MEMBRANE POTENTIAL

ACTION POTENTIAL
 Excitable Cells
Excitable cells are those that can be stimulated to
create an electric current.
Muscle cells
Nerve cells (neurons)

All cells (not just excitable cells) have a resting

potential (membrane potential): an electrical
charge across the plasma membrane
The interior of the cell negative
Typically -70mV (varies -40 to -90 mV depends on the
type of neuron).
 Resting membrane potential
Unequal distribution of ions
across the plasma membrane
[Na+] outside 10x > inside
[K+] inside 20x > outside
[Cl-] and [Ca2+] outside > inside
[Ca2+] high in some intracellular
compartments (green oval)
Resting membrane potential
Na+/K+
pump

1. The Na+/K+ transporter

(ATPase/pump) - active transport
Two K+ into the cell
Three Na+ out of the cell
Result: a net loss of positive charges
within the cell
2. Some potassium
channels (non-gated)
Ca2+ pump in the plasma
membrane are "leaky"
allowing a slow
facilitated diffusion
of K+ out of the cell
(red arrow).

3. Ca2+ is removed via Ca2+ pump

Electrochemical gradients
in out in out in out

membrane permeable only for K+

No net flux of K+ Net flux of K+ Flux of K+ from 1 to 2

From 1 to 2 balanced by opposing
membrane potential
electrochemical equilibrium
 Electrochemical gradients contd
At this electrochemical equilibrium, there is an
exact balance between two opposing forces:
Chemical driving force = ratio of concentrations on 2
sides of membrane (concentration gradient)
The concentration gradient that causes K+ to move from 1 to 2,
taking along positive charge, and
Electrical
driving force = potential difference across
membrane
And opposing electrical gradient that increasingly tends to stop
K+ from moving across the membrane

Equilibrium: when chemical driving force is balanced

by electrical driving force
 Nernst Equation
Predicts the electrical potential generated across
the membrane at electrochemical equilibrium
(the equilibrium potential)
for a single permeant ion

RT [ X ]o 58 [ X ]o
Ex = ln log
zF [ X ]i z [ X ]i
EX = equilibrium potential for any ion X
R = gas constant
T = absolute temperature (Kelvin scale)
z = valence (electrical charge) of the ion
F = Faraday constant
[X] = concentration of ion X on each side of the membrane
What is the equilibrium potential for K+?

in out in out

58 [ K ]o 1
EK = log = 58 log = 58mV
z [ K ]i 10
What would happen if the K+ was
replaced with Na+ ?

in out in out

1mM Na+ 10mM Na+

No potential would be generated

Because no Na+ could flow across the membrane
What sets the membrane potential ?
1. Each ion gradient contributes its Nernst
equilibrium potential to the total
membrane potential.
2. Equilibrium potential across a Neuronal
Membrane:
Na+ contributes +58 mV
K+ contributes -75 mV
Cl- contributes -65 mV
Ca2+ contributes +155 mV
Etc. for Ca2+, Mg2+, PO4-2, etc.
What sets the membrane potential ?

3. The size of the contribution is weighted by

how easily the ion crosses the membrane.
We will use PERMEABILITY as our
measure of this.
If PK>>PNa, PCl, etc. VM EK
If PNa >> PK, PCl, etc. VM ENa
Etc.
 Membrane Potential
Goldman equation

Takes into account both

1) the concentration gradient of the permeant ions and
2) the relative permeability of the membrane ions

RT PNa [ Na ]o + PK [ K ]o + PCl [Cl ]i

EM = ln
F PNa [ Na ]i + PK [ K ]i + PCl [Cl ]o
P = permeability of the membrane to each ion
z = eliminated, therefore [Cl-]s have been inverted
-log (A/B) = log (B/A)
 In summary
The inside-negative resting potential arises
because:

