GABA neurotransmission Flashcards
Name 2 fast inhibitory neurotransmitters in the brain and list their receptors
• GABA GABAA, GABAB • Glycine GlyR o No metabotropic Gly receptor known
GABA(A)R pharmacology - agonists:
- GABA
- Muscimol from Amanita muscaria
- Gabadoxadol
GABA(A)R pharmacology - antagonists:
• Bicuculline (competitive)
• GABAzine (competitive)
• Picrotoxin (non-competitive; combination of picrotoxinin and picrotin)
U93631
GABA(A)R pharmacology - modulators:
- Benzodiazepines (BZDs)
- Barbiturates
- Neurosteroids
- Anaesthetics
- Alcohol
Where are GABA(A)Rs found?
Not on dendritic spines - usually found in the membrane of the postsynaptic neuron. They signal in the normal mechanism for an ionotropic receptor
Explain feed-forward inhibition, using a diagram
• Excitatory neuron stimulates GABAergic and Glutamatergic neurons –> EPSP, then IPSP (caused by GABA release)
diagram: https://neurology.mhmedical.com/data/books/1049/kan_ch2_f008.png
Using a diagram, explain feedback (recurrent) inhibition
• 2nd excitatory cells feeds back onto itself
• Is ‘doubly inhibited’
• There are 3 synapses; 2 of them inhibitory
Diagram: https://neurology.mhmedical.com/data/books/1049/kan_ch2_f008.png
Using a diagram, explain inhibition of inhibition and what it leads to
–> excitation (overall)
GABA neuron inhibits GABA neuron – cancels out release of GABA from 2nd neuron; prevents inhibition ∴ allows excitation
Diagram: https://neurology.mhmedical.com/data/books/1049/kan_ch2_f008.png
Explain why inhibitions are needed
GABA antagonists induce seizures Epilepsy in rats Give PTZ --> seizures ∴ GABA agonists can be used as anticonvulsants Zebrafish assay • Tetanus toxin --> lockjaw/tetanus o Without inactivation with formaldehyde --> muscle spasm o Stops inhibitory synapses from working
Explain how strychnine achieves its inhibitory effects and their consequences:
Antagonises GlyRs Alkaloid Comes from Quaker’s buttons Inhibits synapses of spinal inhibition of motor neurons --> spasms Renshaw cell: Glycinergic Sits in spinal cord α muscle neurons excite these; Renshaw cells feed back and inhibit motor neurons (FF inhibition via Gly)
With the aid of a diagram, explain Glycine receptor structure and subunit composition
‘Cys-loop’ family members 4 TMDs long intracellular loop connects TM3 and 4 either: 5 α subunits Or 2α + 3β
diagram: https://www.researchgate.net/figure/Structure-of-the-glycine-receptor-A-Membrane-topology-of-the-a-subunit-with-the-four_fig1_11904296
List the features of a glycine receptor
Ligand-gated ion channel 5 subunit genes: α1-4 No α4 in humans β1 Large single channel conductance – 50-110pS Anion pore; selects for Cl- ions Activated by Glycine (β-alanine + taurine) Potently inhibited by: Strychnine RU5135 Pore can be blocked by picrotoxinin, as with GABAAR
Explain GABA structure (chirality), synthesis, transport, and receptors
• GABA has no chiral centre – no L/D form
• Not usually on dendritic spines
• Synthesized by GAD67 (in cell) and GAD65 (nerve terminal)
o GAD = Glu decarboxylase
• GAT1 and 3 transport GABA into cells
• VGAT gets GABA (and Gly) into vesicles
• GABAA,B,C receptors bind GABA
• GABA function terminated by diffusion and uptake by GATs
With the aid of a diagram, explain GABA(A)R structure and potential subunit composition, as well as the 3 most common conformations
Ligand-gated ion channel assembly Channel is inherent to receptor Ligand-binding enables channel opening Pentameric - α,β,γ 19 subunits: α1-6 β1-3 γ1-3 δ ϵ π θ ρ1-3 Most receptors have 2α, 2β, 1γ/δ subunit Other conformations include: α1β2γ2, α1βγ2 different subunits = different localisations – α6 is only in the cerebellum
• In the transmembrane domain the terms “plus side”, which is formed by parts of TM2 and TM3 of the principal subunit; and “minus side”, formed by parts of TM2 and TM1 of the complementary subunit, are also commonly used
Diagram: https://upload.wikimedia.org/wikipedia/commons/3/3a/GABAa_receptor.gif
Use a diagram to show where on the GABA(A)R benzodiazepines bind, and explain their potential effects
Bind α-γ subunit interface
Don’t work on α4,6-containing receptors
Classically, are PAMs of GABA-mediate channel activation:
They increase frequency of bursts of channel opening
Are anticonvulsants; promote sleep; reduce anxiety
Can be very addictive
Temazepam sleeping pills
Drug users tend to prefer short-acting and intermediate-acting barbiturates. The most commonly used are amobarbital (Amytal), pentobarbital (Nembutal), and secobarbital (Seconal). A combination of amobarbital and secobarbital (called Tuinal) is also highly used. Short-acting and intermediate-acting barbiturates are usually prescribed as sedatives and sleeping pills. These pills begin acting fifteen to forty minutes after they are swallowed, and their effects last from five to six hours.
