the breakdown of ACh and is taken back up into the presynaptic neuron. Through the action of the enzyme acetyltransferase, acetyl-CoA combines with choline to produce ACh.

Question 5

What is the role of Choline Acetyltransferase (ChAT) in ACh synthesis?

Answer 5

• Enzyme responsible for synthesizing acetylcholine from its precursors.
• There is no selective inhibitor of this enzyme. In conditions like Alzheimer’s disease (AD), amyloid proteins that accumulate may inhibit ChAT, leading to a cholinergic deficit.

Question 6

How is ChAT used as a marker for cholinergic fibers?

Answer 6

• Staining tissue for this enzyme reveals a pattern of staining, indicating the presence of cholinergic nerve endings. This marker helps identify cholinergic pathways and assess cholinergic function in various physiological and pathological conditions.

Question 7

How is ACh stored and concentrated in synaptic vesicles?

Answer 7

• ACh is stored in synaptic vesicles to protect it from metabolizing enzymes and to concentrate it to a high level.
• AchHT (acetylcholine vesicular transporter): a specialized transporter that facilitates the transport of ACh into synaptic vesicles.
• This process is ATP-dependent but does not directly require ATP for pumping ACh.
  
  • Instead, ATP generates a gradient of hydrogen ions (H+) by pumping them up a concentration gradient. This gradient creates an acidic environment inside the vesicles.
  • ACh is then transported into the vesicles in exchange for H+ ions, utilizing the energy associated with the efflux of H+ ions down their concentration gradient. This process is facilitated by an antiporter mechanism.

Question 8

What is the significance of the acetylcholine vesicular transporter (AchHT)?

Answer 8

• Crucial for the packaging of acetylcholine into synaptic vesicles.
• Maintains the cholinergic neurotransmission at the neuromuscular junction and other cholinergic synapses. Additionally, similar mechanisms are involved in the storage of other neurotransmitters in synaptic vesicles.

Question 9

How does vesamicol affect ACh storage in synaptic vesicles?

Answer 9

• Vesamicol is a compound that can poison the vesicular transporters responsible for neurotransmitter storage.
• Specifically, vesamicol inhibits the AchHT, leading to a disruption in the storage of ACh in synaptic vesicles. This results in a gradual development of muscular paralysis. Vesamicol is primarily used experimentally for research purposes and is not employed clinically.

Question 10

What are the key steps involved in neurotransmitter release from the nerve terminal?

Answer 10

• Invasion of an action potential (AP) into the nerve terminal.
• The AP is dependent on voltage-gated sodium channels, leading to rapid depolarization of the membrane.
• This depolarization activates voltage-gated calcium channels, resulting in the influx of calcium ions into the presynaptic terminal
• The release of neurotransmitters is triggered by this influx of calcium ions.

Question 11

How is the action potential (AP) terminated in the nerve terminal?

Answer 11

• The action potential (AP) in the nerve terminal is terminated through two mechanisms:
  
  • Inactivation of voltage-gated sodium channels via depolarization.
  • Opening of voltage-gated potassium channels, causing the efflux of potassium ions from inside the cell and bringing the membrane potential back to baseline.

Question 12

What is the mechanism of action of TTX (tetrodotoxin) and its significance?

Answer 12

• TTX is obtained from pufferfish and is a potent blocker of voltage-gated sodium channels.
• It effectively inhibits the influx of sodium ions during depolarization → preventing the generation of action potentials.
• TTX has a similar mechanism of action to local anesthetics. Its significance lies in its use as a tool in research to block sodium channels and study the role of action potentials in various physiological processes.

Question 13

How does conotoxin affect neurotransmitter release, and what is its source?

Answer 13

• Conotoxin is obtained from predatory snails, and it blocks potassium channels.
• Inhibiting potassium channels → interferes with the repolarization phase of the action potential → prolonged depolarization → preventing the normal release of neurotransmitters [reduced calcium influx]

Question 14

What is the mechanism of action of botulinum toxin, and how is it used clinically?

Answer 14

• Botulinum toxin is derived from bacteria and blocks vesicle fusion by acting as a protease that cleaves proteins involved in vesicular fusion.
• This prevents the release of neurotransmitters, leading to muscular paralysis.
• Clinically, botulinum toxin (Botox) is used for various therapeutic and cosmetic purposes, including the treatment of muscle spasms and the reduction of facial wrinkles.

