02 - Classification of Receptors

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TARGET DECK: MED::I::Signaling Pathways in Health and Disease::Cell Signaling::02 - Cell surface receptors and nuclear receptors

Overview

Lecture Topics

• Types of ligands & receptors
• Principles of ligand-receptor interactions
• Pathways of cellular signaling
• Molecular details of representative signal transduction systems, classified by receptor type

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Classification of Receptors

Membrane Receptors

• Ionotropic receptors (ion channels)
• Metabotropic receptors (G proteins)
• Catalytic receptors (enzymes)

Intracellular Receptors

• Nuclear receptors (e.g., steroid receptors)

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Ionotropic Receptors — Overview

Definition

Ionotropic receptors are the simplest transducers of extracellular signals (ligands). They are ligand-gated ion channels that directly open or close in response to the concentration of a signal ligand or membrane potential.

Examples:

• Nicotinic acetylcholine receptor (nAChR)
• ${\text{GABA}}_A$ receptor
• Glycine receptor
• Glutamate receptor
• Serotonin receptor

What are ionotropic receptors?

Ligand-gated ion channels that directly convert chemical signals (ligand binding) into electrical/ionic changes. They are the simplest transducers of extracellular signals.

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Ionotropic receptors are also called {1:ligand-gated ion channels} and are considered the {2:simplest} transducers of extracellular signals.

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Acetylcholine at the Neuromuscular Junction

The Cholinergic Synapse — Step by Step

1. An action potential travels down the presynaptic axon, triggering opening of voltage-gated ${\text{Ca}}^{2+}$ channels
2. ${\text{Ca}}^{2+}$ influx causes secretory vesicles containing acetylcholine to fuse and release ACh into the synaptic cleft
3. ACh diffuses across the cleft and binds to nicotinic receptors on the postsynaptic membrane (myocyte or next neuron)
4. The ligand-gated cationic channel opens (~1 ms), allowing extracellular ${\text{Na}}^{+}$ and ${\text{Ca}}^{2+}$ to enter
5. Ion influx depolarizes the postsynaptic cell (changes $V_m$), triggering muscle contraction

Role of Ions as Second Messengers

Extracellular ${\text{Na}}^{+}$ and ${\text{Ca}}^{2+}$ entering through the nicotinic channel act as intracellular second messengers, depolarizing the postsynaptic cell and affecting other membrane proteins sensitive to $V_m$.

CC(=O)OCC[N+](C)(C)C

(Acetylcholine)

What is the role of acetylcholine at the neuromuscular junction?

ACh is a key excitatory neurotransmitter that binds to nicotinic receptors (ligand-gated cationic channels) on the postsynaptic membrane, causing channel opening, Na⁺/Ca²⁺ influx, membrane depolarization, and ultimately muscle contraction.

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Nicotinic Acetylcholine Receptor (nAChR)

Found in the post-synaptic surface. The pre-synaptic neuron will produce and release acetylcholine in the cleft between the synapses (extracellular space). They will find a binding site in the post-synaptic cell, advising it to continue the link.

Acetylcholine is not an ion: it does not go through. It opens the channel for positively charged ions, sodium and calcium (specificity).

The channel is a multi-subunit protein, pentameric → $\alpha 2\beta \gamma \delta$.
The alpha subunits get their conformation changed by the binding of acetylcholine, inducing the other subunits to become “larger”.
The $\alpha$ gets rotated.

Localization

• Neuromuscular junctions
• Sympathetic and parasympathetic ganglia
• Adrenal medulla
• Central nervous system

Structure

Molecular Architecture

• Pentameric structure: ${\alpha }_2\beta \gamma \delta$ subunits
• Each subunit contains four transmembrane $\alpha$-helices (M1–M4)
• The M2 helix of each subunit lines the ion-conducting pore

Feature	Detail
Subunit composition	${\alpha }_2\beta \gamma \delta$ (pentamer)
Transmembrane helices per subunit	4 (M1–M4)
Pore-lining helix	M2
Ion selectivity	Cations (${\text{Na}}^{+}$, ${\text{Ca}}^{2+}$, ${\text{K}}^{+}$)
Ligand binding site	Two ACh molecules bind on the $\alpha$-subunits

Key Reference

Miyazawa A, Fujiyoshi Y, Unwin N. Structure and gating mechanism of the acetylcholine receptor pore. Nature. 2003, 423: 949–955.

What is the subunit composition of the nicotinic acetylcholine receptor?

