L14 - Neuronal and Muscle Toxins

Question 1

What toxins is the neuronal Na channel affected by?

Answer 1

Tetrodotoxin

Saxitoxin

Question 2

What toxins is the muscle Na channel affected by?

Answer 2

Tetrodotoxin
Saxitoxin
Conotoxin

Question 3

What toxins is the Ca channel affected by?

Answer 3

Conotoxin

Question 4

What toxins is the Kv K channel affected by?

Answer 4

Dendrotoxin

Question 5

What toxins is the AchR channel affected by?

Answer 5

Tubocurarine

Question 6

What toxins is Ach release affected by?

Answer 6

Tetanus toxin

Botulinum toxin

Question 7

What is tetrodotoxin?

Answer 7

Guanidinium neurotoxin
100 times more lethal than spider toxins
10,000 times more lethal than cyanide
Only nM levels required

Question 8

What does tetrodotoxin inhibit?

Answer 8

Voltage gated Na channels

Question 9

What is the source of tetrodotoxin?

Answer 9

Produced by marine bacteria found in invertebrates, amphibians, fish
- Puffer fish, blue ring octopus

Question 10

What are the symptoms of ingestion of tetrodotoxin?

Answer 10

Numbness of lips and tongue
Facial paraesthesia - abnormal sensation 
Headache
Nausea
Dizziness
Diarrhoea
Vomiting
Increasing paralysis and respiratory paralysis - death

Question 11

What is the rate of onset of symptoms following ingestion of tetrodotoxin?

Answer 11

20 mins to 8 hours

Question 12

What are the symptoms of a bite containing tetrodotoxin?

Answer 12

Same as ingestions but without facial effects

Faster death

Question 13

What are the treatment for tetrodotoxin poisoning?

Answer 13

Mechanical ventilation

No anti-venom - tetrodotoxin binds too strongly to site of action

Question 14

In tetrodotoxin what does the inhibition of voltage gated Na channels lead to?

Answer 14

Failure of neurotransmission – reduction in Ach release at NMJ

• Loss of sensation and then muscle paralysis
• • Because sensory neurons meet the toxin first
• Respiratory paralysis and death

Question 15

What are some examples of tetrodotoxin sensitive channels?

Answer 15

Nav 1.1 SCN1A – CNS and heart
Nav 1.2 SCN2A - CNS
Nav 1.3 SCN3A – CNS and heart
Nav 1.4 SCN4A - skeletal
Nav 1.6 SCN8A - CNS and PNS
Nav 1.7 SCN9A - PNS

Question 16

What are some examples of tetrodotoxin insensitive channels?

Answer 16

Nav 1.5 SCN5A - heart
Nav 1.8 SCN10A - sensory
Nav 1.9 SCN11A - PNS

Question 17

Which channel highlighted the variable sensitivity to tetrodotoxin?

Answer 17

Rat Nav type II

Question 18

Rat Nav type II tetrodotoxin sensitivity?

Answer 18

Wildtype rat channel - tetrodotoxin sensitive
Mutations in E387Q in pore region of rat channel
- Glutamate –> glutamine
- Tetrodotoxin insensitive
- Shows that tetrodotoxin sensitivity is based on one amino acid substitution in pore region

Question 19

Heart Nav 374 tetrodotoxin sensitivity?

Answer 19

Heart Nav 374 – cysteine - tetrodotoxin insensitive

Change heart 374 to tyrosine - tetrodotoxin sensitive

Question 20

What animals shows variable sensitivity to tetrodotoxin?

Answer 20

Garter snakes
- Difference geographical areas
- Different tetrodotoxin sensitivity
- Different Nav sequence IV domain
Both eat salamanders which can contain tetrodotoxin
- Only 1 snake population affected
- The snake population that survived was able to reproduce and pass on this gene

Question 21

Where are dendrotoxins found?

Answer 21

Snake neurotoxins found in Mamba family

Only nM levels required

Question 22

What do dendrotoxins inhibit?