Question 15

How do dendrotoxins function, and what is their source?

Answer 15

• Block voltage-gated potassium channels
• Broadens action potentials → increased calcium influx and enhanced neurotransmitter release.
• Sources: green mamba snake → cause muscular spasms and paralysis in prey.

Question 16

How do proteins associated with synaptic vesicles contribute to neurotransmitter release?

Answer 16

Proteins associated with synaptic vesicles interact with membrane-bound proteins, facilitating the fusion of synaptic vesicles with the plasma membrane. This interaction tightens the bonds between the vesicles and the membrane, bringing them into close proximity. As a result, fusion occurs, leading to the release of neurotransmitters, such as acetylcholine (ACh), into the synaptic cleft.

Question 17

What is the role of synaptotagmin in neurotransmitter release?

Answer 17

• Synaptotagmin serves as a calcium sensor during neurotransmitter release
• When there is an increase in intracellular calcium concentration associated with the influx of action potentials and activation of voltage-gated calcium channels, synaptotagmin undergoes a conformational change
• This change tightens the protein strands, bringing the vesicular membrane in close proximity to the plasma membrane, thereby facilitating vesicle fusion and neurotransmitter release

Question 18

How does botulinum toxin interfere with neurotransmitter release?

Answer 18

• Botulinum toxin binds to synaptotagmin, which is exposed on the surface of synaptic vesicles.
  
  • Once bound, the toxin is internalized into the cell through endocytosis of the synaptic vesicle.
• Within the cell, botulinum toxin undergoes cleavage between its heavy and light chains.
  
  • The light chain then escapes from the synaptic vesicle and acts as a protease, cleaving proteins associated with vesicular fusion.
  • Disruption in vesicular fusion → flaccid paralysis (can be fatal in severe cases of botulinum toxin poisoning)

Question 19

In what conditions is a reduction in muscle activity desired?

Answer 19

• Reduction in muscle activity is desired in various conditions, including:
  
  • Dystonias: muscular spasms (e.g. blepharospasm → involuntary twitching of the eye muscles)
  • Muscle spasticity: increased muscle tone and stiffness, often resulting from neurological conditions like cerebral palsy or stroke.
  • Tremor: involuntary rhythmic movements of one or more body parts, such as essential tremor or Parkinson’s disease tremor.
  • Sialorrhea (drooling) and hyperhidrosis (excessive sweating): conditions where excessive secretion occurs, leading to unwanted symptoms.
  • Botox (cosmetics): botulinum toxin (Botox) injections are commonly used in cosmetic procedures to reduce muscle activity in facial muscles, thereby reducing the appearance of wrinkles and fine lines.

Question 20

What are the effects of α-LTX (α-Latrotoxin) on neuromuscular junction (NMJ) activity?

Answer 20

• α-LTX, found in black widow spider venom, induces significant alterations in NMJ activity:
  
  • Massive Release of ACh: α-LTX leads to an excessive release of acetylcholine (ACh) from synaptic vesicles into the synaptic cleft
  • Electrophysiological Readings: Following α-LTX exposure, electrophysiological recordings become chaotic or messy, likely due to the overwhelming synaptic activity.
  • Muscle Spasms: The excessive ACh release induced by α-LTX can cause muscle spasms or twitching due to the heightened neuromuscular activity.
• After α-LTX Exposure:
  
  • Black Widow Spider Venom Effect: Components of black widow spider venom, such as α-LTX, interfere with endocytosis, the process of reuptake of synaptic vesicles after neurotransmitter release.
  • Emptying of Synaptic Vesicles: As a result, synaptic vesicles remain bound to the plasma membrane instead of being internalized, leading to the depletion or emptiness of synaptic vesicles within the presynaptic terminal.
  • Prolonged Paralysis: With the depletion of synaptic vesicles, there is a cessation or significant reduction in neurotransmitter release, resulting in prolonged paralysis of the affected muscle.

