Lecture 5

Question 1

What is the structure of Cav channels

Answer 1

24 TMDs (four lots of six), 4 pore regions, 4 voltage sensitive regions

Question 2

What regulatory subunits are present on the Cav channels

Answer 2

Variety of regulatory subunits
Beta1-4, alpha 2 and delta subunits interact with the alpha 1 subunit and regulate its activity, sensitivity to voltage and to calcium itself.

Question 3

Which Cav channel types are regulated by high voltages

Answer 3

L type, N type and P/Q type

Question 4

Which Cav channels are regulated by intermediate voltages

Answer 4

R type

Question 5

Which Cav channels are regulated by low voltages

Answer 5

T type

Question 6

What type of channel is Cav1.2

Answer 6

L type

Question 7

What is the gene encoding Cav1.2

Answer 7

CACNA1C

Question 8

Where is Cav1.2 found

Answer 8

Heart brain and lungs

Question 9

how many alternative splicing loci does the CACNA1C gene have

Answer 9

12 - leading to 42 splice variants

Question 10

Where are the Cav1.2 channels found in the heart

Answer 10

On the T tubules close to the sarcoplasmic reticulum

Question 11

Which two types of inactivation does the Cav1.2 channel exhibit and what type of activation?

Answer 11

Voltage dependent activation
Voltage dependent inactivation
Ca dependent inactivation - calcium that enters feeds back on the channel causing it to close. (beta subunit plays important role in regulating inactivation)

Question 12

What are some symptoms of Timothy syndrome

Answer 12

Webbed digits, heart arrhythmia, immune deficient, increased risk of autism etc

Question 13

In the context of LTQS how does Timothy syndrome effect the patient

Answer 13

Patiens show a 2:1 atrioventricular block - 2P waves to each QRS. Meaning there is a mismatch in timing between the atria and the ventricles, as a consequence there is an increased risk of arrhythmia and sudden cardiac death.

Question 14

What changes in the T wave signify a problem with repolarisation in cardiac myocytes.

Answer 14

Alternating T wave polarity - T wave goes from one direction to the other. Sign of a significant repolarisation defect in ventricular muscles.

Question 15

What is ventricular tachycardia

Answer 15

Fast repetitive contractions of the ventricles in a completely uncoordinated manner with relation to the atria.

Question 16

What were the findings of a study into 17 children with Timothy syndrome

Answer 16

10 of the children died during the study at a mean age of 2.5
12/17 experienced life threatening arrhythmias and the 7 survivors had motor and cognitive impairment

Question 17

Why is motor and cognitive impairment common in patients with Timothy Syndrome

Answer 17

Because Cav1.2 is also found in the brain with impacts on development.

Question 18

What mutation of the Cav1.2 channel do patients of Timothy syndrome suffer from.

Answer 18

Not a mutation - instead it is an unusual splice form of Cav1.2
Gain of function seen at position G406R (at the end of the first TMD)

Question 19

What is different to most other Cav channels in the splice variant of Cav1.2

Answer 19

Glycine at position 406 becomes an argenine - glycine seems important due to its conservation between Cav channels.

Question 20

What is the result of the G406R splice variant on IV on inactivation of Cav1.2

Answer 20

Slowing of inactivation - activation is normal.
Less inactivation over the same time period as the WT channel. Effect is on voltage dependent inactivation - with only a small impact on Ca dependent inactivation

Question 21

How does the voltage dependent inactivation differ between the WT Cav1.2 and G406R splice variant at 0mV

Answer 21

In the WT at 0Mv there’s only about 20% of the Ca current left. In the G406R splice variant there’s about 80% left - this means the Cav1.2 channels are staying open longer and prolonging the plateau phase, also prolonging the QT

Question 22

What causes the uplift in both WT and G406R Cav1.2 channels at the lower end of relative Ca current

Answer 22

Relief from the Ca dependent inactivation of the channel.

Question 23

How is Ca dependent inactivation of the Cav1.2 channel prevented experimentally

Answer 23

Using Ba instead of Ca. Ba doesn’t feedback and block the channel, therefore the only inactivation is dependent on the voltage changing. (Cav channels will also allow other divalent cations through its pore)

Question 24

What does the use of Ba show when repeating IV analysis of both WT and G406R Cav1.2 channels

Answer 24

Splice variant shows almost complete loss of voltage dependent inactivation, almost all the inactivation of G406 Cav1.2 channels is therefore mediated by Ca feedback.

Question 25

What percentage of channels would be expected to be the G406R variant if the patient was heterozygous

Answer 25

predicted 11.5% of channels are the G406R variant however this is capable of extending action potentials by 17%

Question 26

Which SQTS types are due to changes in Cav1.2 function

Answer 26

SQTS4 and SQTS5
4 = CACNA1C effecting the alpha subunit of the Cav1/2 channel (loss of function)
5= CACNB2B effecting the beta subunit (loss of function)

Question 27

What changes are seen on a patients ECG suffering from SQTS4/5

Answer 27

Accentuated J-wave (just after the QRS wave

ST segment is also elevated.

Question 28

What is the mutation responsible for beta subunit inactivity

Answer 28

S481L

Question 29

Which two mutations have effects on the alpha subunit

Answer 29

G490R - Between the first loop between TMD 6 and 1

A39V - N terminal mutation

Question 30

What effect does the A39V N terminus mutation have on Ca current

Answer 30

Smaller Ca current, about 25% smaller means the plateau phase will be shorter and repolarisation will occur earlier

Question 31

What effect does the G409R mutation have on Ca current

Answer 31

Only produces a tiny current, they are very slow calcium currents through this specific calcium channel

Question 32

Is the impact of the Cav1.2 mutants likely to be due to gating or trafficking

Answer 32

Gating - location studies showed that Cav1.2 expression was similar in the WT and mutant forms. Only the A39V showed a potential trafficking problem