Neur 3101 / 6101 – 2017

Cellular Neuroscience
Lecture 3: Hodgkin-Huxley & action potential propagation - 1

John Bekkers
John Curtin School of Medical Research
Australian National University
John.Bekkers@anu.edu.au

Outline of lectures

Lecture 1: • Revision: Bioelectricity, the Nernst potential & circuit theory

(20 Feb) • Voltage clamp & current clamp
• Ion channel selectivity & gating

Lecture 2: • Electrophysiological recording techniques

(22 Feb) • A neurophysiologist’s toolbox:
Classes of ion channels and what they do

Lecture 3: • Hodgkin & Huxley’s theory of the action potential

(23 Feb) • Predictions of the HH theory
• Computer lab: 27 Feb, 6 Mar & 27 Mar (and pre-lab tutorials)

Lecture 4: • Initiation & propagation of action potentials

(27 Feb)
• Optical methods in neurophysiology

Lectures 9-11: • Synaptic transmission

(9, 15, 16 Mar)

1!
 Learning objectives – Lecture 3

• Understand Hodgkin & Huxley’s theory of the action

potential, including the ideas of:

• Activation

• Inactivation

• Deactivation

• Understand how the HH theory explains many features of the

action potential, including stimulus threshold and refractory
period (...to be continued in the lab classes)

• Have an understanding of how action potentials are

propagated down an axon

Why are we interested in the action potential (AP)?

• Sagittal section through

a rodent brain

• Tremendous diversity
in AP firing patterns in
different regions

• Information is moved
around by APs, and
somehow encoded in the
rate or timing of APs

“Rate coding”

“Temporal coding”

2!
 Early history of the AP

• Early speculation on the nature of the “nerve impulse”!

• Bernstein (1902): non-selective increase in membrane permeability!
• The sodium hypothesis (1930s)!
• Hodgkin and Huxley (late 40s/early 50s)!
• Nobel Prize (1963)!

Alan Hodgkin! Andrew Huxley! Jack Eccles!

Cambridge! Cambridge! ANU!

The squid axon action potential

Squid axon with internal electrode!

3!
 Current clamp and voltage clamp of a squid axon

Current Clamp:! Voltage Clamp family:!

Clamp Im, measure Vm! Clamp Vm, measure Im!

2 ms!

Whole-cell currents in squid axon

+Tetrodotoxin (TTX)! +Tetraethylammonium (TEA)!

→ Potassium currents! → Sodium currents!

+60 mV! +60 mV!

Vm! Vm!

-60 mV! -60 mV!

-120 mV! -120 mV!

IK! INa!

2 ms! 2 ms!

4!
 How do these currents explain the AP?
Current recorded in voltage clamp: Voltage recorded in current clamp:
ENa
+40
IK!
4

Membrane potential (mV)!

3 0 3
1 2
INa!
-40
1
2 4
Vm! -80 1 ms EK

1 INa & IK small è low membrane permeability è gNa & gK small è resting potential

2 INa activates quickly è increasing Na permeability è gNa activation è upswing of AP

3 INa inactivates & IK activates slowly è gNa inactivation & gK activation è downswing of AP

4 IK declines as K channels close è gK deactivation è return to resting potential

→ The AP can be predicted if we know how gNa and gK vary with Vm and time

Why did HH use voltage clamp?

Current Clamp:!
Ohm’s Law for neurophysiology:
Clamp Im, measure Vm!
Im = g (Vm - Eion)
Vm (mV)!

→ As stated in the previous slide, we want to

measure gNa and gK

1 ms!
Why is it better to fix Vm and measure Im
Voltage Clamp:! (i.e. use voltage clamp)?
Clamp Vm, measure Im!

→ Because gNa (and gK) vary with Vm (and time)

• By fixing Vm with voltage clamp you don’t have

to worry about ‘g’ changing with Vm while
you are measuring it

• Of course, ‘g’ can still change with time in a

voltage clamp experiment

5!
 Measuring activation
Plots of peak INa & IK versus Vm!

IK!

INa!

INa is zero at +40 mV!

• ENa = +40 mV (Nernst potential for Na)!

• INa is zero at this potential, because there is no electrical driving force for Na+!
(i.e. the concentration gradient balances the accumulation of charge, so no more ions flow)!

Measuring activation
Plots of peak INa & IK versus Vm!
Recall: INa = gNa (Vm - ENa)!

∴ gNa = INa / (Vm - ENa)!

ENa! We already know:!

ENa = +40 mV!

At each data point we can read off:!

Vm from x-axis!
INa!
INa from y-axis!

Vm - ENa! Hence, calculate peak gNa!

Vm! (→ lab class)

6!
 Measuring activation

Peak Na conductance (gNa) vs Vm:! Channel open probability (po) vs Vm:!

1.0 po!
1!
gNa!

“Boltzmann!
0.5! curve”!
Normalised

0.5

-80! 0! 80!
Vm!
→ Voltage dependence of gNa is due to the
-40 0 +40
voltage dependence of channel opening!
Membrane potential (mV)!

• gNa determines how many Na channels ‘want’ to open as Vm changes

• The “foot” of gNa helps determine the threshold for eliciting an AP

• A similar plot can be obtained for gK

The Hodgkin-Huxley model of gNa inactivation

Vm!
• INa activates (opens) then inactivates!
INa! (IK does not inactivate, but
it does deactivate)!
Time!
• 2 independent ‘gates’!