Question 21

What was observed in the Nature paper

Answer 21

• AIM: investigate how neurotransmitter was released at the synaptic vesicles.
• METHODOLOGY: synaptic vesicles were observed using electron microscopy (EM).
• OBSERVATIONS
  
  • Deflections were observed after black widow spider venom (BWV) application.
  • Over time, there was an increase in the frequency of events.
• CONCLUSION: each event observed corresponded to the release of one synaptic vesicle.

Question 22

How does dimer formation affect neurotransmitter release?

Answer 22

• Dimer formation: release of calcium from intracellular stores.
• Increases voltage-gated calcium channel activity → increase in calcium concentration at synaptic vesicle release sites and terminals → more efficient vesicular release into the synaptic cleft

Question 23

What role does tetramer formation play in ALTX action?

Answer 23

• Tetrameric ALTX binds to calcium channels and embeds itself into their structure → alters the properties of the calcium channel, rendering it permeable to calcium ions.
• The influx of calcium ions through these modified channels triggers vesicular fusion with the presynaptic membrane, facilitating neurotransmitter release into the synaptic cleft.

Question 24

How do both dimer and tetramer formations contribute to neurotransmitter release?

Answer 24

• Dimer-mediated calcium release enhances calcium influx through voltage-gated channels
• Tetramer-induced modification of calcium channels directly facilitates calcium entry, ultimately promoting neurotransmitter release.

Question 25

What is the relationship between vesicle release and neurotransmitter transmission at the neuromuscular junction (NMJ)?

Answer 25

Vesicle release results in the release of a quanta of neurotransmitter.

Question 26

What is the significance of miniature end plate potentials (mEPPs) in neurotransmission at the NMJ?

Answer 26

mEPPs summate to generate the end plate potential (EPP), which is necessary to depolarize the muscle membrane and initiate an action potential.

Question 27

How does the end plate potential (EPP) contribute to muscle contraction?

Answer 27

If the EPP reaches a sufficient magnitude, it depolarizes the muscle membrane to initiate an action potential, which spreads across the muscle fiber and triggers the release of calcium ions, leading to muscular contraction.

Question 28

What is the role of acetylcholinesterase (AChE) in neurotransmission?

Answer 28

Breaks down ACh into acetate and choline, terminating the action of ACh at the synaptic junction.

Question 29

How is choline recycled in the process of acetylcholine (ACh) synthesis?

Answer 29

Taken back up for the synthesis of new ACh.

Question 30

How do compounds targeting acetylcholinesterase (AChE) affect neurotransmission?

Answer 30

Compounds like anti-AChE drugs (e.g., nerve, neostigmine) inhibit acetylcholinesterase, leading to increased concentrations and effects of acetylcholine (ACh), which can result in muscle spasms.

Question 31

What is the mechanism of competitive non-antagonist (non-depolarizing) blockers?

Answer 31

Block nicotinic receptors at the NMJ → increase in the concentration of ACh at the NMJ.

Question 32

How does tubocurarine induce muscular paralysis?

Answer 32

Blocks the generation of AP at the NMJ

Question 33

How does tubocurarine’s effect on AP generation change in the presence of ACh?

Answer 33

ACh outcompetes tubocurarine, allowing AP generation.

Question 34

What characterizes irreversible compounds like α-bungarotoxin?

Answer 34

• They covalently bind to nicotinic receptors at the NMJ.
• They act as antagonists, preventing ACh from binding to the receptor.

Question 35

How does Tubocurarine affect prey?

Answer 35

• Paralyzes prey, causing muscular paralysis.
• Tubocurarine can also cause respiratory paralysis.

Question 36

What clinical applications did Tubocurarine have?

Answer 36

• Tubocurarine was used clinically, particularly in surgery.
• It was noted for causing a decrease in blood pressure by blocking the ganglion, leading to vasodilation.

Question 37

How do Vecuronium and Rocuronium differ from anesthetics?

Answer 37

• Vecuronium and Rocuronium are not anesthetics; they solely induce paralysis without causing anesthesia.
• Patients administered with these drugs remain conscious and may feel pain.

Question 38

What advantages do Vecuronium and Rocuronium offer in surgery?

Answer 38

• These drugs allow for the reduction of the concentration of gas used during anesthesia.
• They have few side effects compared to other drugs.
• Their effects can be reversed using acetylcholinesterase inhibitors like neostigmine.

Question 39

What are Depolarizing blockers?