The nAChR is a pentamer composed of α₂βγδ subunits, each containing four transmembrane α-helices (M1–M4). The M2 helix of each subunit lines the pore.

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The nicotinic ACh receptor has a {1:pentameric} structure composed of {2:α₂βγδ} subunits, each with {3:four} transmembrane α-helices. The pore is lined by the {4:M2} helices.

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Channel Gating Mechanism

Structural Basis of Gating

In the closed (resting) state: the M2 helices are oriented such that the side chains of five Leucine residues (one per subunit, at position Leu-257 on M2, i.e. α-Leu257) form a hydrophobic ring at the middle of the membrane, physically occluding the channel pore.

Upon binding of 2 ACh molecules to the $\alpha$-subunits, an allosteric conformational change is induced: the M2 helices rotate, moving the Leu side chains away from the pore center and exposing polar residues, thereby opening the passage to cations.

What is the structural basis of nAChR channel gating?

In the resting state, five Leu residues (one per M2 helix, α-Leu257) form a hydrophobic ring that closes the pore. ACh binding induces an allosteric rotation of the M2 helices, moving the Leu side chains away from the center and exposing polar residues, thus opening the channel to cations.

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Functional States of nAChR

State	ACh Binding	Pore	Conductance
Resting	No	Closed	Non-conducting
Activated	Yes (2 ACh on α-subunits)	Open	Conducting (cations)
Desensitized	Yes (usually occupied)	Closed	Non-conducting

Desensitization

The acetylcholine signal is transient: the receptor possesses an intrinsic mechanism that closes the channel even in the continued presence of ACh. In the desensitized state, the ACh binding sites are typically occupied but the pore is closed and non-conducting.

https://doi.org/10.1016/j.neuropharm.2020.108256

What is desensitization of the nAChR?

Desensitization is a state in which the receptor has ACh bound but the channel is closed and non-conducting. It acts as an intrinsic timer that limits the duration of the signal even in the continued presence of the agonist.

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In the {1:desensitized} state of the nAChR, the ACh binding sites are {2:occupied} but the pore is {3:closed} and non-conducting.

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Signal Termination — Acetylcholinesterase

Enzymatic Hydrolysis of ACh

Acetylcholinesterase (AChE) catalyzes the hydrolysis of ACh into choline and acetate, allowing the cholinergic neuron to return to its resting state after activation.

$\text{Acetylcholine}\stackrel{\text{AChE}+H_{2}O}{\to }\text{Acetate}+\text{Choline}$

• Choline is recycled back into the nerve terminal via a choline carrier
• This ensures rapid termination of the cholinergic signal

Reference

Dvir H, Silman I, Harel M, Rosenberry TL, Sussman JL. (2010) Chem Biol Interact. 187:10–22.

What enzyme terminates the ACh signal at the synapse, and what are its products?

Acetylcholinesterase (AChE) hydrolyzes ACh into choline and acetate. Choline is recycled back into the presynaptic neuron via a carrier.

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Acetylcholinesterase hydrolyzes acetylcholine into {1:choline} and {2:acetate}, allowing the cholinergic neuron to return to its {3:resting} state.

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${\text{GABA}}_A$ Receptor

Overview

The ${\text{GABA}}_A$ receptor is an ionotropic receptor gated by GABA ($\gamma$-aminobutyric acid). It is a ligand-gated chloride channel; ${\text{Cl}}^{-}$ influx hyperpolarizes the membrane (inhibitory effect), in contrast to the depolarizing cation channels of nAChR.

Key Concept: Depolarization vs. Hyperpolarization

Important

Ligands that stimulate ion channels can either depolarize or hyperpolarize the target cell membrane, depending on the type of ion that passes through the pore:

• ${\text{Na}}^{+}$ / ${\text{Ca}}^{2+}$ influx → Depolarization (excitatory)
• ${\text{Cl}}^{-}$ influx → Hyperpolarization (inhibitory)

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The GABA_A receptor, upon activation, allows {1:Cl⁻} to flow into the cell, causing {2:hyperpolarization} of the membrane, which is an {3:inhibitory} effect.

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Pharmacological Modulation of ${\text{GABA}}_A$

Allosteric Modulation Sites

Several pharmacological substances can modulate GABA’s effect by binding to sites distinct from the GABA binding site on the ${\text{GABA}}_A$ receptor.

