Lecture 5 & 6 - Neurotransmitter Receptors Flashcards
What are 3 functions of NTs? Explain each and provide examples. Which function is the fastest?
- Neural signaling: mediate comms between neurons = AA NTs (faster than the 2 others)
- Trans-system modulators: modulate large populations of target neurons in multiple systems = biogenic amines
- Within-systems modulators: modulate info by neurons within systems (eg: within the basal ganglia system) = neuropeptides
What does allosteric modulation mean? Example?
Alteration of receptor activity (of binding site conformation) at a site distinct from the NT binding site
Eg: Benzodiazepines on GABAa receptors
Can a ligand be an agonist or antagonist?
YUP
How many subunits in an ionotropic NT receptor? Describe each.
5 subunits with each 4 transmembrane domains (20 crossing overs of the lipid bilayer)
How many subunits in a metabotropic NT receptor? Describe them.
2 subunits with 7 transmembrane domains each
What are the 2 types of ionotropic NT receptors based on their structure?
- Homomeric
2. Hetereomeric
Which are fastest: ionotropic or metabotropic NT receptors?
Ionotropic
What determines whether a particular ion will be able to go through an ion channel? What is this known has?
How that ion interacts with water = the ion’s sphere of hydration
What ion channel is activated by ATP?
P2X4
What is particular about glutamate ionotropic receptors?
They have 4-fold symmetry with a complex extracellular domain that translates the binding of glutamate to the opening of the channel and have evolved differently and during a different period of time than a lot of other ionotropic receptors
What % of clinical drugs act on GPCRs?
50%
What are orphan receptors? How many of them? Describe them.
Large component of the GPCR super family: greater than 100 orphan receptors = we do not know their ligand
What are the 6 components of the GPCR superfamily of receptors?
- Glutamate
- Frizzled/TAS2
- Rhodopsin
- Adhesion
- Secretin
- Orphan
Can G-proteins be stimulatory or inhibitory?
YUP
Which have more diverse postsynaptic effects: ionotropic or metabotropic receptors?
Metabotropic
Time frame of ionotropic synaptic transmission?
Milliseconds
Time frame of metabotropic synaptic transmission/effects?
Seconds to hours
What are 2 ions that a lot of ionotropic receptors are permeable to and cause hyperpolarization?
- Cl-
2. HCO3-
What are 2 ions that a lot of ionotropic receptors are permeable to and cause depolarization?
- Na+
2. Ca++
Describe the binding of NTs to receptors.
High degree of specificity
What determines whether the effect of an NT binding to a receptor will be excitatory or inhibitory?
Selectivity of the ion channel
What determines the length of the ion channel opening upon binding of the NT to the ionotropic receptor? What does this determine?
The kinetics of NT binding (for how long the NT binds)
This determines the duration of the effect
Can a single NT activate more than 1 receptor?
YUP
Can a single NT activate both ionotropic and metabotropic receptors?
YUP, most small ones do
What property describes the degree of specificity of an NT to a receptor?
The dissociation constant, Kd
What are the 5 ionotropic NT receptors?
- Cys-loop nicotinic ACh receptors (nAChRs)
- Serotonin type 3 receptors (5-HT3R)
- GABAaR and GABAcR
- Glycine receptors (GlyR)
- Glutamate ionotropic receptors: NMDA, AMPA, and Kainate
What are the 4 general features of ionotropic NT receptors?
- Pseudo symmetrical arrangement around a central ion-conducting pore
- Non-selective
- Ligand-binding sites at the interface of the subunits (between subunits)
- Exist in 3 states
What are the 3 possible states of ionotropic NT receptors? Describe each
- Resting: unliganded/closed
- Activated: liganded/open
- Desensitized: liganded/closed
What does the transition of the ionotropic receptor from the resting to the activated state dependent on?
Dose of agonist
When is the ionotropic NT receptor in the desensitized state?
When there is too much agonist
What is another name for ionotropic NT receptors?
Pentameric ligand-gated ion channels (pLGICs)
What does the anti-nausea drug ondansetron act on?