Answer 22

Inhibit Kv channels

• Different types block different Kv subtypes
• Inhibition of repolarisation

Question 23

What are the symptoms of infection with dendrotoxins?

Answer 23

Early weakness and numbness in the bitten extremity
Systemic responses 30 minutes to 1 hour after bite
Ptosis - drooping eyelids
Ophthalmoplegia – paralysis of eye muscles
Disphagia - difficulty swallowing
Paresis - mild paralysis
Respiratory failure - death

Question 24

In dendrotoxin what does the inhibition of voltage gated K channels lead to?

Answer 24

Delays repolarisation of NMJ
Excess release of Ach from pre-synaptic membrane leading to Ach depletion
Failure of subsequent neurotransmission
Problems with muscle control - respiratory paralysis and death

Question 25

Where is conotoxin found?

Answer 25

Found in cone snails

Question 26

What are the different types of conotoxin?

Answer 26

Over 8000 different conotoxins - 19 major gene families Multiple and diverse targets - many channel types Varying lethality

Question 27

What are conotoxins being investigated for/

Answer 27

Parkinson’s, MS, lung cancer and chronic pain

Question 28

What are the symptoms of infection with conotoxin?

Answer 28

``` Burning pain Swelling at site of toxin injection Numbness Cardiac and respiratory distress Muscle weakness Loss of coordination Ptosis Headache Nausea Stomach cramps ```

Question 29

What are the symptoms of infection with conotoxin in severe cases?

Answer 29

Lips became stiff Blurred vision Paralysis Coma

Question 30

What are the two conotoxin pain targets?

Answer 30

Found in nociceptive nerve endings Ca 2.2 - Found in pre-synaptic membrane and sensory neurones - Mediates the release of neurotransmitter and therefore propagate pain - Inhibited by Eu1.6 Na 1.8

Question 31

What does Eu1.6 inhibit in HEK cells?

Answer 31

Cav2.2 In control as cell depolarised the Ca currents increase In Eu1.6 the Ca currents are much smaller --> inhibits Cav2.2 --> fewer vesicles fuse --> less neurotransmitter

Question 32

What does Eu1.6 inhibit in rate DRG cells?

Answer 32

Cav2.2 Looking at Cav2.2 in a cell where it is normally expressed Used barium – do not get the negative feedback seen with calcium which inhibits the channel In Eu1.6 the currents are much smaller than in control - blocks 1/3 of the channels - 30% inhibition

Question 33

What experiment was used to see if Eu1.6 suppresses pain?

Answer 33

Rat partial nerve ligation - Rat anaesthetised and sciatic nerve exposed - Pain model – 1/3rd of the sciatic nerve tied off - Application of pressure to hind paw until withdrawal

Question 34

What were the 3 groups tested during the rat partial nerve ligation?

Answer 34

No pain relief – saline - Feel pain with a low amount of pressure - Positive control - morphine and gabapentin - After injection able to withstand higher pressure Eu1.6 - After injection able to withstand higher pressure

Question 35

What is the overall effect of Eu1.6?

Answer 35

Inhibition of voltage gated Ca2+ channels Reduction in release of neurotransmitter at synaptic cleft Reduction in pain signalling

Question 36

What are two examples of tarantula toxins?

Answer 36

µ-TRTX-Tp1a and GpTx-1

Question 37

What do µ-TRTX-Tp1a and GpTx-1 do?

Answer 37

Inhibit Na+ currents through Nav1.7 --> can’t generate action potentials in sensory nerves - Potential to block pain signals Open probability is less in the presence of the toxin

Question 38

What experiment was undertaken to see if GpTx-1 was an analgesic in mice?

Answer 38

Pain model - 40 µl of OD1 injected into hindpaw | GpTx-1 then blocks the Na channels that OD1 activated

Question 39

What is OD1?

Answer 39

Scorpion toxin

Question 40

What does OD1 causer?

Answer 40

40 µl of OD1 injected into hindpaw Activates Nav1.7/1.6 Activates action potential firing in A and C fibres -->channels stay open for longer - Pain felt in paw – shown by lifting, shaking and licking