Substance Class	Effect on GABA Activity	Functional Outcome
Benzodiazepines	Increase	Anxiolytic
Barbiturates	Increase	Sedative/anesthetic
Ethanol	Increase	CNS depression
Pregnenolone sulfate / Allopregnanolone / Allotetrahydrodeoxycorticosterone (neuroactive steroids)	Increase	Sedative
$\beta$-carbolines	Decrease	Anxiogenic
Picrotoxin / TBPS	Decrease (channel block)	Convulsant
Bicuculline	Decrease (competitive antagonist)	Convulsant
Flumazenil	Blocks benzodiazepine site	Reversal agent
Ciclopirroloni / Imidazopiridine	Increase (BZD site agonists)	Anxiolytic/hypnotic

Receptor

Regulation ${\text{GABA}}_A$ receptor activity is also regulated by phosphorylation by PKA and PKC.

Name two classes of substances that increase GABA_A activity and two that decrease it.

Increase: Benzodiazepines, barbiturates (also ethanol, neuroactive steroids).
Decrease: β-carbolines, picrotoxin/bicuculline.

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Flumazenil acts at the {1:benzodiazepine} binding site of GABA_A and is used to {2:reverse} the effects of benzodiazepines. β-carbolines are {3:inverse agonists} that reduce GABA_A activity and are {4:anxiogenic}.

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Comparison of Major Receptor Types

Receptor Type	Mechanism	Second Messenger	Example
Ionotropic (ion channel)	Direct ion flow on ligand binding	Ions (${\text{Na}}^{+}$, ${\text{Ca}}^{2+}$, ${\text{Cl}}^{-}$)	nAChR, ${\text{GABA}}_A$
Metabotropic (GPCR)	G protein → enzyme → second messenger	cAMP, ${\text{IP}}_3$, DAG	mAChR, $\beta$-adrenergic
Catalytic (receptor enzyme)	Ligand binding activates intracellular enzyme domain	Phosphorylation cascades	Receptor tyrosine kinases
Receptor with no intrinsic enzyme	Interacts with cytosolic kinase → gene regulation	Kinase cascades	Cytokine receptors
Nuclear/intracellular	Steroid binds nuclear receptor → regulates gene expression	mRNA / protein	Steroid receptors

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Nuclear / Intracellular Receptors

Mechanism

Steroid hormones (lipophilic) diffuse through the plasma membrane and bind to nuclear receptor proteins. The ligand-receptor complex then acts as a transcription factor, regulating the expression of specific target genes → production of specific mRNA and proteins.

How do nuclear receptors differ from membrane receptors in their mechanism of action?

Nuclear receptors bind lipophilic ligands (e.g., steroids) intracellularly. The ligand-receptor complex directly regulates gene expression by acting as a transcription factor, in contrast to membrane receptors which transduce signals via second messengers or kinase cascades.

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Mnemonic — Ionotropic Receptor Ligands:

“A GABA eats Gly and 5-HT”

• Acetylcholine (nicotinic)
• GABA (${\text{GABA}}_A$)
• Glutamate
• Glycine
• 5-HT (Serotonin)

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TLDR

Lecture A.02 — Key Points

• Receptors are classified as membrane (ionotropic, metabotropic, catalytic) or intracellular (nuclear)
• Ionotropic receptors are ligand-gated ion channels — the simplest signal transducers; they directly convert ligand binding into ion flux and membrane potential changes
• nAChR is a pentameric (${\alpha }_2\beta \gamma \delta$) cationic channel; ACh binding causes allosteric rotation of M2 helices (moving Leu257 away from pore center) → channel opens; found at NMJ, ganglia, adrenal medulla, CNS
• nAChR states: resting (unoccupied, closed) → activated (ACh bound, open) → desensitized (ACh bound, closed) — desensitization terminates the signal intrinsically
• Acetylcholinesterase hydrolyzes ACh → choline + acetate, enabling signal termination; choline is recycled
• ${\text{GABA}}_A$ is an inhibitory ionotropic receptor: ${\text{Cl}}^{-}$ influx → hyperpolarization; can be allosterically modulated by benzodiazepines (↑), barbiturates (↑), ethanol (↑), neuroactive steroids (↑), $\beta$-carbolines (↓), picrotoxin/bicuculline (↓); regulated by PKA/PKC phosphorylation
• Ion type determines effect: cation influx (${\text{Na}}^{+}$/${\text{Ca}}^{2+}$) → depolarization (excitatory); ${\text{Cl}}^{-}$ influx → hyperpolarization (inhibitory)
• Nuclear receptors bind lipophilic ligands (steroids), translocate to nucleus, and directly regulate gene transcription