Action Potentials

Purves et al., Chapters 3 & 4 (review figure from chapter 2)

Summary

Fig. 3.8
what we know about K+ and Na+ permeability during the passing of the action potential
Na+ conductance goes up then down early
K+ conductance goes up then down but much later.
There is something wrong with this figure: K+ conductance is much higher than Na+ conductance during the resting potential.

Bernstein knew about selective K+ permeability
Thought it was lost during the action potential (actually Na+ permeability increases)
Cole and Curtis used the AC Wheatstone bridge to show that the resistance decreased during the action potential
R1 & R2 divide one path, Rv (variable) and Ru divide the other
Galvanometer between two nodes
Ru = Rv x R2/R1
Now it is easy to realize that the Goldman is like the Nernst equation where the relative permeabilities of Na+ and K+ change

Fig. 3.11
An action potential is non-decremental

Alumnus research in neuroscience

Joel Geerling, a chemistry major, took this class (and also graduated) in 2000. He went to Wash U for an MD-PhD. Related to the importance of sodium (covered throughout this course and in this outline) is the hormone aldosterone from the adrenal cortex and its regulation of sodium in the kidney. It is well-known, especially by athletes, that a sodium deficiency leads to increased sodium appetite, Joel's work, in the 4 papers referenced below, addresses this issue at the level of the brain. They illustrate the importance of understanding techniques such as confocal microscopy, as well as brain anatomy.
JCGeerling et al., Aldosterone target neurons in the nucleus tractus solitarius drive sodium appetite. J. Neurosci 26, 411-417, 2006
JCGeerling et al., Aldosterone-sensitive neurons in the rat central nervous system, J Comp Neurol, 495, 515-527, 2006
JCGeerling & ADLoewy Aldosterone-sensitive neurons in the nucleus of the solitary tract: Bidirectional connections with the central nucleus of the amygdala, J. Comp Neurol, 497, 646-657, 2006
JCGeerling & ADLoewy Aldosterone sensitive neurons in the nucleus of the solitary tract: Efferent projections, J Comp Neurol 497, 223-250, 2006

Passive spread of potential along axon

Fig. 3.10 A
If there were not something very special (voltage gating that activates Na+ channel), passive voltage spread would be decremental
Current down along the axon gets smaller because it leaks through the membrane resistance and capacitance.
At any place along the axon, a spike would depolarize the axon to threshold for a spike a certain distance ahead of it, and that distance depends on the square root of the radius.
Spike at one place would also depolarize the axon behind it to threshold, but it does not generate a retrograde action potential because of the refractory period, explained (below) by the inactivation of the Na+ channel.

Fig. 2.2 (again)
TERMS: threshold, generator potential, all-or-none, refractory, unidirectional
NOTE also the membrane acting as a low pass filter

Fig Box 3C [first figure] shows exponential decay and space constant (lambda)
Fig Box 3C [second figure] shows exponential charging of capacitance and time constan (tao)

Cable equation
Here's a pdf of the transparency I'm showing you

Personal reflection. My fellow graduate student, Paul Kottler, and I took Warren Dennis's course (Physical chemistry of cell systems) together; we also studied for our PhD exam together. My mentor, Jerry Wasserman, had been famous for asking a question about the speed of the action potential and asking it each subsequent year because he was never satisfied with the answer. We resolved to retire this question since we knew that our coverage on the cable equation was much more than Wasserman would expect. When I found out that I passed, I asked Wasserman how he liked my answer and he replied that "it was ok, a little theoretical." So Paul and I decided we had done our duty for posterity, retiring the question, by answering it with a derivation that involved differential equations.

Summary:
(1) an action potential at one place depolarizes the membrane ahead of it to threshold.
(2) the spread is passive.
(3) current down the axoplasm leaks out through membrane resistance and capacitance.
(4) solving, space constant varies with square root of radius, time constant independent of radius.
(5) that is why invertebrates use giant axons for fast propagation.
(6) myelinated axons also have faster propagation for larger axons.

