Biased Agonism Flashcards
Which membrane bound enzyme does the Gi-protein inhibit?
Adenylate cyclase -> reduction in cAMP and PKA
Which ion channels/enzyme does the Gi-alpha subunit modulate and what impact does this have on neuronal excitability
Inhibition of adenylate cyclase
Reduces cyclic AMP (cAMP) levels
Leading to decreased activation of protein kinase A (PKA).
This can result in reduced phosphorylation of ion channels, often leading to decreased neuronal excitability
Which ion channels/enzyme does the Gi-beta/gamma subunit modulate and what impact does this have on neuronal excitability
Inhibits the voltage gated Ca channel
○ decreasing calcium influx and neurotransmitter release
○ further dampening neuronal excitability
Activates the K+ channel
○ increasing potassium efflux and hyperpolarizing the neuron
○ which reduces excitability
What is the time frame for covalent modification of a receptor and translocation from the cell membrane?
Covalent modification
○ Phosphorylation -> seconds - minutes
Translocation
○ Reversible (internalisation/sequestration) -> minutes - hours
○ Irreversible (down regulation) -> hours
Explain how GPCR phosphorylation can initiate receptor internalisation and down-regulation including the roles of arrestin, clathrin and dynamin
A ligand binds to GPCR
GRK adds a phosphate group to the GPCR
Beta arrestin binds to the phosphates -> G protein cannot bind
Beta arrestin when bound becomes associated with clathrin coated pits -> can trigger internalisation by recruitment of other molecules
Dynamin dependent fission -> Mediates endocytosis
What are the 4 stages of the pain pathway
Transduction
Transmission
Modulation
Perception
Describe what happens in the pain pathway (transduction, transmission, modulation, perception)
- Transduction - Conversion of noxious external stimulus into an electronic action potential
- Transmission - Action potential transmitted
- Modulation - Body can turn the volume down on pain sensation if necessary
- Perception - Areas of the brain interpreting the action potentials as pain and also motor component (controlling response to pain)
Describe the GPCR pathways activated by the mu-opioid receptor highlighting the impact this has on neuro transmission
Opiate analgesic acts on mu-receptors
Activation of Gi protein -> substrates dissociate
Gb/y
○ Opening of K+ channels -> increased K+ efflux
§ Membrane hyperpolarisation
§ Reduced neuronal excitability
○ Inhibition of VG-Ca channels -> decreased Ca2+ influx
§ Reduced neurotransmitter release
Modulation of pain pathways -> analgesia
Explain the mechanism of action underlying an opioids analgesic effect at nociceptors
Decrease release of neurotransmitters e.g. glutamate
Explain the mechanism of action underlying an opioids analgesic effect at dorsal horn projection neurons
Decrease neuronal firing (pain signal coming in) and therefore intensity of signal sent to the brain
Explain the mechanism of action underlying an opioids analgesic effect at descending pathway at the level of the periaqueductal gray
Disinhibition of the descending pain pathway via blocking the release of GABA from interneurons
○ Decreasing the volume
Opioid receptors are present on GABA interneurons
○ Activates GPCR
○ Decreases Ca+ influx
○ Decreases GABA release
○ GABA receptor isn’t activated therefore the descending neuron from periaqueductal gray is active
○ Therefore pain pathway volume is turned down
What are the side effects experienced when taking an opioid with regards to the respiratory and gastrointestinal systems
Euphoria and sedation
○ Causes dopamine release
Respiratory depression
Constipation
Nausea and vomiting
How can opioids cause respiratory depression
Not well understood but mu receptors are expressed in respiratory centres of the brainstem and carotid bodies
Decreases sensitivity of chemoreceptors to hypercapnia (increased CO2)
Overall effect = reduction in breathing rate, reduced arterial O2 and increased CO2
Most common cause of death in acute opioid poisoning
Can be reversed with naloxone, a mu-receptor antagonist
How can opioids cause constipation
Mu receptors in the enteric system reduce bowel tone and contraction
Most common side effect of chronic opioid treatment (40-95%)
No tolerance
What can opioid related constipation lead to
Faecal impaction
Bowel obstruction
Bowel perforation
Noncompliance with opioid analgesics
Poor quality of life
How can opioids cause nausea/vomiting
The site of action is the area postrema (chemoreceptor trigger zone), a region of the medulla in the brain
Reduced GI motility and secretion
Explain how receptor desensitisation and down-regulation can lead to opioid tolerance
Down regulation and desensitisation can lead to tolerance to drug action
Tolerance means that a higher dose of opioid is required to achieve the same degree of analgesic effect (dose escalation)
Tolerance occurs in people taking long-term opioids
Suggest 2 ways to improve analgesia after tolerance has occurred
Increase dose - but potential for increased side effects
Opioid rotation - switching from one opioid to another
○ Mechanism unknown but cross-tolerance from one opioid to another may be incomplete
○ The equipotent analagesic dose for the new opioid may be decreased by up to 40%
What is biased agonism
A selective activation of cellular pathways
Suggested as the predominant mechanism of GPCR mediated cell signalling
Explain the key differences between classical ‘on-off’ model of GPCR mediated cell signalling and biased cell signalling with regards to G-protein and beta arrestin activation
Unbiased ligands would be expected to have equal levels of efficacy for beta-arrestin and G-protein mediated pathways
Beta-arrestin pathway = side effects
Biased cell signalling would activate one pathway more often than the other
Explain the difference in activation of the G-protein pathway and beta-arrestin pathway between an unbiased agonist such as morphine and a biased agonist such as TRV-130
Unbiased agonist/classic opioid e.g. morphine
○ Activates G-protein and GRK/Beta-arrestin pathways equally
Biased agonist e.g. TRV-130
○ Activated G-protein pathway to a greater extent than the GRK/Beta-arrestin pathway (side effects pathway)
How and why do the analgesic and side effect profile of morphine and TRV-130 differ
Morphine activates both the G-protein pathway (analgesia) and the beta-arrestin pathway (side effects) equally
TRV-130 favours the G-protein pathway therefore the side effect profile is much less severe as it is being activated less frequently
Explain how internalisation assay was used to test whether TRV-130 was a safer therapeutic option for acute pain compared to morphine
Tested: activation of cell signalling pathways
Explain how hot plate studies was used to test whether TRV-130 was a safer therapeutic option for acute pain compared to morphine
Tested: analgesic property of the drug
Animals placed individually on a heated surface and the time interval (seconds) between placement and a shaking, licking or tucking of the hind paw was recorded