1. The membrane of the resting neuron is more

permeable to K+ than to any other ion present
This selective permeability to K+ is caused by K+ -
permeable membrane channels (non-gated) that are
open in resting neurons
2. There is more K+ inside the neuron than
outside
The large K+ concentration gradient is produced by
membrane transporters that selectively accumulate
K+ within neurons
 AP is initiated at axon hillock (Initial Segment = IS)
Why?
1. Lower threshold for AP initiation
2. Small diameter - less current to drive membrane threshold
potential
3. Higher density of VG Na+ channels (20-200 fold) in IS than in
soma / dendrites (SD)
4. Properties of Na+ channels in SD differ from those in IS (slow
inactivation)
Properties of the Action Potential

Threshold All-or-none behavior

Depolarization = inside less negative
Hyperpolarization = inside more
negative
Refractory Period
 Membrane Potential
How to measure membrane potential ?

Threshold
Injections of small amounts of current result in small
shifts in membrane potential.
When the injection stops, the membrane potential
recovers more or less directly to the resting potential.
 pulses
applied
to a cell

the membrane
potential of the
stimulated cell

Electrical stimulation of non-excitable cells

Both hyperpolarizing and depolarizing electrical stimulation result in
graded potentials

The two traces are on the same time scale. Hyperpolarizing stimuli
lead to membrane hyperpolarization, and depolarizing stimuli lead to
membrane depolarization
 pulses
applied
to a neuron

the membrane
potential of the
stimulated neuron

Electrical stimulation of excitable cells such as neurons,

Hyperpolarizing stimulation results in hyperpolarizing graded potentials
proportional in magnitude to the amplitude of the pulse.
Sub-threshold depolarizing stimuli cause graded potentials
Supra-threshold depolarizing stimuli elicit all-or-nothing action potentials

The dashed line represents the threshold voltage (V threshold) of approximately

-50 mV.
 Mechanism of the AP
Resting potential
K+ channels are
always open

Make resting
membrane PK>PNa,
PCl, etc.

Resting membrane
potential near EK

Notice that at the equilibrium potential of a given ion,

there is no net flux of that ion across the membrane.
 Mechanism of the AP
Voltage-gated Na+
channels
respond to a small
depolarization by
opening.
this increases PNa.
Na+ enters the cell
down its
electrochemical
gradient.
EM goes toward ENa
Mechanism of the AP
The depolarization
causes more Na+
channels to open,
Increasing Na+ influx,
Increasing the
depolarization.

Threshold once it
gets started it goes
all the way.
Mechanism of the AP - gating

Depolarization

(1)
At resting potential the
channels are closed
(Ready).
Mechanism of the AP - gating

(2)
Upon depolarization the
channel first opens. Na+
can enter. EM ENa
Mechanism of the AP - gating

(3)
A couple of msec later the
channel shuts.
(Refractory/inactive,
not ready)
 Mechanism of the AP
The inactivation of the Na+ channels
causes the refractory period.
The channels must recover before they
can respond to a depolarization. This
takes time.
Na+ channels can be
closed and ready,
open,

closed and not ready (refractory).

 Mechanism of the AP

Voltage-gated K+ channels in many cells

open in response to depolarization.
Increase PK.

Open more slowly than the Na+ channels.

Increased K+ efflux helps restore the

resting potential.
 Mechanism of the AP
60
40
20
Depolarization
Repolarization The increase in
0
PNa drives the
mV

-20
-40 depolarization.
-60
-80 The fall in PNa
0 2 4 6 8 10
msec and PK drive the
PNa rises quickly then declines
repolarization.
PNa or PK

PK rises slowly, declines

with repolarization

0 2 4 6 8 10

g (conductance) = p (permeability)
 Mechanism of the AP
The Na+ channels
60 inactivate during the
40
Depolarization
Repolarization
action potential.
20
0 PNa declines.
mV

-20
-40
Inactive Na+ channels
-60
cannot respond to a
-80
0 2 4 6 8 10
stimulus.
msec
The cell is refractory
PNa rises quickly then declines
until the channels
PNa or PK

