Inhibitory GABBA and glycine receptors

Question 1

What is the molecular mechanism of GABBA?

Answer 1

Enzymes (GABA-T, GAT) produce GABA from a-ketoglutarate.

The receptor is a heterodimer with GABAB1 providing the GABA binding domain and GABAB2 the G-protein coupling to Gi & Go.

Presynapse: the activation of GABA b causes the Go complex to decrease
the probability of Ca2+ channels opening, and so decreasing the quantal release of neurotransmitter and hyperpolarisation. This consequently decreases
GABA release upon a second stimulus.

Post synapse: GABA acts post-synaptically to activate chloride-conducting
GABAaRs which activates Cl- channels, cl- influxes, causing an IPSP (EPP equivalent)

The activation of GABA b causes Gi complex to OPEN K CHANNEL which in turn causes calcium channel to open, calcium leaves the cell, and cell HYPERpolarises MORE AND FOR LONGER and produces a PROLONGED IPSP

So the IPSP produced is controlled by both GABA a and b receptors

Question 2

What is the molecular mechanism of GYCINE?

Answer 2

There are specific transporters (GlyT1, and GlyT2)

Question 3

How has molecular biology technology discovered isoforms of the receptor

Answer 3

Mice knockouts,

Chimeras

Question 4

What are the properties of the isoforms?

Answer 4

knock out mice show:

alpha 1: sedation

alpha 2 and 3: anti anxiety and anti pain

alpha 5: cognition; without this subunit there was a lack of learning and memory

Beta 2: sedation and anaesthesia

Beta 3: immobility and anaesthesia

A SINGLE mutated amino acid would cause the malfunction of any of these subunits; prospect for future drug targets.

Question 5

What was shown through a comparison of the pharmacological properties of GABAA receptor isoforms in conjunction with the use of site directed mutagenesis?

Answer 5

revealed binding domains on the protein for GABA and benzodiazepines.

Different isoforms in different localisation in the CNS and therefore mediate different behaviours; allowing for selective drugs.

Question 6

In what way do NEUROSTERIODS act as regulators

Answer 6

may act as endogenous regulators of GABA-ergic transmission.

Patch clamp experiment: shows neurosteriod keeps channel open for longer so inhibition is prolonged.

Question 7

List types of drugs that effect GABBA.

Answer 7

Drugs that enhance GABA AR function and neural inhibition are:

1) Anxiolytic
2) Anticonvulsant
3) Analgesic
4) Amnestic
5) Sedative/hypnotic
6) Anaesthetic.

For example, propofol, steroids ans anaesthetics and alcohol

Question 8

Describe the differences between the drugs Barbiturates & benzodiazepines

Answer 8

They both enhance but Benzodiazepines increases the probability of the channel opening

Barbiturates promote open states of LONGER and directly activate the receptor.

Question 9

Describe how GAMMA A additional subunits were found other than subunits a and b?

Answer 9

In frog oocytes were injected with alpha and beta subunits in receptor which was activated by GAMMA and blocked by GAMMA inhibitors, enhanced by Barbiturates but IT WAS NOT ENHANCED BY benzodiazepines.

Therefore, there must be additional subunits that are enhanced by benzodiazepines such as subunit gamma2.

Question 10

What is the structure of the GABA receptor

Answer 10

Pentamer of 2 alpha and 2 beta subunits and gamma subunits,

2 GABA binding sites between alpha and beta subunits, a central Cl- pore and a beta2 site between alpha and gamma subunits.

It has 4 transmembrane domains

The important residues found. the interface between alpha and beta has 3 DISCONTINUOUS beta LOOPS and 3 DISCONTINUOUS alpha LOOPS which form a binding pocket.

Alpha subunit, is negative, beta is positive side of the interface.

Question 11

How was the binding site of benzodiazepine found?

Answer 11

The receptor insensitive to benzodiazepine and the receptor sensitive to benzodiazepine drug were compared using a chimera of the two receptor; it was found a mutation at Histamine 101 to Arg responsible for insensitivity (so this H amino acid) needed for enhancing

Question 12

What is The effect of Strychnine on the Glycine receptor?

Answer 12

Strychnine is a potent competitive antagonist; competing against glycine.

Stops inhibition

Enhances the perception of pain & causes convulsions.

The receptor is ANION selective and also accepts taurine.

Question 13

What functions do glycine receptors regulate in the body? Where is it distributed?

Answer 13

1) respiratory rhythms
2) motor control
3) muscle tone
4) sensory & pain processing.

Limited distribution in CNS (less than GABA), found in the cerebellum and dorsal raphe.

Question 14

List some subunits in GABA and glycine

Answer 14

GABA: alpha 1-6, beta 1-3, gamma 1-3

Glycine alpha 1-3, beta 1-4

Different combinations of subunits make up different receptors.

Question 15

What is baclofen?

Answer 15

Baclofen – a GABABR agonist used clinically to treat spasticity.

Question 16

Where is GABA produced?

Answer 16

In the GABA interneurone.

There are many different types of GABA interneurons,
that differ in
1) their firing rates

2) the part of the principal neuron
they innervate

3) the subtype of synaptic GABAaRs they activate.

Question 17

In what way is the glutamate EPSP govered by the GABA and shows the coordinated activity of principal neurons & interneurons?

Answer 17

The EPSP has a limited time summation to reach A.P (depolarised) before GABA inhibits with IPSP (hyperpolarisation)

The release of GABA to the pyramidal neuron controls how long cell can be excited for in a feed forward manner.

However Feed-back inhibition is shown as The firing of the pyramidal neuron activates an inhibitory
interneuron, which in turn inhibits the pyramidal cell. Once the inhibition decays, the pyramidal cell can fire again, initiating another cycle of inhibition.

Question 18

Where is glutamate produced?

Answer 18

The pyramidal neurone

Question 19

Give an example of GABA interneurone diversity

Answer 19

CCK = cholecystokinin 
interneurons. 
Asynchronous
release of GABA.
Express CB1 
receptors

PV = parvalbumin
fast spiking interneurons.
Synchronous GABA 
release.
Express µ opioid receptors

CCK more prolonged than PV due to receptor types.

Question 20

What effect does adding steroid progestrone have on fish? How is this seen in humans?

Answer 20

There is rapid induction of general anaesthesia.

In small concentrations (10nM) steroids GABA response starts to increase, in pregnancy steriods concentration increases greatly up to 100nM to maximise GABA activity (counteract pain)

Question 21

How does the brain have auto control?

Answer 21

It makes local neurosteroids that influence synaptic & extrasynaptic GABAA receptors

Overall theselink to brain activity regulation; they are not produced at constant levels and can be changed by drugs (prosac-antidepressant) or alcohol.