Exam 2 Powerpoint 10

Question 1

H and H experiments predicted

Answer 1

existence of channels

Question 2

Predictions (4)

Answer 2

Channels must allow ions to move across the membrane at a high rate

Channels make use of electrochemical gradients

Discriminating –specific for specific ions

Voltage sensitive; inactivating in some cases

Question 3

Technical impediment H and H faced

Answer 3

Voltage clamp allowed recording of
aggregate currents (thousands of channels): needed a new
method

Question 4

Patch Clamping was invented in

Answer 4

1976

Question 5

Patch clamping was made to

Answer 5

record activity of a single ion channel or whole cell currents

Question 6

Patch clamping forms a

Answer 6

high resistance seal between cell membrane and glass wall of
the micropipette (apply suction)

Question 7

Patch clamp recording allows

Answer 7

Allows recording from one ion channel

No ions escape b/c it is a very tight seal

Low level of background noise

Can alter the potential across the patch simply by applying
voltage to the pipette

 https://www.youtube.com/watch?v=mVbkSD5FHOw
 https://www.youtube.com/watch?v=TUoCQTwewVo

Question 8

Sodium currents Recorded in the
presence of _____ which blocks
__________ channels

Answer 8

Cs+

Potassium

Question 9

Macroscopic recordings are from

Answer 9

multiple channels

Question 10

Microscopic recordings are from

Answer 10

1 channel

Question 11

Depolarization increases the probability of

Answer 11

channel opening

Question 12

Potassium currents
in in presence of _____
which blocks __________________

Answer 12

TTX

sodium
channels

Question 13

___genes are associated with K channels

Answer 13

78

Question 14

____genes are associated with Na channels

Answer 14

29

Question 15

More than _____ genes encode for channels. Can vary due to RNA editing and splice variants.

Answer 15

230

Question 16

Why is it difficult to study specifics in vivo by patching onto neurons

Answer 16

Hard to tell individual channels apart (lots of noise)

 Challenging in telling different types of neurons apart

 Splice variants

 Pharmacological tools are not always great, although not always available

Question 17

The molecular structures of ion channels as individual proteins were determined after the cloning of genes encoding specific ion channels, as the revolution in
molecular biology spread to neuroscience in the 19__s.

Answer 17

80s

Question 18

Voltage-gated ____ channel proteins were first purifed from the electric eel Electrophorus electricus, whose electric organ is densely packed with ___ channels

Answer 18

Na+

Question 19

Xenopus System

Answer 19

1980s first demonstration that they can express ion channels

 Harvest eggs from females

 1 mm diameter

 Once you inject mRNA (you synthesize in vitro), makes a lot of protein

 Powerful—combine physiology and molecular methods

Question 20

Xenopus Oocytes

Answer 20

Immature eggs from the clawed African frog (Xenopus laevis) used as
expression machines

Channel genes can come from any organism (the beauty of a common
central dogma)

Question 21

How do we study channels

Answer 21

Use of well defined systems
(heterologous systems)
-Cell lines (cells in a dish)
-Frog oocytes

Genetic mutations in model
organisms
-Mice
-Fruit flies

Question 22

Natural toxins for sodium channels

Answer 22

 TTX (puffer fish)- paralyze
 Saxitoxin (red tide dinoflagellates)-
similar to TTX
 Scorpion toxins-alpha toxins
(paralysis, arrhythmias)-slow
inactivation of VGSC

Question 23

Natural toxins for potassium channels

Answer 23

x- TEA, cesium
x- Dendrotoxin (from wasps, snakes)
(causes convulsions)
- Block repolarization—prolong
action potentials
x- Apamin (bee)

Question 24

Major channel structure feature

Answer 24

Plasma membrane

Question 25

Channels insert

Answer 25

between membrane spanning structures, pores

Question 26

Selectivity filter:

Answer 26

Size of cation is a big factor

Question 27

Multiple subunits:

Answer 27

Repeating motifs within
these subunits

Question 28

Voltage gated channels have specific functional states:

Answer 28

Open
Inactivating
Inactivated
Closed

Question 29

States of a Voltage gated Na+ channel

Answer 29

Closed
open
inactivating
inactivated
closed

Question 30

States of a Voltage gated K+ channel

Answer 30

Closed
Closed
Open
Open
Closed

Question 31

Voltage sensing

Answer 31

Depolarization changes the structure

Question 32

How voltage sensing result in channel opening

Answer 32

 X-ray Crystallography studies: structure
 Mutational analysis: understand the important amino acids
 Combine mutational analysis with electrophysiology

Question 33

Positively charged amino acids in protein channels are

Answer 33

voltage sensors

Question 34

Na+ and Ca2+ channels:

Answer 34

1 large subunit: 4 repeat motifs with 6 membrane
spanning regions each (total: 24 transmembrane segments)

Question 35

K+ channels:

Answer 35

4 subunits come together to make a functional channel

Question 36

Cl- channels structurally distinct:

Answer 36

2 separate pores made by two sets of 2 identical subunits

Question 37

Auxiliary subunits such as beta, gamma are

Answer 37

Often important for channel functionality, such as inactivating