Lecture 16: Ion Channels

ZAP

Conducting ions is a _____ and ______ process

Passive and rapid

Ions are surrounded by…

Waters of hydration

Pore size compensates for what?

Waters, they are sifted/filtered

All ion channels have what?

An aqueous pore that spans the entire width of the membrane

Trimeric Channels (P2X)

Feature a cysteine-rich loop along with two transmembrane domains

The cysteine-rich loop has what? What modifies it?

A ligand binding site (asparagine), it is modified by lysine (k)

How many transmembrane domains does a trimeric channel have?

Two transmembrane domains.

What goes in a trimeric p2x channel? What goes out?

Cations of calcium and sodium go in, potassium goes out.

How does a trimeric channel get the name p2x?

P2 (two transmembrane domains) X (Cystine-rich loop)

What regulates a P2X channel?

Glycine

The N-terminous for the P2X, what is it’s function?

Crucial for phosphorylation of threonine, desensitizing or closing the channel

How many different subunits combine to form receptors of P2X?

7, with 11 studied

Tetrameric Channels are _____ gated __ Channels

Voltage, Na

What percentage identical are transmembrane domains?

>50%

What forms the voltage sensitive region in a tetrameric channel?

S1-4

What is the role of S4 in a tetrameric channel?

A voltage sensor

Where does the tetrameric channel get its name?

4 domains, each one made up of 6 transmembrane segments

What causes the desensitization and closing of D3/D4 in a tetrameric channel?

The IFM motif interacts with it, aided by the C-terminus

The n-terminous is intra or extracellular?

Intra

What does the N-terminous itneract with?

Microtubule proteins

Each alpha subunit gets…

2 beta subunits

What are the roles of the beta subunit?

• Cell adhesion

• Axon fasciculation

• Neurite outgrowth promotion

• Channel kinetics promotion

How many isoforms are there of voltage gated Na channels?

There are at least 9 known isoforms of voltage-gated Na+ channels, each encoded by different genes and exhibiting varied properties.

AMPARS are the primary receptor of what?

Glutamate

Most AMPARs are

GluA1/2 and GluA2/3

AMPAR receptors are heteromeric, which means

They are made up of 2 or more subunits

In tetrameric channels, what forms the binding sites?

S1 and S2

S2 has a “R/G” site, what is another name for it?

Flip/flop

What does a flop variant do faster?

Desnsitize

What does a Hairpin loop create in an AMPAR variant?

The pore

What is the symbol for current?

I

What is the symbol for voltage?

V

Negative I means what?

Ions flowing in

What study characterized cocaine-induced changes in AMPARs in the Nucleus Accumbens (NAc)?

Conrad et al., Nature (2008)

According to Conrad et al. (2008), what specific changes were observed in NAc AMPARs after cocaine administration?

Changes in electrical conductance at both negative (-) and positive (+) voltages.

Changes in the shape of the Current-Voltage (IV) curve.

Evidence pointing towards the presence of Calcium-Permeable AMPARs (CP-AMPARs).

How was the presence of CP-AMPARs verified in the Conrad et al. (2008) cocaine study?

1. Pharmacologically: Inhibition by NASPM (a specific blocker of CP-AMPARs).

2. Biochemically: Direct biochemical analysis confirming changes in subunit composition consistent with CP-AMPARs

What are the functional consequences of cocaine-induced CP-AMPARs in the NAc?

• Produce greater current influx (pass both Na+ and Ca2+).

• Increased Ca2+ influx strengthens depolarization.

• Increased depolarization increases the likelihood of opening nearby NMDARs.

• Rectification properties potentially keep the channels open longer under certain conditions.

What is the role of the Nucleus Accumbens (NAc) in addiction, particularly regarding environmental cues and CP-AMPARs?

• The NAc is activated by environmental cues associated with drug use.

• This activation is increased in addiction.

• Further studies (e.g., Wolf) showed that drug craving effects can be mediated by these cocaine-induced CP-AMPARs in the NAc.

What type of channel are Nicotinic Acetylcholine Receptors (nAChRs), and what subunits make them up?

• Pentameric Channels (made of 5 subunits).

• Subunits include α, β, γ, δ, and ε.

• The α subunit is obligatory (must be present).

What are the most common subtypes of nAChRs found in the brain and muscle?

• Brain: α4β2 and α7 subtypes are most common.

• Muscle: (α1)2β1γδ subtype is most common.

Describe the key structural features of nAChRs related to ligand binding and pore formation.

• Ligand Binding Site: Located on the N-terminus, characterized by a Cys loop, supported by the C-terminus.

• Transmembrane Domains: Four domains (M1-M4) per subunit.

• Pore: Formed by the M2 transmembrane domain from each of the 5 subunits. Subunits rotate to open the pore.

