GABA and Benzodiazepines Flashcards
What does GABA stand for?
Gamma-aminobutyric acic
How is GABA made?
GABA is an amino acid neurotransmitter
Made from glutamate by the enzyme glutamic acid decarboxylase
What is the main function of GABA?
Inhibitory
Makes the neuron less likely to fire an AP or release neurotransmitters
GABA Receptors
2 types:
GABAA and GABAB
GABAA Receptors
Ionotropic
Opens chlorine channel > chlorine enters neuron > hyperpolarises neuron
GABAB Receptors
Metabotropic/GPCR
Activation of receptor opens different potassium receptor > potassium leaves neuron > neuron hyperpolarises
Termination of GABA
GABA transporters take up GABA from the synapse into the neuron or glial cells, where GABA is degraded primarily by mitochondrial enzymes
Sedative effects of GABA
Increased GABA activity = reduced neural transmission = sedative effects
Alcohol and benzidiazepines (ie diazepam) increase activity at the GABA receptor
What are the (side)effects of drugs enhacing GABAaR function?
Effects:
Anxiolytic (reduces anxiety)
Anticonvulsant
Analgesic (reduces pain)
Amnestic (loss of memory)
Sedative/Hypnotic
Anaesthetic
Side effects:
Impair cognition
Produce tolerance
Become addictive
What drugs enhance GABAaR function?
Benzodiazepines, steroids, barbiturates
What are PAMs of the GABAaR? Give examples
PAMs are Positive Allosteric Modulators (enhance function)
Bind at the benzodiazepine binding site
Bind between the alpha and gamma subunit interface
Examples: diazepam, flunitrazepam
What are NAMs of the GABAaR? Give examples
NAMs are Negative Allosteric Modulators (decrease function)
Bind at benzodiazepine binding site
Bind between the alpha and gamma subunit interface
Examples: DMCM (a beta-carboline that is pro-convulsant and anxiogenic)
What are antagonists of the GABAaR? Give examples
Antagonists do not influence the action of GABAaR but prevent the action of PAMs and NAMs
Bind between the alpha and gamma subunit interface
Example: flumazenil (inert)
What are the most commone GABAaR subtypes?
alpha1,beta-gamma2 > alpha3, beta-gamma2 > alpha2, beta-gamma2
GABAaR with apla1-beta1-gamma2 subunits
Benzodiazepine sensitive
Benzodiazepines bind between the alpha and gamma subunit interface
Benzodiazepine only enhances GABA responses for the alpha1-beta1 subunits joined with gamma2 (for other gammas it does not)
What does diazepam need to facilitate (high affinity) binding?
For high affinity bidning, the binding site must contain a gamme subunit, usually gamma2
Gamma2 + alpha1,2,3, or 5 are diazepam sensitive (have H residue at position 101)
Gamma2 + alpha6 or 4 are diazepam insensitive (have R residue at position 101)
alpha1H101R mouse
Causes loss of 3H-flumazenil binding binding in areas known to express alpha1
Sedative (not myorelaxant) effects of diazepam are blunted (not blunted in alpha2H101R mouse)
Anxiolytic, not sedative
alpha2H101R mouse
alpha2-beta-gamma2 GABAaRs mediate the anxiolytic effects of diazepam
Can be tested with the light/dark box, or elevated plus maze
Anxiolytic effects do not occur in alpha2H101R
Sedative, not anxiolytic
Function of benzodiazepines in inflammatory and neuropathic pain? What are the testing methods?
Intrathecal (injected into the spinal cord) diazepam supresses response to heat and mechanical stimuli
Primarily mediated by alpha2/3 GABAaRs
These effects are maintained in the alpha1H101R mouse but not the alpha2H101R mouse
Heat hyperalgesia - paw withdrawal latency upon exposure to defined radiant heat
Mechanical sensitisation - paw withdrawal with Von-Frey filaments
Cold allodynia (pain from harmless stimulation) - time spent lifting, shaking, licking paw after drop of acetone
alpha5-NAMs as cognitive enhancers?
Diazepam/flunitrazepam impair cognition (esp. + alcohol)
NAMs of alpha5-beta-gamma2 subtype exhibit pro-cognitive properties in mouse models of:
HD
Alzheimer’s
Down’s syndrome
Also enhances impaired hippocampal CA1 long term potentiation (LTP)
How is cognition assessed in mice?
Morris water maze
Novel object recognition task
Relation between alpha5-GABAaRs and cognition?
Expressed extrasynaptically in hippocampal CA1 neurons (mediate tonic current)
alpha5 knockout mouse shows improved performance in water maze
Cognitive impairment associated with diazepam is absent in alpha5 knockout mice
NAMs of alpha5 GABAaRs improve cognition and LTP in mouse models of HD, alzheimer’s and Down syndrome
Overview of the function of benzodiazepines on different alpha subunit GABAaRs
Alpha1 - sedative
Alpha 2 - anxiolytic, analgesic
Alpha5 - impairs cognition
How are novel GABAaR subtype-selective benzodiazepines identified?
- Compound synthesis
- Displacement of 3H-flunitrazepam from rat brain membranes (determines affinity for receptor)
- Displacement of 3H-flunitrazepam from cell lines expressing recombinant human GABAaRs (affinity for human receptor and various subtypes)
- Functional electrophysiological studies with a) cell lines expressing recombinant GABAaRs and b) neuronal GABAaRs (determines appropriate efficacy)
- Appropriate pharmokinetics:
CNS receptor occupancy
Changes in EEG activity (mouse/rat) (determines whether it can cross the blood/brain barrier, and the duration of action) - Behavioural studies:
Sedation/self-administration - alpha1
Anxiolytic/analgesic - alpha2/3
Cognition - alpha5
(behavioural profile)
