3-4. Ion Channel Diversity and Control

Lecture 3 + 4

What are the 3 important properties of ion channels?

1. Conduct ions across membranes

2. Recognize & select specific ions

3. Open/close in response to mechanical, electrical, or chemical signals

What are the phases of an action potential and which channels are involved?

1. Resting (–60 mV): all Na⁺ & K⁺ gated channels closed

2. Depolarization: Na⁺ voltage gated channels open and enters cell→ membrane becomes +40 mV

3. Repolarization: Na⁺ channels inactivate, K⁺ channels open slowly → back to negative potential

4. Hyperpolarization (undershoot): K⁺ channels stay open → membrane more negative than resting

What makes ion channels so diverse?

• Open by different stimuli (voltage/electrical, ligand, stretch, temp, pH)

• Different kinetics: open/close quickly, stay open longer, one-way movement, or always open

What is the structure and function of ion channels?

• Function: aqueous pore for ions to cross via diffusion

• Structure: large transmembrane proteins with hydrophobic membrane-spanning domains

• Can be hetero-oligomers, homo-oligomers, or single polypeptides with repeating motifs

• Some have auxiliary subunits that assist gating

What structural features do ion channels share?

• Alpha helices span lipid bilayer

• Hydrophobic amino acids face lipids

• Hydrophilic amino acids line pore

• Q: Which is monomeric? → C) Single polypeptide w/ repeating motifs

  • Which is not? → A) and B) are individual subunits

Example: How is a voltage-gated Na⁺ channel structured?

• Monomeric protein with 4 homologous transmembrane domains (motifs)

• Helix 4 in each domain = voltage sensor

  • Has more positively charged AA to provide the voltage sensor to open and close the channel

• Non-helical P-loops between helices 5 & 6 in each domain = selectivity filter

  • Makes channel specific to certain ion

• Loop between domains 3 & 4 = inactivation gate

How do conformational changes open ion channels?

Gating involves twisting & bending of alpha helices → enlarges pore, creates hydrophilic surface for ions

A) Conformational change in one region

B) General structural change

C) Blocking particle = inactivation gate

Why do ions need channels to cross membranes?

• Ions interact with water’s dipoles (O = slightly negative, H = slightly positive)

  • Oxygen attracts electrons; Hydrogen loses electrons

• Forms hydration shell, making lipid bilayer crossing energetically unfavorable

How do ion channels achieve selective passage of ions?

• Hydration cloud prevents ions crossing the hydrophobic lipid bilayer

• Channels provide aqueous pore with selectivity filter

• Filter binds ions briefly (<1 μs), plus pore diameter & electrostatic forces → ensures selectivity

Why does a potassium channel not also allow sodium ions to pass through, even though sodium is smaller?

• Sodium has a smaller atomic radius → charge more concentrated → forms a larger, stronger hydration cloud

• Potassium’s charge is more spread out → weaker interactions with water → smaller hydration cloud

• Functionally, sodium acts “larger” than potassium due to its hydration shell

• Selectivity filter amino acids favor ions that can shed their hydration cloud → K⁺ passes, Na⁺ blocked

What was revealed by the first crystal structure of an ion channel (K⁺ channel, 1998)?

• Tetrameric structure (4 subunits around central pore)

• P-loops from helices form narrow selectivity filter

*Don’t pay too much attention to this slide

What amino acid sequence enables K⁺ channel selectivity?

• Signature sequence = TVGYG in selectivity filter

• Hydroxyl of the threonine & carbonyl groups of other AA form 4 K⁺ binding sites

• Less conserved sequence → reduced K⁺ selectivity (e.g., nonselective HCN channels allow K⁺ & Na⁺)

How do ion channel selectivity filters vary across channel types?

• Many channels share similar amino acid sequences in selectivity filter region

• Greater conservation → stronger ion selectivity

• Less conservation → broader ion permeability

Why doesn’t sodium (Na⁺) pass through potassium (K⁺) channels even though it’s smaller?

Because Na⁺ has a larger hydration cloud (stronger interaction with water due to higher charge density), making it functionally larger than K⁺.

What are the four main types of ion channels and what activates them?

• Resting K⁺ channel/leak channel → always open

• Voltage-gated → open with change in membrane potential

• Ligand-gated → open/close with neurotransmitter binding

• Signal-gated → open/close with intracellular molecules

• Temperature-gated → open/close based on temperature

How do ion channels open and how is current measured?

• Channels open/close in all-or-none fashion

• Opening allows brief ion passage → measurable current

• Current size: a few picoamperes (pA)

  • Channel closed: current is 0

  • Channel open: 5 pA

What is a gigaseal and why is it important in patch clamp recordings?

• Gigaseal = very tight seal (≥10 GΩ resistance) between glass pipette and membrane

• Ensures all ion flow enters pipette → higher seal resistance = lower background noise, improves accuracy

How can patch clamp be used to study ion channels in muscle?

• Pipette filled with acetylcholine solution

• Records current through a single channel

• Can study leak, voltage-gated, ligand-gated, mechanically-gated channels across many cell types

• Can measure voltage changes inside and outside the cell

An ion channel has 4 pA current when open. About how many ions pass through per second?

Things to consider:

• 1 ampere = 1 Coulomb/sec (Coulomb = measure of electric charge)

• The charge of a monovalent cation = 1.6 × 10^-19 Coulomb

Set up calculation:

1pA = 1 × 10^-12 A

Rewrite to: 1pA = 1 × 10^-12 Coulomb/sec

So: 4pA = 4 × 10^-12 Coulomb/sec

Also:

If 1 ion = 1.6 × 10^-19 Coulombs

Then:

4 × 10^-12 Coulomb/sec x 1 ion/1.6 × 10^-19 Coulombs

= 2.5 × 10⁷ ions/sec (25 million ions/sec)

What does inside-out patch clamp allow researchers to study?

• Electrode quickly withdrawn from cell

• Exposes cytoplasmic (inside) surface to external solution

• Useful for testing intracellular factors that regulate channel activity

What does outside-out patch clamp allow researchers to study?

• Electrode slowly withdrawn from cell

• Exposes extracellular surface of membrane

• Useful for studying ligand-gated channels (ligands applied to bath solution)

How do voltage-gated Na⁺ and K⁺ channels behave during depolarization?

• Rest and hyper-polarized: both voltage gated channels closed

• Depolarization activates voltage sensors (+) → Na⁺ channels open first, then K⁺ channels

• Na⁺ channels inactivate during depolarization, K⁺ often stay open

How do pumps/transporters differ from ion channels?

• Require energy: ATP (ATPases) or ion gradients (co-transported ions)

• Transporters move ions slower than channels

• Energy needed to move ions against their electrochemical gradients

How do toxins affect ion channels?

• Bind specific sites on voltage-gated sodium channels

• Can block/inhibit function → also useful for studying channel mechanisms

Channelopathies - What happens in hyperkalemic periodic paralysis?

• Mutation in voltage-gated Na⁺ channels prevents inactivation gate from closing

• Triggered by high blood K⁺

• Leads to periodic muscle paralysis (muscle cells become inexcitable)