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ZOO4670/5670 Lecture No. 6
Neurotoxins as Tools for Studying Ion
Channel Structure and Functions
Qian-Quan Sun Ph.D ([email protected])
Univ. Wyoming
Today’s outline
• 1. Basics about channel block
• 2. Snake toxins and potassium channels
• 3. Sodium channels: toxins and local
anesthetics
• 4. Sea snails, spiders and calcium
channels
1
Apamin
Honey bee
Blocks Ca2+ activated potassium channels
Honey bee
Stichodactyla Toxin
Sea Anemone
Blocks voltage-gated potassium channels
sea anemone
2
3
Palytoxin
Capsaicin
Soft coral
Activates sodium channels
Cayenne Pepper
Excites peripheral nerve endings
4
Batrachotoxin
Maculotoxin
Poison Arrow
Frog
Blue-Ringed
Octopus
Prevents sodium channels from closing
Blocks sodium channels
5
Model or
scheme
normal function
State 1
State 2
k21
open
closed
units: M-1s-1
units: s-1
simple block
k21
closed
all molecules
begin here at
t= 0
open
k23 = k+[Drug]
drug
blocked
current
time constant
= 1/k21
time constant
= 1/(k21+ k23)
none
Ligand binding and dose response curve
Concentration of toxin (nM)
6
use-dependent block= open channel blocker
inside
“Trapped” or
“Use-Dependent”
Blocker
Functioning channel
Antiarrhythmics (heart)
“use-dependent blocker”
example: (procainamide)
Antiepileptics / anticonvulsants (brain)
“use-dependent blocker”
(phenytoin = Dilantin® )
A use-dependent blocker
stimuli
impulses fail
impulses
(voltage)
channel
population
(currents)
threshold
pronounced block
at brief intervals
little block
at long intervals
7
Green mamba snake
Pore region sequence of ShaKv1.1.
The dendrotoxin receptor
surface of ShaKv1.1.
KcsA
The surface color represents the change in binding
energy of dendrotoxin associated with mutation
8
Conclusions: dendrotoxin binds near the pore entryway
but does not act as a physical plug.
Hypothetical binding orientation for dendrotoxin
Scorpion toxin fits even better to the pore entryway!
Binding orientation of scorpion toxin on the K. channel.
9
C11H17N3O8
beta-scorpion toxins
Tetrodotoxin
alpha-scorpion toxin
Sea anemone
Motif III
b-scorpion toxins trap
IIS4 in the outward,
activated position.
Channel must be
activated for toxin
effect.
a-scorpion toxin trap IVS4
in an inward, partially
activated position.
Prolonged depolarization
drives the toxins off their
receptor site
Kd =0.1nM
Motif I
Motif IV
10
Neurotoxins that block Na+ channels
Antiarrhythmic
Drugs
Lidocaine
Mexiletine
Quinidine
Flecainide
Local Anesthetics
Etidocaine
Lidocaine
Many others
Anticonvulsants
Phenytoin
Carbamazepine
Lamotrigine
use dependent block
11
Baldomero “Toto” Olivera Ph D
[email protected]
Distinguished Professor
Olivera lab directory
Geographer’s cone
Magician’s cone
Leopard cone
Cloth-of-gold cone
12
A conotoxin:
27 amino acids
held together by disulfide bonds
individual conotoxins specifically block
individual ion channels.
13
The lethal sea snail toxin is a
promising pain killer .
Ziconotide is the synthetic equivalent of a 25-amino-acid, polybasic peptide from the venom of Conus
magus, a marine snail. It is the first selective N-type, voltage-sensitive calcium-channel blocking agent to
reach clinical trials.
In the current study, patients aged 24 to 85 years with refractory pain from advanced cancer or AIDS were
randomized to ziconotide or placebo in a 2:1 ratio. Intrathecal ziconotide was titrated over 5 to 6 days,
followed by a 5-day maintenance phase. Mean pain scores improved 52.9% in the ziconotide group,
compared with 17.5% in the placebo group, with no reduction in efficacy during the maintenance phase.
Five patients receiving ziconotide achieved complete pain relief.
Ziconotide also seems to improve the sleep pattern in some patients with chronic severe pain. Patients
experienced positive effects on nocturnal sleep duration and believed that remaining pain symptoms
interfered less with their daily life.
In experimental studies of absorption, distribution, metabolism, and elimination, intrathecal ziconotide
appeared and diminished rapidly in plasma and resulted in relatively little plasma protein binding.
Intravenous ziconotide degraded in animal brain tissue in 2 to 24 hours, produced no detectable
intermediates, and was cleared quickly from both cerebrospinal fluid (CSF) and the circulatory system.
This accelerated clearance rate likely means that the distribution of ziconotide throughout CSF and
metabolism within CSF are also rapid.
14
Cerebral cortex (layer II/III)
Thalamus
Trigeminal nucleus
15
Voodoo
• Ancient haitian traditions used the puffer fish for its voodoo
practices.
– The tetrodotoxin is used to create zombies
– It puts the people in a death-like state
• The towns people would take
some to be put into this state
– While under this state, they
would be forced to do odd jobs.
- The zombies were used as a
social damper to scare towns
people to keep them in line.
Toxins that act on voltage-gated ion channels
Name
Source
Neuronal Action
Agatoxin
Funnel Web
Spider
Blocks calcium channels
Agitoxin
Scorpion
Blocks potassium channels
Apamin
Honey bee
Blocks potassium channels
Atracotoxin
Blue Mountains
Funnel Web
Spider
Blocks voltage-gated calcium channels
Batrachotoxin
Poison Arrow
Frog
Prevents sodium channels from closing
Capsaicin
Cayenne Pepper
Excites peripheral nerve endings
Charybdotoxin
Scorpion
Blocks potassium channels
Ciguatoxin
Dinoflagellate
Opens sodium channels
Conotoxin
Marine Snail
voltage-sensitive calcium channels; sodium channels
Dendrotoxin
Green Mamba
Blocks voltage-gated potassium channels
Grammotoxin
SIA
South American
Rose
Tarantula
Blocks calcium channels
Gonyautoxin
Dinoflagellate
Blocks sodium channels
Iberiotoxin
Scorpion
Blocks potassium channels
Kaliotoxin
Scorpion
Blocks potassium channels
Kurtoxin
South African
Scorpion
Blocks calcium channels
16
Toxins that act on voltage-gated ion channels: continued
Maculotoxin
Blue-Ringed
Octopus
Blocks sodium channels
Margatoxin
Scorpion
Blocks potassium channels
Noxiustoxin
Scorpion
Blocks sodium channels
Palytoxin
Soft coral
Activates sodium channels
Phoneutriatoxin
Banana spider
Slows sodium channel inactivation
Phrixotoxin
Chilean fire
tarantula
Blocks potassium channels
Robustotoxin
Funnel web
spider
Opens sodium channels
Saxitoxin
Dinoflagellate
Blocks sodium channels
SNX-482
African Tarantula
Blocks calcium channels
Stichodactyla
Toxin
Sea Anemone
Blocks voltage-gated potassium
channels
Taicatoxin
Australian Taipan
snake
Inhibits voltage-gated calcium channels
Tetrodotoxin
(TTX)
Pufferfish
Blocks sodium channels
Tityustoxin-K
Brazilian
Scorpion
Blocks potassium channels
Versutoxin
Funnel web
spider
Opens sodium channels
17