5. Lionfish Venom Flashcards
lionfish in the atlantic ocean
lionfish are native to the pacific ocean –> ended up in the atlantic as an invasive species bc atlantic fish not used to eating it with its venom
describe the lionfish stinging mechanism
- 13 spines on the back
- venom glands within spines produce venom but the rest of the fish is not poisonous
- even just 1 spine or the skin brushing against the spines can inject venom
what is nociception?
pain sensation –> activation of the pain pathway
how can we identify which part of venom is responsible for activating pain pathway?
identify behaviour effects to find cellular targets and pathways
isolate + identify the toxin to understand toxin-receptor interactions and learn how predators are resistant
what is the pain measurement in mice?
inject some venom into feet of mice and record how long the mice lick their feet
why do we know that a protein component is responsible for the pain response from venom?
add trypsin or heat to denature proteins before injection –> lose pain response
why do we know that the protein component must have cysteine bridges?
add reducing agent before injection –> lose pain response
describe the transgenic mice used
pain sensing neurons express fluorescent Ca2+ sensor when Ca2+ is bound/channel is activated
does venom target peptidergic or non-peptidergic nociceptors?
non-peptidergic
describe the use of capsaicin
induces Ca2+ response and activates peptidergic pain receptors to cause fluorescence
2 results of fluorescent Ca2+ from applying venom
- Ca2+ fluorescence rapidly rises and slowly decreases
- Ca2+ fluorescence has a delayed increased and oscillates
why does venom cause oscillation of Ca2+ fluorescence?
When the channel is activated, ATP is activated to induce oscillatory response in secondary responders
what receptor does venom act on? data to support using pharmacological blockers
venom acts on P2X3
- Inject venom + saline = high pain response
- Inject venom + P2X3 blockers = reduced pain response
what is the second aim?
to identify the algogenic/pain-causing component of the venom –> i.e. the toxin
is the toxin EDTA-sensitive?
no, but other parts of the venom are
workflow for identifying the toxin
A. grind tissue and collect venom gland
- make cDNA library
- sequence to get transcriptome and predict proteome
B. collect whole venom
- fraction the venom
- MS
- predict proteome
C. compare the proteomes from the gland and whole venom
- find parts that are the same in both
D. identified 4 toxins that could activate P2X3 in high content screening –> observe pain response in animals
what is the 3rd aim?
characterize receptor target and evolved resistance mechanisms
why do venom-resistant mechanisms exist in animals?
animals are able to eat other venomous animals so they have mechanisms of resistance against the toxins
4 mechanisms of resistance
- target-site insensitivity
- toxin scavenging molecules
- off-target repurposing
- auto-resistance
what is target-site insensitivity?
mutation in receptor so it cannot bind toxin anymore
target-site insensitivity with P2X3
P2X3 binds ATP to give response but toxin can also induce this response –> mutate P2X3 so it can still bind ATP but not toxin
what are toxin scavenging molecules?
molecules that sequester the toxin so it cannot act on its target
what is off-target repurposing?
use a decoy receptor that is not associated with the toxin function but can bind the toxin
what is autoresistance?
resistance to toxin from their own species bc animals often cannibalize themselves
what animal can eat the lionfish? why?
moray eel
- their wall epithelium is so thick that the spikes cannot enter
describing cloning of venom receptors
cloned venom receptors from:
- Zig zag eel –> not in same environment as lionfish so no evolutionary pressure to modify receptor
- Honeycomb moray eel –> in same environment as lionfish so there is evolutionary pressure to modify receptor for resistance
- Lionfish
then tested each receptor’s sensitivity to lionfish venom
what is the goal of looking at various venom receptors?
find differences between human and eel receptors then swap domains to see if we can confer sensitivity in eel receptors or remove sensitivity in human receptors
