3. Lectures 6, 7, 8

Question 1

What are the properties of an action potential?

Answer 1

AP is a transient depolarization triggered by a depolarization beyond a threshold

1. Threshold
2. All or none event
3. AP is conducted without decrement. It had a self-regenerative feature that keeps the amplitude constant, even when it is conducted over great distances
4. Refractory period

Question 2

What are the 4 phases of the action potential?

Answer 2

Rising phase- depolarizing (positive going) phase is rapid and smooth increase in Vm from the negative resting potential to a maximum positive value that typically lies between +10 and +40 mV
Repolarizing phase- negative going, the repolarization phase vary considerably among different excitable tissues and cells
Afterhyperpolarization- when repolarization undershoots to a voltage minimum, more negative than Vrest
Overshoot- the part of the action potential that lies above 0mV
Slide 3 lecture 6

Question 3

What are the 3 things that affect the threshold, amplitude, time course, and duration of an action potential?

Answer 3

1. The gating and permeability properties of ion channels
2. The intracellular and extracellular concentrations of the ions
3. Membrane properties (cap, res, geometry of cell)

Question 4

What are the absolute and relative refractory periods of action potentials?
Where do they arise from?

Answer 4

Absolute- impossible to fire another action potential, goes from initiation of the spike to when repolarization is almost complete
Due to Na+ channel inactivation

Relative- the minimal stimulus necessary for activation is stronger or longer than predicted by the first AP
Due to high K+ conductance and minimal Na+ conductance

Aria from hating properties of Na+ and K+ channels and the overlapping time course of their currents
Slide 4 lecture 6

Question 5

What are Hodgkin and Huxleys big discovery with ions and APs?

Answer 5

Discovered what ions contributed to action potential
Changes in membrane potential depend on flow of ions into or out of the cell
These flows are caused by openings of ion channels, which are proteins that form pores through membrane
Found this in squids
Slide 6-7 lecture 6

Question 6

What do the sodium and potassium channels conductance have to do with action potential?

Answer 6

Slide 7-8 lecture 6
Sodium conductance is fast
Potassium conductance takes time so it’s used for the second part of the action potential (K+ peaks in repolarizing phase)

Both channels activate at same time potassium just takes longer to reach max

Summation of Na and K currents on slide 11-12 lecture 6

Question 7

What is the positive feedback for Na+ channel activation?

Answer 7

Depolarization of the membrane causes Na+ channels to open rapidly (increase in gNa), resulting in inward Na+ current.
This current, by discharging membrane capacitance, causes further depolarization, thereby opening more Na+ channels, resulting in a further increase in inward current. This regenerative process drives the membrane potential toward ENa, causing the rising phase of the action potential.

A slower negative cycle caused by opening of K+ channels repolarizes the cell membrane
Slide 9-10 lecture 6

Question 8

When do we see inactivation or sodium channels?

Answer 8

Over time!!

TIME MUST BE ON THE GRAPH

Question 9

How do the selective blocking of the sodium and potassium channels with pharmacological agents (tetrodotoxin, tetraethylammonium) change the current over time graphs of the current over time?

What does a current (Im) over voltage (Em) graph look like for potassium and sodium?

Answer 9

Slide 13 lecture 6
Slide 17 lecture 6

We cannot see where K+ reverses direction because Ek is much more negative than the threshold for K+ channels

Question 10

What is the time course of Na+ current on a graph of current over time?
What about K+?

Answer 10

Slide 14 lecture 6

Immediately after depolarizing voltage step to a Vm of -30mV, the inward INa (downward going) reaches a peak value and then returns to zero. The initial phase of this time course (before the peak) is called activation, and the later phase (after the peak) is called inactivation

Slide 19 lecture 6

Question 11

What are the 3 possible states of Na+ channels?

Answer 11

1. Resting
2. Activated
3. Inactivated

Activation and inactivation are voltage dependant processes triggered by membrane depolarization
Activation occurs more quickly than inactivation (Na channels can open and allow Na influx before the processes of inactivation close the channel)
Inactivated state can only be reverse by repolarizing the membrane to its negative resting potential

Slide 15-16 lecture 6

Question 12

What is open probability?

Answer 12

The open probability (Po) is a measure of the proportion of the total recording time that an Jon channel is in open state

At each voltage there is a specific open probability, the total conductance (G) for each ion increases with increases in voltage

Slide 21 lecture 6

Question 13

What determines the amount of current passing through each open channel?

Answer 13

Ohms Law
The current passing through each open channel will depend on the conductance (g) of that channel (ability to let ions pass through) and the driving force for that ion (determined by the difference between the membrane voltage and equilibrium potential)

Question 14

What are ion channel superfamilies?

Answer 14

Most of ion channels that have been described in nerve and muscle cells fall into a few gene superfamilies. Members of each gene superfamily have similar amino acid sequences and transmembrane topology and, importantly, related functions
Superfamilies look the same crossing the membrane

Each superfamily is thought to have evolved from common ancestral gene by gene duplication and divergence

Question 15

What are the 4 different types of ion channels’ structure?

Answer 15

Hetero-oligomers

Homo-oligomers

Single polypeptide

Channels with α subunits and with auxiliary subunits (β or γ) that modulate the hating of the pore

Slide 5 lecture 7

Question 16

What is the structure of the voltage-gated K+ channel and just the α subunit from the channel?