What ions pass through nAChRs, and what is the function of the intracellular loop?

• Ion Permeability: Allow Na+ and Ca2+ to enter the cell, and K+ to exit.

• Intracellular Loop (M3-M4): Involved in receptor assembly, trafficking (targeting to specific neuron compartments like dendrites vs. axons), and regulating desensitization/closing (often driven by phosphorylation).

Define EC50 as used in dose-response curves.

EC50 (Half maximal effective concentration) is the concentration of a ligand (e.g., drug or neurotransmitter) that produces 50% of the maximum possible effect or response. It's used as a measure of ligand potency.

What other neurotransmitter receptors belong to the same structural family (Cys-loop pentameric channels) as nAChRs?

• GABAA receptors

• 5-HT3 receptors (Serotonin)

• Glycine receptors

What are the three key properties shared by all ion channels?

Signal sensitivity, ion selectivity, and ion conductance.

Why are ion channels considered passive conductors of ions?

Because they allow ions to move down their electrochemical gradient without using ATP.

How does the molecular filter of an ion channel influence its selectivity?

It excludes ions that don't match the correct size, charge, or ability to shed water.

Why can larger ions sometimes pass through narrower channels compared to smaller ions?

Larger ions may more easily shed their hydration shell, allowing them to fit through the filter.

Explain how ion channel conductance influences neuronal signaling speed and reliability.

Higher conductance allows for rapid membrane potential changes, essential for fast signaling.

What type of ions do P2X receptors primarily allow into the cell?

Sodium (Na⁺) and calcium (Ca²⁺).

What does the G342 residue in the TM2 domain regulate in P2X receptors?

Ion selectivity and conductance.

How did MTSET help researchers study P2X channel function?

MTSET reacted with engineered cysteines in TM2, revealing which regions were accessible in the open state.

How does phosphorylation at the P2X N-terminal threonine site influence receptor behavior?

It causes receptor desensitization or closure.

Why might different combinations of P2X subunits result in different physiological effects?

Subunit combinations alter ligand affinity, kinetics, and ion selectivity.

What domain of voltage-gated sodium channels contains the voltage sensor?

The S4 segment.

What feature of GluA2-lacking AMPA receptors makes them calcium permeable?

They retain glutamine at the Q/R site, allowing calcium influx.

How does the IFM motif contribute to sodium channel desensitization?

It rapidly blocks the channel pore after opening, inactivating the channel.

Compare how AMPA receptor subunit composition influences ion flow and synaptic strength.

Subunits affect calcium permeability, desensitization rate, and synaptic plasticity.

Explain how changes in AMPA receptor composition might contribute to pathological states like addiction.

Upregulation of calcium-permeable AMPA receptors increases excitability in reward circuits.

What subunit is required in all nicotinic acetylcholine receptors?

The alpha (α) subunit.

What happens to the pore when nicotinic receptor subunits rotate?

It opens to allow Na⁺ and Ca²⁺ in, and K⁺ out.

How does nicotine affect the EC50 of α4β2 nicotinic receptors?

It lowers the EC50, making the receptor more sensitive.

Why does nicotine have a stronger effect at α4β2 receptors than at α7 receptors?

α4β2 has a higher affinity and is more responsive at physiological levels.

Based on receptor subtype expression, why might nicotine affect reward but not memory at normal doses?

Reward areas express α4β2 receptors; memory-related areas use α7, which are less sensitive to typical nicotine exposure.

What protein family helps AMPA receptors reach the membrane?

TARPs (Transmembrane AMPA receptor regulatory proteins).

What does the "flop" splice variant do to AMPA receptor desensitization?

It increases the rate of desensitization.

How does phosphorylation influence nicotinic receptor desensitization?

It can enhance or reduce desensitization depending on the site and kinase.

How do intracellular loops influence ion channel localization?

They interact with trafficking proteins to direct channels to specific regions (axon, dendrite).

How might impaired receptor trafficking lead to neurological dysfunction?

Mislocalized receptors could disrupt signal balance, impairing cognition, plasticity, or movement.

What does an I-V curve measure?

The relationship between membrane voltage and ionic current.

What is rectification in an ion channel?

A tendency to conduct ions better in one direction than the other.

How was NASPM used to identify CP-AMPARs in cocaine-exposed rats?

NASPM blocked current, confirming the presence of calcium-permeable AMPA receptors.

Why do CP-AMPARs show greater inward current at negative voltages?

Their rectifying properties favor ion entry at hyperpolarized potentials.

How could upregulation of CP-AMPARs contribute to drug craving during withdrawal?

Increased Ca²⁺ and Na⁺ influx strengthens reward circuitry responses to drug cues.