Answer 39

Agonists of nicotinic receptors at the neuromuscular junction

Question 40

Describe the mechanism of action of Suxamethonium during Phase I.

Answer 40

• During Phase I, Suxamethonium persistently activates the nicotinic receptors at the NMJ.
• It is not broken down and remains at the NMJ, continuously activating nicotinic receptors.
• This leads to the depolarization of the end plates, activating voltage-gated sodium channels necessary for generating action potentials (APs).
• Ultimately, this results in muscular paralysis.

Question 41

What characterizes Phase II of Suxamethonium action?

Answer 41

• Phase II is characterized by the persistent activation of the nicotinic receptors leading to desensitization.
• Continuous activation of the receptor causes desensitization, meaning the receptor can no longer respond to acetylcholine or Suxamethonium.
• This desensitization leads to the repolarization of the muscle membrane, as no more sodium ions enter the cell due to receptor inactivation.
• Ultimately, this maintains blockade and paralysis of the muscle.

Question 42

What are the clinical uses of Suxamethonium?

Answer 42

• Suxamethonium is used clinically for procedures requiring rapid onset muscle relaxation.
• It is particularly useful in tracheal intubation to facilitate airway management in patients.
• Additionally, it is employed in electroconvulsive therapy (ECT) to prevent muscular spasms associated with the procedure.

Question 43

What are the characteristics of Suxamethonium’s action?

Answer 43

• Suxamethonium has a rapid onset of action, making it suitable for situations where quick muscle relaxation is required.
• It is broken down by plasmacholinesterases, contributing to its short duration of action.

Question 44

What are the side effects associated with Suxamethonium?

Answer 44

• Bradycardia [effects on muscarinic receptors in the heart → a slowing of the heart rate]
• In cases of trauma or extensive muscle damage, Suxamethonium administration may cause the release of potassium ions from cells, potentially leading to hyperkalemia, cardiac dysrhythmias, and even cardiac arrest.
• Prolonged paralysis can also occur as a side effect of Suxamethonium administration.

Question 45

What types of receptors are present at ganglia in the autonomic nervous system (ANS)?

Answer 45

Ganglia in the ANS contain nicotinic receptors, which are composed of alpha and beta subunits.

Question 46

Describe the transmission between preganglionic and postganglionic neurons at autonomic ganglia.

Answer 46

• The preganglionic neuron releases acetylcholine (ACh) at the ganglia’s synaptic junction, which binds to nicotinic receptors on the postganglionic neuron.
• This binding of ACh leads to the generation of an action potential (AP) in the postganglionic neuron.

Question 47

What happens if the ganglia in the autonomic nervous system are blocked?

Answer 47

Blocking ganglia in the ANS reduces both sympathetic and parasympathetic actions [transmission of signals between preganglionic and postganglionic neurons is inhibited]

Question 48

Describe kappa-bungarotoxin

Answer 48

• It is an irreversible antagonist that targets ganglionic nicotinic receptors.
• Selective for ganglionic nicotinic receptors.

Question 49

What is the mechanism of action of trimethaphan?

Answer 49

• Competitive antagonist.
• It is occasionally used in surgery.

Question 50

Explain the difference between hexamethonium and tubocurarine in terms of their mechanism of action.

Answer 50

• Tubocurarine is competitive at the NMJ but non-competitive at the ganglionic nicotinic receptor.
• Hexamethonium, once used for high blood pressure, affects both the NMJ and ganglionic nicotinic receptors.
• However, its side effects include wiping out the actions of the autonomic nervous system.

Question 51

What are nicotine and lobeline?

Answer 51

• Lobeline is selective for ganglionic and chromaffin cells nicotinic acetylcholine receptors (nAChR).
• Both are nAChR agonists.

Question 52

How do nicotine and lobeline affect receptors?

Answer 52

They repeatedly stimulate receptors → inactivation of voltage-gated Na+ channels and desensitization of nAChR receptors.

Question 53

Why are nicotine and varenicline used clinically?

Answer 53

• They are used for tobacco addiction treatment.
• They provide the “buzz” associated with nicotine without exposure to additional harmful chemicals.

Question 54

Does suxamethonium affect ganglionic nicotinic receptors?

Answer 54

No