Answer 16

Slides 6-7 lecture 7
K+ ion channels are composed of 4 α subunits as well as other accessory subunits (β, γ, δ)

Each alpha subunit has voltage sensor domain to detect when membrane is being depolarized and a pore domain
Has six alpha helices domains

Question 17

How do Na+ and Ca2+ ion channels compare to K+ ion channels structure?
What are the 2 key structural elements?

Answer 17

Slide 8 lecture 7

Na and Ca channels are composed of a single pore-forming α subunit with 4 domains

Made of:

1. The S4 segment- positively charged residues every third amino acid and involved in voltage sensing
2. P region- dips into the membrane but does not cross it and responsible for ion selectivity of the channel (4 P domains)

Question 18

What is the selectivity filter?

Answer 18

Every ion is surrounded by water and has charge, at the selectivity filter an ion must shed most of its waters of hydration to traverse the channel; in their place weak chemical bonds (electrostatic interactions) form with polar (charged) amino acid residues that line walls of channel

Interaction with selectivity filter must compensate for the loss of the energy of interaction with water of hydration

Question 19

What are waters of hydration?

Answer 19

Ions in solution are surrounded by a cloud of water molecules that are attracted by the net charge of the ion
The smaller the ion, the more highly localized is it’s charge and the stronger it’s electric field
Smaller ions attract water more strongly
Na+ has a larger water shell than K+, because of the larger water shell, Na+ behaves as if it is larger than K
Slides 11-13 lecture 7

Question 20

What is the K+ selectivity filter?

Answer 20

Contains 4 positions that mimic waters of hydration with which potassium interacts with the channel
Potassium comes to channel presenting itself with waters of hydration but when it meets the port of channel it sheds water and becomes naked potassium and gets waters back once it’s through

Slide 14 lecture 7

Question 21

What is the Na+ selectivity filter?

Answer 21

Negatively charged carboxylic acid groups of glutamate or aspartate residues at the outer mouth of the pore perform the first step in the selection process by attracting cations and repelling anions
These substitute as water of hydration

Energetic cost of dehydration is not as great as in K+ channels since Na+ channel is large

Slide 15-16 lecture 7

Question 22

What are voltage gating channels structure?

Answer 22

Voltage gated cation channels are composed of pore forming α-subunits that each contain a motif consisting of six transmembrane segments (S1-S6), seventh hydrophobic region (P region) connects the S5 and S6 segments

Voltage gating channels have a gating charge, and a change in Vm causes this charge to move across the electrical field of the membrane, resulting in conformational changes that open/close channels

Slide 18 lecture 7
Slide 3 Lecture 11!!

Question 23

What is the voltage sensor?

Answer 23

The S4 segment has pattern of amino acids where every 3rd position has positive charged arginine or lysine. This pattern is absent in channels that are not voltage gated it is thought this is the voltage sensor

Voltage sensor is the part of the protein that transduces depolarization of the cell membrane into a conformational change that opens the channel

Slide 20-21 lecture 7

Question 24

Why do all of these voltage gated cation channels have similar structural components?

Answer 24

They are thought to have evolved from a common ancestor channel billions of years ago

Question 25

Where did Ca2+ channels originate from?

Where did Na+ channels originate from?

Answer 25

Theory that Ca channels arose from K+ channels through internal duplication events followed by specialization

Na channels arose from Ca channels through duplication of a Ca channel gene
Supported by observation that the first domain of the Na channel is most similar to the first domain of the Ca channel
Voltage gated Na channels are less diverse

Slide 3-4 lecture 8

Question 26

How do you record Nav channel current?

Answer 26

Cel attached configuration
Extracellular solution in pipette
Most Na channels activated by depolarization
Slide 5 lecture 8

Question 27

What type of patch clamp configuration was used on the diagram slide 6 lecture 8?
What solution is in pipette?

Answer 27

Whole cell configuration
Intracellular solution in pipette
Large smooth currents

Question 28

What are channelopathies?

Answer 28

Neurological diseases caused by altered function of ion channel subunits or the proteins that regulate them. These diseases may be either congenital (mutations or epigenetic mechanisms) or acquired (often resulting from autoimmune attack on an ion channel)

Something happened to channel that it can’t function properly

Slide 9 lecture 8

Question 29

What is hyperkalemic periodic paralysis?

Answer 29

A trigger stimulus of increase [K]o leads to depolarization, which activates Nav channels
The mutated Nav channels fail to inactivate and cause persistent Na inward current which sustains depolarization and K leaks out
Sustained depolarization inactivates normal Nav channels leading to loss of excitability and muscle paralysis

Question 30

How can low levels of Ca lead to neuronal excitability?

Answer 30

The voltage dependence of Na channels is highly dependant on concentration of Ca in the extracellular fluid and for this reason low Ca levels can lead to neuronal hyperexcitability

Question 31

What is tetrodotoxin (TTX) and Saxitoxin (STX)?

Answer 31

Tetrodotoxin is produced by certain marine bacteria, it’s accumulate in tissues of various invertebrates, amphibians, and fish. The internal organs of the puffer fish contain lethal amounts of TTX

Saxitoxin is produced by dinoflagellates that are responsible for “red tide” and by freshwater Cyanobacteria, which can poison ponds and rivers