Coding
frequency of action potentials, not size since they are all-or-none
sometimes action potentials come in bursts
or at beginning of depolarization because of "adaptation"

Hodgkin Huxley experiments

Fig. 3.12
Propagation of action potential (spike) shown with opening and closing of Na+ and K+ channels drawn in

Note relative opening and closing of channels
"sodium pump" already established the ion gradients
oscilloscope essentially graphs voltage as a function of time
action potentials can also be listened to on a loud speaker
activation, inactivation, voltage gating

Fig. Box 3A
general recording "geometry" -
differential amplifier compares 2 voltages and puts out current
operational amplifier is a differential amplifier to clamp voltage
space clamp - really just do whole axon at once

Fig. 3.2
voltage clamp data
voltage clamp - change voltage then pump and monitor current needed to keep it there
I - t curves

Fig. 3.3
divide into early and late components as I - V curve
Ohm's law: E=IR, thus, the axes of an I-V curve are reversed and the slope is
conductance = 1/R in units of Siemens (formerly "mho")
NOTE: iNa = gNa(V-VNa) - driving potential

Fig. 3.4
experiment with low Na+ to show early current is Na+

early fast sodium tetrodotoxin (TTX) sensitive (see box C, Chapter 4, on toxins)
TTX from puffer fish, puffer fish is a delicacy in Japan, but careful preparation is importantto prepare sushi, best if enough TTX left to make mouth numb
saxitoxin from dinoflagellates (red-tides are "blooms" and filter feeding shellfish can become poisonous
Other experiments (e.g. dose of TTX) show few sodium channels, works only if applied to outside of axon

late slow potassium TEA (tetraethyl ammonium) sensitive

Channels

Fig. 4.3
In summary, resting potential is based on predominant K+ permeability
then Na+ channels activate
then Na+ channels inactivate
then a late K+ channel activates

GENERALIZATION - action potential is based on Na+ and K+
there are MANY other channel types

Figures Box 3B
for 20 years they have been studied by "heterologous expression" in cells like Xenopus oocytes
inject exogenous mRNA into clawed African frog egg

Channels are at a low "concentration" (except in post-synaptic membrane).
It takes little tetrodotoxin to block action potential so they are proteins that are not highly expressed.

Thus, channel mutants might be lethal, so they used tricks to get genes like conditional mutants (like temperature sensitive with permissive and restrictive temperatures)

Fig. 4.5 ABC
KV2.1 (A) is like "delayed rectifyer" K+ channel of action potential. "Rectifier" means that it only allows current in one direction. KV4.1 and HERG have inactivation.

One famous conditional channel mutant in Drosophila, ether-a-go-go, shakes under ether anesthesia. A hunan homologue was found, HERG = human ether-a-go-go-related-gene. HERG inactivates so quickly that it only opens after voltage is over. Contributes to long action potential in cardiac muscle. Long QT syndrome is sometimes mutation of HERG, QRS in EKG (electrocardiogram) is ventricular depolarization, T is repolarization.

A lot of work was done with KV4.1 (B)

Fig. 4.6C
K+, A-type conductance, not at all like K+ channel of action potential
sea slug Anisodoris, Drosophila fruit fly - Shaker mutant
Tetramer makes channel, each component crosses membrane 6 times with hydrophobic domains, S1-S6, inactivation is "stopper" on chain at N-terminal, voltage gating is S4 with + charged arginines or lysines every 3 or 4 amino acids, rotates and moves. Pore is between S5 and S6, not so hydrophobic.

Fig. 4.6G
This figure shows a 12-transmembrane protein for the Cl- channel
A famous Cl- channel is the cystic fibrosis transmembrane conductance regulator (CFTR)
cystic fibrosis is most common genetic disorder in Caucasians (1/2000), lungs fill up with thick mucus. One presumes the channel is two components.

Fig. 4.6A
Sodium channel, now diverse (human 10 genes)
electric eel Electrophorus electricus 600 V
Huge - 1820 amino acids - "pseudotetramer"
S4 - gating - positively charged (basic) arginine (R) or lysine (K)

Fig. 4.7
rotation
Pore between 5 & 6 (not hydrophobic)

Fig. 4.1B
low current (1-2 pA)
low conductance - 10 pS
stopper to inactivate.

There are different types of Na+ channels, and some are targets of local anesthetics benzocaine and lidocaine.

