Week 3- Glutamate Flashcards
what is glutamate? how is glutamate synthesises and where?
-its in all cells and its an amino acid • Glutamate can be synthesised in the pre-synaptic neuronal terminal • from glucose (via TCA cycle) • from glutamine (by action of glutaminase)
there are 2 types of glutamate receptors what are they?
-Metabotropic glutamate receptors (mGluRs)
• GPCRs
-Ionotropic glutamate receptors
• Ligand-gated ion channels
• 3 classes of receptor called NMDA receptors, AMPA receptors and kainate
receptors
how are ionotropic glutamate receptors separated?
• Three subtypes are separated on
responsiveness to synthetic analogues
what activates ionotropic glutamate receptors?
• All activated by L-glutamate
what are the properities and structure of the ionotropic glutamtae receptors?
• Different pharmacological properties
• Different biophysical properties and
functional effects in neurons
• All have similar structures; tetramers
what are the main facts about NMDA receptors? sub units, shape
- Assembled from 7 potential subunits encoded by 7 different genes
- GluN1, GluN2A, GluN2B, GluN2C, GluN2D, GluN3A, GluN3B
- The receptor is a tetrameric complex
- Hetero-tetramers – mixture of subunits
- Typically 2 GluN1 and 2 GluN2 subunits come together
- Alternative splicing can affect the GluN1 gene (8 variants identified)
what is the NMDA receptors structure like?
• Each subunit of the tetramer has: • Extracellular N-terminal domain labelled ATD • An extracellular ligand-binding domain (LBD) - binds the agonist • Three transmembrane spanning α-helical domains (M1, M3, M4) and • One re-entrant P-loop called M2 • An intracellular C-terminus (CTD)
the subunits that make up then NMDA receptor affect the properities? which properties?
• Agonist and coagonist potency • Deactivation rate • Magnesium • Ion permeation • Channel conductance • GluN2 subunits affect pharmacology of NMDAR
how many drug binding sites are there for NMDA receptors?
5
what are the 5 different drug binding sites for NMDA receptors?
- Glutamate (agonist site)
- Glycine site
- Polyamine binding site
- Mg2+ site
- Channel blocking site
how is the NMDA receptor activated at the glutamate binding site?
-glutamate is the ligand
• Each tetramer binds two molecules of glutamate
• EC50 value for glutamate is 0.5-3.3 µM (dictated by GluN2 subunit)
• NMDA is a synthetic agonist that will act at the same binding site
-for full activation a co-agonist is needed
• 2 molecules of glycine are required (co-agonist) - EC50 value 1 µM
• D-serine, D-alanine can also act as a co-agonist
• GluN1 is the glycine-binding subunit
what are the 5 different drug binding sites for NMDA receptors?
- Glutamate (agonist site)
- Glycine site
- Polyamine binding site
- Mg2+ site
- Channel blocking site
what are some competitive antagonists at the glutamate site?
- drug called D-AP5
Limited drug development due to conservation of glutamate binding site
between NMDAR, AMPAR, KainateR
how is the NMDA receptor activated at the magnesium binding site?
• Mg2+ site • Channel is blocked by Mg2+ at resting neuronal membrane potentials • This feature gives NMDA receptor voltage dependent channel block -prevent ions moving down channel -ion is blocked at resting ion potential whihc is -60-70mV and is membrane dependant blocked as membrane becomes depolarised it removes the block
how is the NMDA receptor activated at the magnesium binding site?
• Mg2+ site • Channel is blocked by Mg2+ at resting neuronal membrane potentials • This feature gives NMDA receptor voltage dependent channel block -prevent ions moving down channel -ion is blocked at resting ion potential whihc is -60-70mV and is membrane dependant blocked as membrane becomes depolarised it removes the block
how is the NMDA receptor activated at the drug binding site? type of drugs?
-Channel-blocking drugs
-relies on the fact that the channel is open and can fit in to block it
• ketamine - a dissociative anaesthetic and analgesic, drug of abuse
• phencyclidine – a psychomimetic, drug of abuse
• MK-801 (dizocilpine)
-Developed for treatment of epilepsy
• memantine – binds to M2 domain deep inside channel pore
-Alzheimers disease; neuroprotection
17
-Neurosteroids
Can be positive or negative allosteric modulators
e.g. pregnenolone sulfate (PAM at GluN2A receptors)
how do signals travel across a synapse
-electrical signal is converted into chemical signal to cross the gab and then convert back in post synapic neurone
what is a excitatory postsynaptic potential (EPSP)?
-time delay due to depolarisation
when the membrane is at resting potential of -70mV and then goes up to threshold but not yet reaching threshold
what are the different types of signals a post synaptic cell can receive?
inhibitory or excitatory
where do the 2 synapses contact the neurone as they are different locations?
type 1 is at the spine
type 2 at the cell body and dendritic shaft
where and how are receptors clustered?