Activation gate!
Inactivation gate!

• Opposite voltage dependence:!

• depoln opens activation gate!
• depoln closes inactivation gate!

Closed! Open! Inactivated!

7!
 Measuring gNa inactivation
• Activation & inactivation gates are independent!

∴  Inactivation gate may have closed already, depending on the starting potential!

Starting Potential (-120 to -40 mv)!

Peak Na conductance (gNa) vs Vm:!
Strongly!
hyperpolarized! 1.0
(e.g. -120 mV)!

gNa!
Normalised
Less strongly! 0.5 If step Vm to 0 mV the
hyperpolarized! activation gate will open!
(e.g. -80 mV)!

Weakly! -40 0 +40

depolarized! Membrane potential (mV)!
(e.g. -40 mV)!
Activation gate is still shut at -40 mV!

Measuring gNa inactivation

• Activation & inactivation gates are independent!
∴  Inactivation gate may have closed already, depending on the starting potential!

Starting Potential (-120 to -40 mv)! Test Potential (0 mV)!

( At 0 mV activation gate is open)!

Strongly!
hyperpolarized! Large!
(e.g. -120 mV)! current!

Less strongly!
hyperpolarized! Small!
(e.g. -80 mV)! current!

Weakly!
depolarized! Zero!
(e.g. -40 mV)! current!

8!
 Measuring gNa inactivation
• INa evoked by a fixed Test pulse!
• Vary amplitude of Starting Potential (Vpre)! Steady-state inactivation curve!

All inactivation gates!

Test pulse = 0 mV! already ‘open’ at Vpre!
Prepulse (Vpre) =!
-40 mV!
I/Imax! 1!

-120 mV! 0.5!

0.5 nA! -120! 0!

Starting potential Vpre (mV)!

4 ms!

All inactivation gates!

already ‘closed’ at Vpre!

Measuring gNa inactivation

Na conductance (gNa) vs starting Vm:!
(“steady-state inactivation curve”)!

→ This is also a Boltzmann curve –

gNa!

a backwards one!!
Normalised

This is because the inactivation gate

also has a voltage-dependent
probability of being open (po) –!
but with the reverse polarity!

-120 -80 -40 0

Starting potential (mV)!

• This curve determines how many Na channels are ‘available’

to open from different starting membrane potentials

• It explains how, if the resting membrane potential becomes too

depolarised, Na channels are inactivated and APs fail

9!
 Summary: gNa activation and inactivation

Inactivation Activation
plot plot

gNa!
Normalised

-120 -80 -40 0 +40

Membrane potential (mV)!

• Opposite voltage dependence:!

Inactivation: depoln → fewer channels available to open!
Activation: depoln → more channels want to open!

We also need to know the kinetics

Vm! Potassium current:!

• Measure the speed at which IK turns on!

!(activates) and off (deactivates)!
IK!
• Do this at different voltages!

Vm! Sodium current:!

• Measure the speed at which INa turns on!

!(activates) then turns off (inactivates)!

• Do this at different voltages!

INa!

10!
 The HH equations

dVm!
I = Cm + INa + IK + IL!
dt!

dVm!
= Cm + gNa m3h (V - ENa) + gK n4 (V - EK) + gL (V - EL)!
dt!

where! dm!
= αm(1 - m) - βmm! [gNa activation parameter]!
dt!

dh!
= αh(1 - h) - βhh! [gNa inactivation parameter]!
dt!
dn!
= αn(1 - n) - βnn! [gK activation parameter]!
dt!

For a non-propagating AP, I = clamp current = constant!

Hodgkin & Huxley J Physiol 117: 500-544 (1952)!

Calculating the AP from the Hodgkin-Huxley theory

Stimulus
current:

Membrane
potential (Vm):

gNa • gNa increases and

gK decreases quickly!
Membrane
conductance (g): • gK increases and
decreases slowly!

Potassium
current (IK):
• IK is slower and
outward (+ve)!
Sodium • INa is faster and
current (INa): inward (-ve)!

11!
 Another way of thinking of the HH model of the AP

The action
• Inward INa drives a fast
potential is driven positive feedback cycle!
by two cycles:

• Outward IK drives a slower

negative feedback cycle!

Fig. 3.9, Purves et al “Neuroscience”

Include propagation along the axon

Na+ Na+

‘Positive
feedback’

12!
 Include propagation along the axon

K+ Na+ Na+

‘Negative
feedback’
AP propagation = ‘a burning fuse’

Putting it all together: The HH action potential

• AP is due to a fast inward flow of Na+ and a delayed outward flow of K+!

• The theory shows how to calculate 2 conductances, gNa and gK!

• gNa and gK both depend on Vm and time!

• gNa and gK both activate (↑) with depolarization!

! !(gK activates with a delay)!

• gNa (but not gK) also inactivates (↓) with!

!depolarization at later times!

• The AP propagates along axons like a

!burning fuse!

13!
 The theory is a triumph!
• HH theory and predictions for APs:!

The theory explains:!

• Why APs have a

Calculated! threshold!

• Why they fail if the cell

loses its resting
potential!

• How fast they

propagate down an
Measured! axon!

• How their properties

change with
temperature!

• BUT more recent research shows that some details of the theory are not correct!
• e.g. the activation and inactivation gates are not strictly independent

• Despite this, the theory is broadly correct, and widely used

Some historical movies

http://www.science.smith.edu/departments/NeuroSci/courses/bio330/squid.html!

14!