PK rises slowly, declines

with repolarization recover from
0 2 4 6 8 10
inactivation.
 The afterhyperpolarization (APH)
There is a hyperpolarizing
60
overshoot at the end of the AP
40 It takes a few miliseconds for all the
20
0
voltage-gated K+ channels to return
to the close state
mV

-20 hyperpolarization
During this time the efflux of K+ from
-40
-60
-80 the cell is greater than the resting
0 2 4 6 8 10
msec state
PNa rises quickly then declines
As a result, VM is hyperpolarized
PNa or PK

PK rises slowly, declines

with repolarization
slightly with respect to its normal
resting value
0 2 4 6 8 10
 Refractory Period
The short time immediately after an AP in which the
neuron cannot respond to another stimulus, owing
to an increase in potassium permeability.

This limits the number of APs that a given nerve cell

can produce per unit time

The membrane must recover from the first AP before

the second can be elicited.
At any time however, there either is an action
potential or not.
 Refractory Period
Two
Two phases
phases
The absolute refractory period
Comes immediately after the AP;
During this period it is impossible to excite the cell no matter how large a
stimulating current is applied
The relative refractory period
During which it is possible to trigger an AP, but only by applying stimuli that
are stronger than normal.
These periods are
caused by
1. the residual
inactivation of
Na+ channels and

2. opening of K+
channels
BREAK
 Propagation of the AP
active and passive current flow

(1)
Na channels locally open in
response to stimulus generating
and action potential
(Active, voltage-gated Na+ current)
(2)
The resulting inward current
flows passively along the axon

PROPAGATION
t=1
 Propagation of the AP
active and passive current flow
t=2
(3)
Local depolarization causes
neighboring Na+ channels to open
and generates an action potential

Membrane repolarized depolarized resting

(4)
Upstream Na+ channels inactivate,
While K+ channels open. Membrane
potential repolarizes. Axon is refractory here. PROPAGATION
 The rate of conduction depends on:
Passive and active flow of current

Size of the cell larger cells conduct faster (passive

flow)

Myelin - increased axonal insulation, reduced ability

of current to leak out of the axon (passive flow)

Sizeof the Na+ influx larger Na+ influx means faster

conduction. This depends on the number of Na+
channels that are opening (active flow).
 The action potentials
jump from node to
node.
Voltage-gated Na+
channels are present
only at the nodes of
Ranvier

1mm
1-2 m
Saltatory Conduction
Speed of AP in myelinated and
unmyelinated axon

Unmyelinated axon conduction : 0.5 to 10 m/s

Myelinated axon conduction up to 150 m/s
 Membrane conductance
Essential to understanding current flow through ion
channels
Conductance is an electrical term, the reciprocal of
resistance, and can be thought of as a measure of
permeability
Ions crossing the membrane also constitute an electrical
quantity, namely current, so it is natural that conductance
and the electrochemical gradient predict current

Iion = gion (VM Eion)

The electrochemical driving
force acting on the ion

Iion = ionic current

VM = membrane potential
Eion = equilibrium potential
Hodgkin and Huxley Voltage Clamp

The neuronal membrane potential (VM) is

controlled by the experimenter

The Voltage-Gated Channels open or close

in response to the predetermined Vc
(command voltage)

The current (fluxes of Na+ and K+) is

measured
 Membrane conductance
Hodgkin and Huxley used this equation to
calculate the dependence of Na+ and K+
conductance on time and membrane
potential
they knew VM set by voltage clamp
they could determine ENa and EK from the
ionic concentrations on the two sites of the
axonal membrane (i.e. Table 7-1)
they knew INa and IK from recordings of the
membrane currents

Iion = gion (VM Eion)

 Voltage Clamp
Internal electrode
measures membrane
potential (Vm) Voltage clamp amplifier
Sets the command potential Vc