Potassium channels (lots of them, 100)
Leak (resting potential) 20 pS
Delayed rectifier (repolarization of action potential) 10 pS
Anomalous rectifier - maintain depolarization - cardiac, fertilization
HERG human ether-a-go-go related gene

Calcium channels
16 genes
Ca2+ regulation by parathormone, calcitonin and vitamin D important
Ca2+ channel in synaptic terminal vesicle release - very important, also many others
Ca2+ channel is receptor for IP3 (inositol trisphosphate "second" messenger) on smooth endoplasmic reticulum
Ca2+ channel in muscle sarcoplasmic reticulum
Ca2+ channel in t- (transverse-) tubule in muscle

Box 4C
Toxins
Tetrodotoxin puffer fish (saxitoxin dinoflagellates) block Na+ channel
scorpions
and many others

Fig. Box 4D
genetic diseases of channels
myotonia (stiffness from too much excitation) from Cl- channel defect

Fig. Box 4D
paralysis from Ca2+ channel defect
CSNB from Ca2+ defect Congenital (i.e. genetic) stationary (as opposed to degeneration) night blindness (would affect rods)

Fig. Box 4D
myotonia, paralysis or stiffness from Na+ channel
Long QT syndrome from Na+ or K+ channel defects EKG (electrocardiogram) has PQRST waves, P from atrial depolarization, QRS from ventricualr depolarization and T from ventricular repolarization
K+ channel from HERG = human ether-a-go-go(EAG) related gene EAG - Drosophila twitch under ether anesthesia

Fig. 4.8
Structural studies on bacterial K+ channel - it takes a lot of protein to do X-ran crystallography

Fig. 4.8
selectivity by pore size
interesting that non-hydrated ion passes.
Hydrated - size is inverse
Li > Na > K > Rb > Cs (lyotropic series)

Fig. 4.6
There are a lot of configurations of channels

Exam questions from 2005 - 2007 relating to this outline

How did the electric eel Electrophorus electricus assist in the isolation of a channel?

sufficient concentration of sodium channel to allow characterization

In theory, and in data, what is the direction and amount of Na+ current when the voltage in the axon is clamped to the Na+ equilibrium potential?

none

Paving the way for the Nobel Prize winning Hodgkin and Huxley work, what could you conclude from the Cole and Curtis finding that the AC bridge went out of balance as the
action potential goes by?

conductance increased

In terms of amino acid sequence, how does the S4 of the voltage-gated Na+ channel differ from the typical transmembrane alpha helix?

charged arginines or lysines every 3 or 4 amino acids

To hold the voltage of a squid giant axon at a clamped level of 0 mV, Hodgkin and Huxley had to pump current out through the membrane at 0.5 ms (fairly early) to compensate for
what?

sodium current

What does tetrodotoxin block?

the sodium channel

"Long QT syndrome can be caused by a mutation of HERG." Translate.

genetic long myocardial action potential

How does the space constant of an axon relate to the axon's size?

with square root of radius

Some potassium channels do show inactivation. What part of the molecule is responsible?

stopper on the N-erminus

CSNB is a channelopathy. S=stationary (not progressive degeneration). NB=night blindness (affecting rod photoreceptors). Why is the term C=congenital applied?

it's genetic

An action potential depolarizes the axon ahead of it to threshold, and that is why the action potential propagates. It would also depolarize the axon behind it. Why does it not cause a
backward action potential?

refractory potential, inactivation

Selecting for conditional channel mutants, like temperature sensitive mutants, has been especially useful in Drosophila. Why not just isolate regular mutants?

they might be lethal

Why would two resistors in series connected to a battery be called a Voltage divider?

Two sources of voltage (Ohm's law) E=IR, E = IR1 + IR2, used in Wheatstone bridge

In discussing the passive properties (i.e. without an action potential), the current gets smaller with distance from the stimulus going along the axoplasm (down the inside of the axon). Why does it get smaller?

along the way, it is lost through the membrane (the membrane's resistance and capacitance)

The Shaker K+ channel is a tetramer of proteins that each cross the membrane 6 times. Why is the Electrophorus Na+ channel called a pseudotetramer instead?

because one huge molecule has 4 repeated domains each the size of one shaker channel protein

When (or why) is there a negative conductance region in the I-V curve in Voltage-clamp data?

early, when Na+ channels activate then inactivate

What would be the cause of death if you ate a puffer fish?

Na+ channel block, no action potentials

What does an alpha helix, namely S4, with positively charged (basic) amino acids [arginine (R) or lysine (K)] every 3 or 4 amino acids do for the action potential's sodium channel?

it detects Voltage to gate the activation of the channel

Cole and Curtis already knew that the conductance went up as the action potential went by. Hodgkin and Huxley went a bit further. Which conductance(s) went up?

Na+ and K+

Give the name of at least one "channelopathy" (genetic disease resulting from a mutation affecting a channel protein).

paralysis, myotonia, long QT syndrome, congenital stationary night blindness

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This page was last updated 1/28/08