Binds to 5-HT3Rs and blocks it
Where are a lot of GlyRs found?
Spinal cord
How fast after ACh binding is the nAChR activated?
Instantaneously
How fast do neuronal nAChRs desensitize? What does this explain?
Rapidly (msecs-secs)
Explains nicotine addiction
Are all neuronal nAChRs the same? Why/Why not? Explain.
No, they are structurally diverse because multiple genes coding for these so different channels have unique properties, like:
- Desensitization rates
- Ca++ permeabilities
At what ratio does ACh bind its receptor?
2 ACh molecules : 5 molecules of receptor (pentameric)
Describe how ion current varies throughout receptor desensitization.
First the current is strong and reaches the peak current, and then decreases little by little
To what can we compare NT receptor desensitization? What is different?
Na+ VG channel inactivation
Desensitization is slower
Which causes stronger desensitization of AChRs: nicotine or cytisine? What does this explain?
Nicotine (explains addiction and abuse )
What ions go through nAChRs?
Na+/K+ mainly and some Ca++
What ions go through AMPA receptors?
Na+ and K+
What ions go through NMDA receptors?
Na+, K+, and Ca++
What ions go through Kainate receptors?
Na+
What ions go through 5-HT3 receptors?
Na+
What ions go through GABAa receptors?
Cl-
What ions go through Glycine receptors?
Cl-
What is a strong antagonist of GlyRs?
Strychnine poison
Why are nAChRs called Cys loop receptors?
Because they contain a disulfide bond between 2 Cys
What are the 2 families of ionotropic receptors based on? What does each contain?
Based on structural differences:
Family 1: nAChRs, GABAaRs, 5HT3Rs, GlyRs = N-terminal and C-terminal are both extracellular and 5 subunits each containing 4 transmembrane helices
Family 2: AMPA, NMDA, Kainate = N-terminal extracellular and C-terminal cytosolic and 4 OR 5 subunits each containing 3 transmembrane helices (completely transverse) plus 1 incomplete pore loop
What is the purpose of the cytosolic C-terminal of ionotropic glutamate receptors?
Contain sites for phosphorylation and binding of intracellular proteins
Why are NMDA receptors also permeable to Ca++ and the other glutamate receptors are not?
They have a voltage dependent Mg++ block
What are the 2 types of neuronal nAChRs?
- Pentahomomeric receptor
2. Pentaheteromeric receptor
Why don’t our muscles contract when we smoke a cigarette?
Because muscle nAChRs are different from neuronal ones and nicotine does not bind as well to these
What are the 4 genes that encode AMPA receptor subunits?
- Glu A1
- Glu A2
- Glu A3
- Glu A4
What is the ratio of NR1 and NR2 NMDA receptor subunits?
2:2
What are the 7 genes encoding for NMDA receptor subunits? What are the 2 main ones?
- NR1
- NR2A***
- NR2B***
- NR2C
- NR2D
- NR3A
- NR3B
What are the 5 genes encoding for Kainate receptor subunits?
- Glu R5
- Glu R6
- Glu R7
- KA1
- KA2
What are the 6 genes encoding for GABA receptor subunits? How do these mix?
- Alpha 1-7
- Beta 1-4
- Gamma 1-4
- Delta
- Epsilon
- Rho
Mix in a very heterogenous fashion to produce many different types of GABAa receptors
How many GABA molecules need to bind to the GABAa receptor to open the ion channel?
2
What is picrotoxin?
GABAa receptor antagonist that induces seizures by blocking the ion channel
How do benzodiazepines and barbituates affect the GABAa receptor?
They increase the flux of Cl- through the GABAa receptor channel in the presence of GABA
What drugs can be used against epilepsy?
Barbituates
How do steroids affect GABAa receptors?
They have the same interaction as benzodiazepines and other allosteric modulators
Why are glutamate ionotropic receptors not very good drug targets?
They are expressed by all neurons so there would be a very wide range of side effects
What are the 4 pathological processes in which glutamate is involved?
