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1. Action Potential
2. Ionic Basis of Action Potential
Action Potentials
• What are they?
– Rapid reversal of the resting membrane
potential
0 mV
cell
-80 mV
3 ms
Electrophysiology Techniques
Lolligo pealeii
Squid giant axon
-1 mm in diameter
-1000X larger than most
axon
Reference electrode
Recording electrode
stimulus
Action Potentials
Important concepts:
• Threshold
• All or none
• Regenerative
• Conduction along axons
Action Potential
Rising phase or
depolarization
0 mV
Overshoot
Falling phase or
Repolarization
Threshold Potential
-80 mV
Resting membrane
potential
Undershoot or afterhyperpolarization
Reminders…
1. Action potential is a rapid reversal of Vm
2. Vm is dominated by the equilibrium
potential of the most permeable ion
3. Permeability controlled by ion channels
Ionic Basis of Action Potentials
PNa>>PK
PK>>PNa
PK>>PNa
ENa
0 mV
-80 mV
EK
Time 
Na+
K+ leak channel
K+
K+
Membrane Potential
Na+
Section of
Squid Axon
Voltage-gated channels
time
Membrane Potential
Na+
Na+
time
K+
K+
1. At rest only K+ leak channels open,
PK>>PNa
2. With stimulus, voltage-gated Na
channels open, PNa>>PK
Na+ flows into the cell carrying positive
charge
3. Delayed opening of voltage-gated K
channels, PK>>PNa
K+ flows out of cell removing positive
charge
Na+
Na+
K+
K+
How do we know Na+ important for
depolarization?
Replace Na+ in extracellular bath with
impermeable cation - choline
Normal
0 mV
-80 mV
Low Sodium
Ion currents underlying the AP
• Use voltage-clamp technique to measure
currents
• Measure currents in the presence and
absence of Na+
• What are ionic currents?
– So far, voltage (V)
– When ions move  current (I)
– Movement through channel is resistance (R)
• Reciprocal is conductance (g)
Ohm’s law
I=V/R or I=gV
Where g = 1 / R
• More properly
Iion = gion X emfion
• Iion is ionic current
• gion is ionic conductance
• emfion is the electromotive force acting on an
ion
• emfion = Vm - Eion
Total membrane potential
Nernst potential for the ion
Therefore,
Iion  gion (Vm  Eion )
• gion is controlled by ion channels
• If all channels closed, g = 0 and no ions
flow
• if Vm = Eion then emf = 0, and no ions
flow
• How are ion currents measured?
• Voltage-clamp
Electrophysiology Techniques
Voltage clamp
Control
amplifier
+
Command
Signal
Squid axon
Voltage output
Reference electrode
Recording electrode
Current
electrodes
Im
Current output
Membrane currents
• Measure ionic currents from squid axon
– To determine contribution of Na+ and K+
Measure in normal saline with Na+ and Na+free saline
Ion currents underlying the AP
Membrane
Potential
K+ current
Na+ free saline
outward
Ionic
Currents
inward
Total current
normal saline
Ion currents underlying the AP
K+ current
Na+ free saline
outward
Ionic
Currents
inward
Total current
outward
normal saline
Subtract K+ current from total
Ionic
Currents
inward
Na+ current
Ion currents underlying the AP
1. The Na+ current activates quickly and
then inactivates quickly
2. The K+ current activates more slowly
and persists longer
Ionic Basis of Action Potentials
PNa>>PK
PK>>PNa
PK>>PNa
ENa
0 mV
EK
-80 mV
Time 
Ion currents underlying the AP
Membrane
Potential
Ionic
Currents
Pk(leak) + Pk(volt)
Pk(leak)
After-hyperpolarization
K+ current
• The after hyperpolarization coincides
– with the persistent K+ current and absent Na+
current
Stimulus & Threshold
• The stimulus depolarizes the membrane
– Experimentally applied current
– Synaptic potential
– Receptor potential
Threshold
• The membrane potential at which
Na flowing into the cell exactly equals the
K flowing out of the cell
• A fraction more stimulus depolarization is
required to ‘fire’ an action potential
Threshold Potential
Larger stimulus
Reaches threshold
0 mV
-80 mV
Small stimulus
Below threshold
Positive Feedback
Na+ entry
Increased Na
permeability
Membrane
depolarization
The AP is regenerative and
displays all-or-none behaviour
Why does the AP stop rising?
ENa
1. As VmENa,
Na+ inflow stops
2. Na+ channels
inactivate
3. K+ channels
open, K+ outflow
starts
Refractory Period
1. A second stimulus very soon after the
first will not fire an AP (Absolute)
2. With a delay, a second stronger stimulus
will cause a small AP (Relative)
3. With longer delay a second AP can be
fired
Absolute refractory
period
A
B
C
Relative refractory
period
Why is there Refractory Period?
• The Na channel stays inactivated for a
short period of time after it closes
Closed
Active
Open
Inactivated
Closed
Active
Summary & Key Concepts
1. The AP is controlled by rapid changes in
ionic permeability
2. Permeability is a function of voltagegated ion channels
3. Threshold potential
4. Positive feedback
5. Refractory period has two phases