BIO448 - Ion Channels

myelination

__ increase conduction velocity in axons, aka the current passively flow farther and farther due to the subsequent saltatory conduction.

leakage

myelination prevents ___ of current/positive charge through the leaky K+ channels as it passively flows along axon.

action potential

When losing myelin sheath, we have a loss of the ___ because too little current/positive charge makes it to the next region of concentrated Na+ channels, thus none can be made/propagated.

redistribution

There is some signaling between the myelin sheath and the axon, causing a ___ of the Na+ channels, leading to them being concentrated in the nodes of Ranvier regions.

Leaky K+ channels are still found everywhere along the axon, the myelin sheath just makes them leak less.

function

In demyelinating diseases, sometimes, ___ can be recovered over time.

Voltage-dependent Na+ channels slowly redevelop in the unmyelinated region, so function is recovered as the impulse alternate between fast conduction in the myelinated region, and slow conduction in the demyelinated one.

voltage clamp

the current injected by the amplifier is equal but opposite in sign to the current that is flowing through the cell

Patch clamp method

allows recording of current from single channels & can detect conformational changes in a single protein

Uses recording pipette and mild suction to isolate a single channel.

Made up by Erwin Neher and Bert Sakmann

whole cell recording

Patch clamp method

recording pipette and strong suction to break the isolated part of plasma membrane

• cytoplasm is continuous with pipette interior

• allows diffusional exchanges with interior of the cell

  • can inject substances to modulate interior of cell

• measurement of Vm and current from the entire cell

inside out recording

patch clamp method

free of the complications imposed by rest of the cell

strong suction and pull the recording pipette to take off a segment of the plasma membrane with channel on it

• yield a small patch with the former intracellular surface exposed

• can change the medium to which the intracellular surface is exposed

• valuable when studying the influence of intracellular molecules on ion channel function

study characteristics of a SINGLE channel

outside out

patch clamp method

strong suction, pull recording pipette out to rip a segment of plasma membrane off, let the new ends fuse together

• membrane patch has its extracellular membrane exposed

• cytoplasmic domain pointed to the interior

• optimal for studying how channel activity is influenced by extracellular signals (neurotransmitters)

• single channel recording

fluctuations

patch clamp method allows us to see the ___ between open and closed states of a single channel.

outward

__ current, peaks point up on voltage clamp, ions flow out of cell, depolarization

inward

___ current, peaks point down on voltage clamp, hyperpolarization, positive ions flow in the cell

gating

what is responsible for the opening and closing of the ion channels

• voltage

• ligand (neurotransmitters)

• temperature

• light (photons)

• stretch

activation curve

graph showing the probability of open state of the ion channel in the presence of a stimulus

Does NOT tell us when it is open, just gives us a probability.

macroscopic

voltage dependence of ion channels has ___ effects

half

K+ and Na+ activation curves are very similar: as the membrane potential increases (depolarization) the probability of ion channels being open increases.

At 0mV the channels are open __ of the time

volatge sensor

transmembrane domain of channel detecting and reacting to the change in voltage/membrane potential

macroscopic

single channel currents build/add up to the whole cell current, aka the ___ current

Many single channel currents summate to create the whole cell (___) current

Individual channels behave slightly differently from each other, they do not behave perfectly, thus we CANNOT know WHEN they will be open. They aren’t all open at the same time. Adding up all the individual behaviors of the whole channel population gives you a __ current

Po, probability of open channel

___ is dynamic, regulation is due to gating and modulation

unevenly

ion channels are ___ distributed throughout the cell

axon hillock

region of neuron with high concentration of Na+ channels, thus the highest probability of firing an action potential, as the threshold id very low. On the opposite hand, we have the dendrites that have a low Na+ channels concentration, thus a higher threshold, and very little chance to fire an action potential.

properties of ion channels

• membrane spanning

• allow rapid ion movement

• selective

• gated

• modulated

membrane spanning

several transmembrane domains

integral protein going from one side of the plasma membrane to the other.

allow rapid ion movement

diffusion of ions is passive (does NOT require energy), and happens DOWN electrochemical gradients

Selective

selective to either anions or cations or to specific ions (K+, Na+, etc.).