Are also BDZ-site inverse agonists
Anxiogenic
Seizure-promoting
E.g. DCM
And competitive antagonists:
Flumazenil
Treats overdose
Some BDZ-site ligands have subtype selectivity:
Can lead to more focused therapeutic use with fewer side-effects
α1,5 –> sedation (ataxia, amnesia)
α2,3 –> anxiolytic (stop anxiety)
Barbiturates and (neuro)steroids
Different binding site to BDZs:
PAMs at [low]
Activate receptors directly at [higher]
Tend to increase duration of bursts of channel opening
Sodium thiopental/pentobarbital – used in lethal injections (as with midazolam)
Briefly summarise the structure and pharmacology of GABA(B) and GABA(C) receptors
GABAB:
GPCR
Signal via intracellular G-proteins to 2nd messenger cascades
Obligate dimer
GABAC:
AKA GABAA-ρ
Insensitive to bicuculline and GABAzine (unlike GABAA)
Structurally similar to GABAA – Cys-loop pentameric Cl- channel
How many types of GABAergic cells are there?
• There are many kinds of GABAergic cell types
o 3 types of pyramidal cell are accompanied by at least 21 classes of interneuron in hippocampal CA1 area
• Basket cells mediate FF and FB inhibition
o Depends on which branch of the basket cell is being activated
Explain the immediate potentiating effect of GABA(A)R-mediated synaptic transmission, how this can be blocked and enhanced, and why this has no effect on the long, slow portion.
GABAAR-mediated synaptic transmission is the most common form of GABAergic synaptic signalling
–>IPSP; short-lived. Completely blocked by bicuculline, GABAzine, picrotoxin. Enhanced by BDZs, barbiturates, neurosteroids
Some GABAergic IPSPs are longer, and biphasic
Blocking GABAARs does not block the long, slow portion of the IPSP
GABABRs mediate this long slow IPSP via G-protein linked Inwardly Rectifying K+ (GIRK) channels
Initially identified by pharmacology – a receptor which was activated by GABA, not blocked by GABAAR antagonists and activated by a selective antagonist:
Baclofen:
Hyperpolarises postsynaptic cells via increased K+ conductance (∴ causes IPSP)
Reduces NT release from presynaptic terminals (via inhibition of VDCCs)
Reduces production of cAMP by adenylyl cyclase
∴ coactivation of GABAARs and GABABRs causes long-lasting, biphasic IPSPs
With the aid of a diagram, describe GABA(B)R structure and the effect of knocking out the subtypes
• Formed from 2 interacting proteins
o Both proteins have ‘classic’ GPCR structure
7TMDs
o GABA binds GABABR1; G-proteins interact with GABABR2
o 2 N-terminal splice variants of GABABR1
o KO of either Receptor subtype blocks all GABAB-type responses
diagram: https://www.google.com/url?sa=i&url=https%3A%2F%2Fwww.semanticscholar.org%2Fpaper%2FTargeting-GABAB-receptors-for-anti-abuse-drug-Phillips-Reed%2Fa049643bb25c8631e0267a40182e6d310d1f31fb%2Ffigure%2F0&psig=AOvVaw3ajPOqHuT8aaF5_1c8tkgU&ust=1577221072651000&source=images&cd=vfe&ved=0CAIQjRxqFwoTCIDWj8DUzOYCFQAAAAAdAAAAABAO
List 2 GABA(B)R agonists
• Agonists:
o GABA
o Baclofen
o SKF-97541
List 2 GABA(B)R antagonists
• Antagonists:
o Saclofen
o CGP-55845
o SCH-50911
List 2 GABA(B)R positive allosteric modulators
o Rac BHFF
o GS-39783
Name a GABA(B)R negative allosteric modulator
o CLH304a
Which signalling pathway are GABA(B)Rs linked to? What downstream effects does this have?
Are Gi/o linked, ∴ GABABRs:
Inhibit cAMP production
Activate GIRK K+ channels –> long-lasting IPSP
Inhibit VDCCs
Where are GABA(B)Rs found on a synapse? What are the subsequent effects?
• Are found pre- and post-synaptically
o Presynaptic GABABRs can inhibit VDCCs on the presynaptic neuron (autoreceptor), inhibiting further GABA release from the presynaptic neuron
o Presynaptic GABABRs can depress Glu release
What effect can GABA(B)Rs have on CA1 pyramidal cells?
GABABRs can mediate short-term plasticity in CA1 pyramidal cells:
If 2 identical stimuli are given in rapid succession (30-1000ms of each other), the 2nd IPSP is smaller
This is because the first GABA ‘wave’ inhibits the cell’s VDCCs ∴ i[Ca2+] is decreased
∴ Ca2+ less available; so can’t cause cytoskeletal rearrangement and vesicle release
∴ less GABA is able to be released, so the second IPSP is smaller
GABABR antagonists can block this!
What do neurogliaform cells produce?
• Neurogliaform cells (NGFs) produce GABAB-mediated IPSPs
o GABAB-mediated synaptic responses normally come from these
• Applying a GABAAR antagonist blocks the short, fast portion of the biphasic response
o Leaves the long, slow portion – GIRK-mediated
o Shown with paired cell recordings (recording pre- and post-synaptic cells at the same time)
• NGFs can affect other NGFs in this manner too! (inhibition of inhibition)
• NGFs utilise GABAergic volume transmission (sprinkler, not hosepipe-like wired transmission)
o This can activate astroglia
Explain GABA(B)R-mediated heterosynaptic short-term plasticity
• Glu synapse with GABAB
• You can get released GABA to bind to and activate presynaptic GABABRs on Glutamatergic nerve terminals
• Activity at 1 synapse can transiently affect activity at another synapse
o Due to GABA ‘spill-over’ during conditioning (short-term plasticity)
o GABA diffuses between synapses; accesses GABABR on presynaptic neuron of glutamatergic nerve terminal heterosynaptic short-term plasticity
• GABABR antagonist removes this depression – Glu release remains the same when GABABR antagonist is applied
