Lecture 3

What is acetylcholine (ACh)?

Acetylcholine (ACh) is a neurotransmitter.

What physiological functions and diseases is acetylcholine (ACh) involved in?

ACh is implicated in a range of physiological functions and diseases.

Physiological functions include neuromuscular junctions, gastrointestinal regulation, cardiovascular regulation, lung function, and bladder control.

Diseases associated with ACh include Alzheimer's, schizophrenia, Parkinson's, and addiction.

What are the steps involved in acetylcholine synthesis and storage?

Choline enters the cell through a Na+ channel alongside Na+. It is then turned into ACh by the enzyme Acetyl CoA. The vesicular ACh transporter located on the vesicle membrane pumps ACh into vesicles to be stored for later use.

What happens during acetylcholine release?

When an action potential runs down and depolarises the neuron, it opens voltage-gated Ca2+ channels. This causes movement of vesicles to the axon terminal, where they fuse with the plasma membrane and release ACh into the synaptic cleft.

How does acetylcholine activate receptors?

ACh can bind both presynaptically and postsynaptically to nicotinic ionotropic or muscarinic metabotropic receptors.

How is acetylcholine deactivated and how are ACh receptors regulated?

ACh esterases break down ACh into its constitutive components, choline and acetyl group (precursors), which can then be taken up by the presynaptic cell to be recycled. ACh can also bind to pre- and postsynaptic receptors and cause them to desensitise and internalise, decreasing ACh signalling.

What are the ways to modulate the acetylcholine system?

The ACh system can be modulated by: 1. Altering synthesis. 2. Altering release. 3. Altering the time spent in the synapse. 4. Targeting ACh receptors.

How can acetylcholine synthesis be altered?

Increased dietary choline intake can theoretically lead to higher ACh levels.

How can acetylcholine release be altered?

Botulinum toxin prevents ACh-containing vesicles from fusing with the membrane, inhibiting ACh release.

How can the time acetylcholine spends in the synapse be altered?

Acetylcholinesterase inhibitors block the breakdown of ACh in the synapse, leading to increased and prolonged ACh activity.

What are the two main types of acetylcholine receptors?

The two main types of ACh receptors are nicotinic receptors and muscarinic receptors. Receptor expression differs throughout the brain and between different cell types within the same brain region.

What are nicotinic acetylcholine receptors?

Nicotinic receptors are ligand-gated ion channels (LGICs) that bind nicotine and ACh. There are many different arrangements of 17 different subunits, grouped into 4 large families. They increase neuronal excitability by allowing influx of Ca2+.

What are muscarinic acetylcholine receptors?

Muscarinic receptors are G-protein coupled receptors (GPCRs) that bind muscarine and ACh. There are 5 key receptors: M1–M5.

What are the functions of odd-numbered muscarinic receptors (M1, M3, M5)?

Odd-numbered muscarinic receptors (M1, M3, M5) are Gq-linked GPCRs. They activate PLC and are therefore involved in gene regulation.

What are the functions of even-numbered muscarinic receptors (M2, M4)?

Even-numbered muscarinic receptors (M2, M4) are Gi-linked GPCRs. They inhibit cyclic AMP production and decrease neuronal excitability.

Why is selective targeting of muscarinic receptors important?

Each muscarinic receptor has distinct physiological effects, so there needs to be a way to selectively target specific receptors.

What is an ideal drug target for schizophrenia involving muscarinic receptors?

The ideal drug target for schizophrenia is to agonise M1 and M4 receptors and antagonise M5 receptors.

What is xanomeline and how does it act on muscarinic receptors?

Xanomeline is an M1 and M4 agonist that is relatively selective for M4 and M1, with a ~30-fold preference in potency and an additional preference in efficacy.

What are the limitations of xanomeline?

Despite being relatively selective, xanomeline has many side effects, primarily due to actions of peripheral muscarinic receptors. The desired actions are in the brain, but peripheral actions cause severe gastrointestinal, salivary, and cardiovascular issues.

What is Cobenfy and how does it work?

Cobenfy is a combination drug containing xanomeline and trospium chloride. Xanomeline is an agonist that can pass the BBB, while trospium chloride is a non-selective muscarinic antagonist that cannot pass the BBB. Xanomeline reaches the brain and activates the wanted M1 and M4 receptors, while trospium chloride antagonises xanomeline in the periphery, stopping negative side effects.

When was Cobenfy FDA approved and for what condition?

Cobenfy was FDA approved in 2024 for schizophrenia.

What are orthosteric ligands?

Orthosteric ligands bind to the same site as the endogenous neurotransmitter. Because this binding site is highly conserved across related receptor subtypes, drugs often activate or block multiple receptor types, increasing the risk of off-target effects.

Why are orthosteric ligands difficult to make selective?

Orthosteric ligands bind to conserved binding sites, making it difficult to create ligands that specifically target closely related receptors without affecting other receptor types.

What is meant by blanket targeting of orthosteric ligands?

Orthosteric drugs target all receptors they can bind to until they are eliminated, producing a strong on or off response.

When are orthosteric ligands useful?

Orthosteric ligands are useful when the endogenous ligand is lost, such as in Parkinson's disease.

Why do orthosteric agonists reduce spatial and temporal precision of neurotransmission?

Orthosteric agonists bind to highly conserved binding sites, so they often activate multiple closely related receptor subtypes, resulting in widespread (blanket) receptor activation. This overrides the normal spatial and temporal control of neurotransmission, reducing the precision of endogenous neurotransmitter signalling.

What are allosteric ligands?

Allosteric ligands bind to sites outside the conserved orthosteric pocket, allowing drugs to selectively target specific receptor subtypes and reduce unwanted side effects.

What are the advantages of allosteric ligands?

Allosteric ligands tweak endogenous signalling, maintaining the spatial and temporal nature of the ligand. They provide greater selectivity and can fine-tune endogenous signalling patterns for therapeutic gain.

When are allosteric ligands useful?

Allosteric ligands are useful when the endogenous ligand is preserved, such as in Alzheimer's disease.

How do allosteric modulators affect receptor activity?

Allosteric modulators can alter the affinity of orthosteric ligands, alter the efficacy of orthosteric ligands, or activate the receptor alone.

What is a positive allosteric modulator (PAM)?

A positive allosteric modulator (PAM) binds to an allosteric site and enhances the receptor's response to the endogenous ligand. PAMs can have their own agonist activity (PAM-agonist) or have no agonist activity (ago-PAM).

What is a negative allosteric modulator (NAM)?

A negative allosteric modulator (NAM) binds to an allosteric site and reduces the receptor's response to the endogenous ligand, decreasing receptor activity.

What is a neutral allosteric ligand (NAL)?

A neutral allosteric ligand (NAL) binds to an allosteric site but does not alter receptor activity or the response to the endogenous ligand. Instead, it blocks other allosteric modulators from binding.

Why does allostery provide opportunities for targeting muscarinic receptors?

Muscarinic receptors have highly conserved orthosteric binding pockets, so allostery offers an opportunity to develop more specific compounds.

What is VU0467319?

VU0467319 is a PAM designed for the M1 receptor. It has little effect as an agonist alone, meaning it is a PAM rather than an ago-PAM. It increases the potency of ACh at the M1 receptor and is selective, having no effect on ACh activity at other receptors