- Epilepsy
- Anxiety
- Addiction
- Ischemic brain damage
What are the 3 physiological processes in which glutamate is involved?
- Neuroplasticity
- Neuronal development
- Learning and memory
What are some mutations in the NR2A and B involved with?
Below 100 IQs
What 2 areas of the brain are associated with addiction?
- Basal ganglia
2. Nucleus accumben
Explain how glutamate is involved in ischemic brain damage.
NMDA receptors have a tendency to become over active when they lack O2 leading to Ca++ influx which will lead to further damage of brain cells (eg: apoptosis)
How could brain damage in stroke victims be prevented?
Given NMDA receptor blockers during the acute phase of the stroke
What type of ion channels are NMDA receptors?
Both ligand and voltage gated
Describe the kinetics of the AMPA receptor.
FAST
Describe the kinetics of the NMDA receptor.
SLOW
What co-agonist is required for NMDA receptor activation?
Glycine
How is the Mg++ block of NMDA receptors removed?
Initial depolarization to -30 mV
How fast do AMPA and Kainate receptors desensitize? What is the mechanism?
Rapidly
Disruption of agonist binding domain dimer assembly
How fast do NMDA receptors desensitize?
Slowly
Are AMPA receptors permeable to Ca++?
Just a little
Why is the NMDA receptor classified as a glutamate receptor and not glycine?
Because ambient levels of glycine usually saturate the binding sites on NMDA receptors under normal circumstances
For what 2 conditions in the NMDA receptor used as a pharmacological target?
- Schizophrenia
2. Acute effects of ischemic event
What is the 4th very rare type of ionotropic GluR?
DeltaRs
What do Glu NR3A and B form when joined? How frequent are these?
Cl- ion channels, meaning they are inhibitory
Rare
What are the 4 pathological processes in which NMDA receptors are involved?
- Stroke
- Seizures
- Traumatic CNS injury
- Neurodegenerative disorders
What is a hypothesis for the cause of Huntington’s disease?
Too many NMDA receptors in the basal ganglia leading to the destruction of cells
Describe the structure of a G protein and how it interacts with NT receptors.
Lipid modification on the alpha subunit and the beta-gamma dimer to hold it in close proximity to the NT receptor
Describe the basal state of a G protein.
GDP bound to alpha subunit
Describe the size of the subunits of the G protein.
Alpha subunit is much larger than the beta and gamma together
Describe the 4 steps of operation of G proteins. Timeframe?
- Inactive NT receptor does not interact with G protein
- NT binding = G protein association and GDP exchange for GTP on alpha subunit
- Activated G protein splits and both alpha and beta-gamma subunits activate effector proteins
- Alpha subunit removes phosphate from GTP to terminate its own activity
Milliseconds
Can G proteins directly activate ion channels?
Yes
What is an example of a G protein directly activating an ion channel?
mAChR binding of ACh => G-protein activates K+ channel to hyperpolarize
What can G proteins do once activated?
- Directly activate ion channels
- Activate membrane bound enzyme
- Activate second messenger cascades
Effect of G alpha s activation?
Increase in intracellular cAMP
Effect of G alpha i activation?
Decrease in intracellular cAMP
What are 6 second messengers that can be activated/inhibited by G proteins?
- Ca++
- cAMP
- cGMP
- IP3
- Diacylglycerol
- Nitric oxide
What is the function of the second messenger IP3?
Ca++ release from ER
What is the function of the second messenger DAG?
PKC activation
What is the function of the second messenger NO?
Guanylyl cyclase activation
What is the function of the second messenger cAMP?
PKA activation
What is the function of the second messenger cGMP?
PKG activation
Effect of G alpha q activation?
PLC => DAG + IP3
Norepi on alpha adrenergic 1-2 receptors: what G protein activated?
G alpha i
Glutamate on mGluR: what G protein activated?
G alpha q
Dopamine on D1 and D2 receptors: what G protein activated for each?
D1: G alpha s
D2: G alpha i
What determine the output of a neuron?