Can be permeable to multiple ions too.

Gated

Open and closed states activated by voltage, ligand binding, light, or physical changes

modulated

modulated/changed by auxiliary subunits, usually B subunits, G proteins, or neuromodulators

Most selective

Voltage gated ion channels, Ca2+ activated potassium channels, cyclic nucleotide activated ion channels are the ____

ONE

All voltage-gated ion channels are selective for ___ ion, and only ___

less selective

permeable to more than one ion

Ligand gated ion channels (nAChR, GluR, GABA A etc.).

cations/anions permeable

least selective

gap junctions

let all ions through, even small metabolites

largest pores

voltage-gated channels

selectively permeable to each of the major physiological ions

all contribute to the resting potential
all control excitability
responsive to changes in membrane potential

ligand-gated channels

essential for synaptic transmission

usually less selective in ion permeability than voltage-gated channels

sensitive to intracellular signals (Ca2+, cGMP, cAMP, phosphorylation etc.).

large

ion channels are ___ transmembrane proteins made from 640 to 2000 amino acid residues

multimeric

made of multiple parts/subunits

homomeric multimer

made of multiple parts/subunits made of the same protein

heteromeric multimer

made of multiple parts/subunits that are different proteins

heteromeric

ligand-gated channels such as the ones responding to acetylcholine are ___ multimers

homomeric

voltage gated K+ channels are ___ multimers

single polypeptide

Voltage-gated Na+ channels are ___ aka a single protein spanning the membrane multiple times and has multiple transmembrane domains

protein

amino acid chain encoded by mRNA

subunit

part of a channel. Single protein may contain 1 or more domains. Subunit = 1 protein that needs to interact with another to be functional.

domain

functional “unit” of a channel

transmembrane segment

section of a domain that spans the membrane

N & C tails

allow subunit to associate with other proteins or ligands to modulate channel function

20

__ amino acids (alpha helices or B sheets) are necessary to traverse the membrane

8

a MINIMUM of __ transmembrane crossings are necessary to form a functional channel

X-rays analysis

gets us the 3D structure of proteins

proteins need to be crystalized

help finding binding sites, location of pores, identifying amino acids etc.

24

voltage gated ion channels have __ transmembrane regions

repeated motifs of 6 transmembrane spanning regions (times 4)

this type of channels has a distinct type of transmembrane helix that possesses a number of positively charged amino acids serving as voltage sensors

4

Voltage-gated K+ channels have __ subunits of the same type (homomeric), 6 transmembrane regions.

pore loop

formed by the chain of amino acids connecting the 2 helices of one subunit, all of them point toward the inside of the channel creating the pore, which stabilizes ion as it loses its hydration shell to go through the channel.

negatively

the amino acids in the pore loops are ___ charged to stabilize the ion going in (in the potassium voltage-gated channels)

non-hydrated

ion channels only allow the passage of ___ ions, hence the amino acids of the pore loops stabilizing ions as they go through

spacing

the amino acids chains forming the pore loops are specific enough and create a specific pore aka unique __ which is the reason behind selectivity of ion channels

3

Voltage-gated potassium channel is occupied by __ K+ ions. Passing through does not require energy aka passive.

phospholipid bilayer

The transmembrane part of the K+ voltage-gated channels is sitting in the ___

hinge

In between the transmembrane domain and the T1 domain are the ___ regions, which are the parts that are going to be moved by the voltage sensor domain, causing it to go up or down depending on membrane potential.