• Receptors are clustered by lots of regulatory proteins into areas called
a “postsynaptic density”
• A typical PSD is 350nm in diameter and can contain 20 NMDAR (blue)
and 10-50 AMPAR (green)
what is PSD also a hold of?
shank and homer which are responsible for localising other membrane receptors
what do NMDAR and AMPAR RESPond to?
- both present in postsynaptic memebrane
- can respond to glutamate being released into the synaptic cleft
what is an EPSC?
-excitatory postsynaptic current
-it is what can be measured to find out the which channel are active and the response
-Each type of glutamate receptor contributes to
different phases of the EPSC
• NMDAR unique in high Ca2+ entry
• Translates electrical signal into biochemical signal
what are the key facts about LTP?
- In normal transmissions with infrequent synapse activity glutamate mainly activates the AMPA receptor
- the nmda WILL mostly be blocked by Magnesium so the glutamate will mostly activate the AMPA cause depolarisation and excitation
- after conditioning train WHERE IT MIMICS FREQUENT synapse activity
- so they keep firing and there maybe more glutamate released into the synapse so more receptors can get activated, there will be sufiienct delpolarisation of the post synaptic membrane mediated by AMPA receptors which allow the unblocking of magnisum on the NMDA recptors allowing it to be activated by glutamate
what are the key facts about LTP?
- In normal transmissions with infrequent synapse activity glutamate mainly activates the AMPA receptor
- the nmda WILL mostly be blocked by Magnesium so the glutamate will mostly activate the AMPA cause depolarisation and excitation
-after conditioning train WHERE IT MIMICS FREQUENT synapse activity
how does an LTP work?
• Postsynaptic AMPA receptors are activated NMDAR requires a sustained depolarisation (mediated by AMPAR) to relieve Mg2+ block - allow channel opening and Ca2+ entry • Ca2+ entry can activate other enzymes including kinases (PKC, CaMKII) which phosphorylate AMPAR increasing responses • Phosphorylation increases AMPAR conductance and can increase expression
how does excitotoxicity work?
• High Ca2+ permeability of NMDAR can cause problems
• High glutamate concentrations Ca2+ overload toxicity
• Glutamate excitotoxicity can cause neuronal cell death
• Attempts to block NMDAR to reduce toxicity in situations such as
ischaemic stroke (excessive glutamate release)
• Many other receptors contribute to excitotoxicity
• Thought to be a major factor in ischaemic brain damage and in
neurodegenerative diseases
what are the 4 genes that encode for different subunits of AMPA recptors?
GluA1, GluA2, GluA3, GluA4
-can be a homo or hetra tetrama to make the ion channel
what are the 5 genes that encode for different subunits of kainate recptors?
GluK1, GluK2, GluK3,
GluK4, GluK5
-
what are the 5 genes that encode for different subunits of kainate recptors?
GluK1, GluK2, GluK3,
GluK4, GluK5
what ion is the AMPA receptors impermeable for?
calcium
but permeable to sodium and potassium
-calcium selectivity is due to amino acid in the M2 domain
-RNA editing changes single amino acid to arginine causing impermeability
what are AMPA and kainate receptors important for? distribution and expression
-excitatory transmissions
Also important ion channels in the brain
• AMPA receptors mediate fast excitatory transmission
• Typically AMPAR are co-expressed with NMDAR
• Kainate receptors are more limited in distribution
what are AMPA and kainate receptors important for?
Also important ion channels in the brain
• AMPA receptors mediate fast excitatory transmission
• Typically AMPAR are co-expressed with NMDAR
• Kainate receptors are more limited in distribution
what are the other type of glutamate receptors? properties?
-Metabotropic glutamate receptors
• GPCRs (7TM domains) belonging to Class C family
• Large extracellular N-terminal domain where glutamate
binds
• 8 subtypes – mGluR1 to mGluR8
-the ligand bindingisnt with the transmembrane like other GPCRS its on the outside
what are the other type of glutamate receptors? properties?
• GPCRs (7TM domains) belonging to Class C family
• Large extracellular N-terminal domain where glutamate
binds
• 8 subtypes – mGluR1 to mGluR8
what are the properties of group 1?
- Includes mGluR1 and mGluR5
- post-synaptic (excitatory)
- Gαq/11 linked – increase Ca2+
what are the properties of group 2 and 3?
- Group 2 - mGluR2, mGluR3,
- Group 3 - mGluR4, mGluR6, mGluR7, mGluR8
- pre-synaptic location – neuromodulation (inhibitory)
- Gαi linked - decrease cAMP
what is the pharmacology of mGluR?
• Drugs that distinguish between subtypes have been developed • Antagonists for Group I mGluR – hold potential for epilepsy, pain, Parkinson’s disease, neuroprotection • Antagonists for Group 2 mGluR – potential as cognition enhancers • Positive allosteric modulators for Group 3 mGluR – potential for Parkinson’s disease and anxiety