Concentration and timing of NTs binding at its receptors
ACh on muscarinic receptors: what G protein activated?
G alpha q
GABA on GABAb: what G protein activated?
G alpha s OR
G alpha q
Histamine on H1-3 receptors: what G protein activated?
G alpha s OR
G alpha q
Serotonin on 5-HT1-2 and 4-7: what G protein activated?
G alpha s
Norepi on beta-adrenergic 1-3 receptors: what G protein activated?
G alpha s
What is an exogenous agonist to beta receptors? What is it used for?
Isoproterenol
Increase heart contractility
What is an exogenous antagonist to beta receptors?
Propanolol
What is an exogenous antagonist to alpha 2 receptors?
Phentolamine
What do a lot of the names of beta adrenergic receptor antagonists have in common?
End in -olol
What 2 types of adrenergic receptors do we have in the brain?
- Alpha2
2. Beta1
What are the 8 metabotropic receptor subtypes?
- Glutamate
- GABAb
- Dopamine
- NE and EPI (adrenergic)
- Histamine
- Serotonin
- Purines
- Muscarinic
What are the 3 classes of glutamate metabotropic receptors? Which G protein does each activate?
- Class I: G alpha q
- Class II: G alpha i
- Class III: G alpha q, i , and o
What NTs bind purine metabotropic receptors?
ATP and GTP
What drugs were given in the early treatment of Parkinson’s disease? Why?
Anti-muscarinic receptor drugs because patients have elevated levels of ACh in basal ganglia
What 2 receptors are working against each other in the basal ganglia?
Metabotropic dopamine and muscarinic receptors
Can metabotropic receptor subtypes for a single NT activate different G proteins? Examples?
YUP
Eg: glutamate metabotropic receptors
Describe the concept of amplification with metabotropic NT receptors.
The activation of a single receptor with a single NT bound can result in the activation of many G proteins (same ones)
What are the 2 MAIN types of voltage-gated calcium channels? Describe each.
- L-type (long-lasting): less sensitive, open slower, and remain open longer
- T-type (transient): more sensitive, open faster, and remain open for shorter periods of time
Which VG calcium channels are high voltage gated?
- L type
- P/Q type
- N type
- R type
Which VG calcium channels are low voltage gated?
T type
Which VG calcium channels are expressed in the presynaptic terminal?
- P/Q type
- N type
- R type
Which VG calcium channel is expressed in the postsynaptic terminal?
L-type
Can VG calcium channels inactivate?
Yes
What is EGTA? What does its effect demonstrate?
A molecule that chelates calcium and therefore inhibits inactivation of VG calcium channels because the inactivation is calcium-dependent
How would an extracellular environment rich in Ba++ instead of Ca++ affect a calcium VG channel?
Ba++ would flow in the cell but Inactivation of the channel would be inhibited because the inactivation is calcium-dependent
What is a method to test the functionality of different VG channel genes? 6 steps
- Dissect superior cervical ganglion near carotid bifurcation in rats
- Enzymatically dissociate and isolate the sympathetic neurons and place them on poly-lysine coated culture dish
- Intranuclear injection of cDNA plasmids encoding the protein of interest
- Wait 12-24 hours
- Use fluorescent die to determine which neurons have successfully expressed the gene
- Voltage clamp experiment on cells that express the gene and cells that didn’t
What is voltage-dependent VG calcium channel inhibition? How can this be overcome?
The beta-gamma subunit of the G-protein binding the N type VG calcium channel will inhibit the channel when voltage is applied by having charge-charge interactions with the VG channel
A very large voltage (+100mV) will disrupt the interaction and the dimer will dissociate
Does voltage-dependent VG calcium channel inhibition happen pre or post synaptically?
Both
How can metabotropic receptors produce long-term changes?
By regulating gene expression
What other neuronal receptors are involved in long term effects?
Tyrosine kinase receptors
What is the orthosteric binding site?
The site where the NT binds
What is cystisine?
nAChR agonist
How many serotonin receptors? Which is the only ionotropic one?
7
5HT-3R