Beta

___ subunits bind to ligands to modulate protein aka K+ voltage-gated channel

up

depolarization, positively charged amino acids of voltage sensor move __ making the hinge regions go __ as well, opening the K+ voltage-gated channel

Depolarization : pushes sensors outward (because more + inside)

down

hyperpolarization, positively charged amino acids of voltage sensor move __ making the hinge regions go __ as well, closing the K+ voltage-gated channel

Hyperpolarization : pulls sensors inward (because more - inside)

positive charges

___ enable movement within the membrane in response changes in membrane potential

S4

special helix in voltage-gated channels acting as sensor

conservation in sequence for it between many different species

is absent from NON voltage dependent K+ channels

relative to equilibrium potential only comes down at equilibrium, but even small depolarization is enough to move them up

ball and chain model

model for the inactivation mechanism of voltage-gated Na+ channels

modulation

__ shifts the probability curve of ion channel being open to right or to the left

when a channel is modulated, the same stimulus will give a different response

many different mechanisms and time scales of modulation

-phosphorylation and second messenger (rapid and reversible)

-change in subunit composition or receptor trafficking (slower, longer lasting)

-change in gene expression (slower, long lasting)

faster

Na+ channels inactivation is ___ in the presence of beta subunits, aka ___ repolarization

binding of beta subunits changes the kinetics of the ion channels
Example: scorpion toxin affects beta subunits of Na+ channels and make inactivation time longer, leading to paralysis

channelopathies

genetic diseases affecting ion channels\

arise from mutations in critical amino acids in the ion channels

Ca2+

volatge-gated __ ion channels implicated in

-Familial hemiplegic migraines (FMH) due to mutation in pore forming region

-Episodic ataxia (EA2) truncating (nonsense) mutations leading to improper assembly of channel at membrane

-Congenital night blindness truncating (nonsense) mutations affect retinal rod photoreceptors

ALL of these lead to decreased __ currents and thus decreased neurotransmitters release

Na+

diseases implicating the __ voltage-gated channels

-generalized epilepsy with febrile seizures (GEFS)

-mutation in alpha subunit = chromosome 2
-mutation in beta subunit = chromosome 19

ALL of them lead to slowing of __ channels inactivation, which causes hyperexcitability

active transporters

maintain ion concentration gradients across the membrane

restore ions displaced during current flow from electrical signaling or continual leakage that occurs at rest

against, energy

active transporters move ions ___ their electrochemical gradient. which requires ___

ATP

active transporters use energy in the form of ___ or other high energy phosphate molecules or using another ion’s concentration gadient

2 classes of transporters

• ATPase pumps

• ion exchangers

ATPase pumps

acquire energy from hydrolysis of ATP

ex: Na+/K+ pump & Ca2+ pump

• change conformation after binding high energy phosphate molecule in order to move ion across membrane

ion exchangers

use electrochemical gradient of other ions as energy source

ex: usually Na+ like Na+/Ca2+ or Na+/H+ transporters

• coupling movement of one ion to move another against its gradient

• exchange intracellular for extracellular ion

OPPOSITE

ion exchangers move ions in ___ directions across the membrane

co transporters

move ions in the SAME direction across the membrane

type of active transporters

• carry multiple ions in the same direction

• usually regulate Cl- ion concentration, can also pick up neurotransmitters

Ex: K+/Cl- transporter, Na+/GABA or dopamine transporter, Na+/K+/Cl- transporter

Na+/K+

co-transporters and ion exchangers ultimately depend on the ___ ATPase pump.

Na+/k+ pump

ATPase pump

horseshoe shaped

pumps Na+ from inside to outside the cell & K+ from outside to inside the cell

1. 3 Na+ ions bind to the pump from the inside

2. protein phosphorylated by ATP

3. change in conformation with the binding site now facing outside the cell

4. the sodium gets out and 2 K+ ions bind to the pump

5. dephosphorylation means conformation change and protein back to facing the inside of the cell

6. potassium gets out

Every time it’s working, the inside gets more negative (hyperpolarize) because it gets 3 positive charges out but only brings back 2.

electrogenic

create an electrochemical gradient

ex: Na+/K+ pump

inseparable

the opposing flow of Na+ and K+ in the Na+/K+ pump are operationally ___, meaning they do not work without the other present.

reducing the concentration of one reduce the movement of the other.