The current flowing

Inject current
across the axon is
to make
measured here.
Vm=Vc

This feedback arrangement causes the membrane potential

to become the same as the command potential
Box 9-1
 If the membrane potential (Vm) and the command potential (Vc) are identical
Vm = -65mV and Vc is set at -65-mV
Membrane current is at the resting state
There is no change in voltage
No current is injected into the axon to oppose any changes in membrane current
 Vm = -65mV

Vc is set at 0mV

Vm = -65mV and Vc is set at 0mV

At Vc = -0mV clamp amplifier injects current to push the Vm to 0mV
A very brief capacitive current is seen because the step from one potential to
another alters the electrical potential difference across the membrane
Different types of ion channels open and close
An early inward current is seen followed by
A delayed outward current
 Vm = -65mV and Vc is set at 0mV
The transient early inward current corresponds to the Na conductance (INa)
carried out by Na+ ions (via voltage-gated Na channels) = causing
depolarization during an action potential. VGNaC activate rapidly to the
depolarization
The delayed outward currents corresponds to the K conductance (IK) carried
out by K+ ions (via voltage-gated K channels). VGKC activate more slowly
 Tetrodotoxin (TTX) Tetraethylammonium (TEA)
a neurotoxin found in puffer fish and block K+ currents without
in the skin of tropical frogs, and affecting the Na+ current
salamanders
blocks the Na+ current without
affecting the K+ current
 Voltage clamp at various Vc
Na+ and K+ conductances
change over time

As the size of the

depolarization increases,
the permeability and rate of
opening for Na and K
channels increase

At all levels of
depolarization
Na channels open more
rapidly
K channels open more
slowly

Fig 9-6
 Multiple Sclerosis

Voltage-gated Na+ Partial recovery

due to an increase in
channels the number of Na
a high density channels along the
clustered at the nodes demyelinated parts of
the axon,
of Ranvier
partially restoring axon
conduction
Voltage-gated Na+ channel expression
At least 9 genes : NaV1.1 - NaV1.9

NaV1.1, NaV1.2, NaV1.3, and NaV1.6 channels are

expressed widely within the NS

NaV1.7 - NaV1.9 channels are expressed

selectively within dorsal root ganglion and
trigeminal ganglion neurons

NaV1.4 and NaV1.5 channels are expressed within

somatic and cardiac muscle, respectively
Waxman et al. 2004
 Voltage-gated Na+ channels
At least 9 genes : NaV1.1 - NaV1.9

NaV1.2 channels
distributed diffusely along non-myelinated axons
support AP conduction that is known to occur in pre-
myelinated axons
after myelination, there is a loss of NaV1.2 channels

NaV1.6 channels
distributed along myelinated axons
cluster at the Nodes of Ranvier
not detectable under myelin
preferentially associated with axonal injury
Larger current that NaV1.2 channels
Waxman et al. 2004
 Multiple Sclerosis
NaV1.6

NaV1.6
Very low Na+ channels density
cannot support secure AP
conduction
NaV1.6 & NaV1.2
Some demyelinated axons acquire
higher than normal densities of Na+
channels in demyelinated regions
(restoration of conduction)

Degeneration of axons also

occurs in MS permanent loss of
function
Waxman et al. 2004
 Multiple Sclerosis
NaV1.2 channels in demyelinated axons

Increased NaV1.2
channel expression
 Multiple Sclerosis
NaV1.6 channels in demyelinated axons

Both NaV1.2 and NaV1.6 channels

produce rapidly activating and
inactivating currents that can support
action potential but

NaV1.6 channels produce larger Na+

current than NaV1.2 channels
 Multiple Sclerosis
NaV1.6 channels in demyelinated axons

Na+ channels and Na+/Ca2+ exchanger

co-localize in demyelinating axons (usually
with NaV1.6)
The activity of Na+ channels can trigger
Ca2+ - mediated injury of axons,
Na+ influx through Na+ channels and
reverse activity Na+ efflux and Ca2+
influx through Na+/Ca2+ exchanger
 Multiple Sclerosis
NaV1.6 channels in demyelinated axons