L08 The GABAergic System

The GABAergic System

• Main inhibitory transmitter in the CNS

• Acts on GABA receptors: GABA_A , GABA_B and GABA_C

• GABAARs are most well-studied, will be our focus

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• GABA type A most important

GABA synthesis

• Made from glutamate via glutamic acid decarboxylase (GAD65 + GAD67)

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• key synthesizing enzyme

• GAD65 + GAD67 marker of GABAergic neurons (key in many studies)

GABA levels

• Abnormal levels associated w/mood disorders (e.g. depression, evidence inconsistent)

• Interest in modifying GABA levels w/supplements

• Research problematic

  • Conflicts of interest are apparent

  • Passage across blood-brain barrier questioned

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• controversial to say OTC supplement would max/min GABA levels

• supplement unlikely to reach the brain → doesn’t cross BBB

The basics of GABA

• Receptor subtypes

• Receptor localization

• GABA synthesis and metabolism

• Transport

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GABA_A receptors

• Ionotropic, generally ligand-gated channels

• Permeable to Cl- ions

• Fast inhibition (hyperpolarizing and/or shunting)

• Composed of 5 subunits from a family of many members (16+)

  • 2 α (1-6), 2 β (1-3) and 1 of γ (1-3), δ, ε, π or θ

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• targets for GABA

• receptor is a transmitter and a channel → fast inhibitory signal

• key feature: permeable to Cl- ions

  • conductance of -ve ion hyperpolarizing doesn’t make it inhibitory

• clamps/fixes neuron from reaching MP (threshold)

• 5 subunits from subunit pool → makes GABA_A receptor

• 2 alpha, 2 beta, 1 miscellaneous

review

Native GABA_A receptor abundance

• Preferred stoichiometry of 2α:2β:γ or δ. Though many are possible, only a handful exist. Some are more common than others!

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• some are most abundant

• a1, a3, b, gamma are possible but far less common → possibly restricted expression

• significant for learning, memory, mood, and more (ones highlighted)

  • show diff pattern of exp + functionality than other receptors (next slide)

GABA_A receptor localization

Synaptic or extrasynaptic regions

Synaptic:

• αxβxγ2

• αxβxγ3

• “strong but transient”

Extrasynaptic:

• α4βxδ

• α6βxδ

• α5βxγ2

• αxβx*

• “Weak but always on”

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• GABA binds to receptor → brief, strong current

• focus: receptors that are outside (extra synaptic) → operate in low levels of GABA, not action-dependent release of GABA

• middle = phasic (action-dependent release of GABA associated); extra synaptic + tonic currents

• review

Our discussion of learning and memory will focus on α5 and δ receptors.

• lower affinity + higher efficiacy

• opposite true for extrasynaptic (bc low GABA and would otherwise not be on)

Excitatory and inhibitory signals

• more signallng, less likely firing thru hyperpolarization/shunting

Summation of signals

• Any one neuron generally receives many signals

• All these signals are summated

• If the sum exceeds a given threshold (excitatory >>> inhibitory), the neuron will fire

• Inhibitory signaling decreases the likelihood of firing

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• excitatory + inhibitory signals incoming simultaneously

• what is relative amount of each?

• good amount of GABA keeps it from firing, but lots of ex would also may cause it to fire

GABA_ARs reduce excitability

• Top: Activating GABA_ARs with THIP decreases excitability → less APs / time unit

• Bottom: Blocking GABA_ARs with picrotoxin increases excitability → excitability would skyrocket

  • blocking → less pA needed to fire neuron

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• excitability = amount of AP / time unit

• less excitable = less APs/ time unit (and vice versa)

GABA and neuroactive steroids

• The activity of GABA at its receptor can be modified by many compounds, including neuroactive steroids.

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• steroids can act on receptors; tuning role increasing activation of receptor in presence of GABA (pregnenolone, progesterone etc.)

• positive, allosteric modulators b/c of delta receptors (they will tune up in presence of GABA)

Neuroactive steroids

• Steroid hormones naturally produced by the body

• Levels vary during development (e.g. puberty), reproductive cycle (e.g. postpartum) and with stress

• Generally increase activity of GAB_AA receptors, particularly δGABAA receptors

  • Positive allosteric modulators

• Some have anxiolytic and antidepressant properties

  • Brexanolone/Zulresso for post-partum depression (IV) and Zuranolone/Zurzuvae (oral)

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• more activated by agonists

GABA transporters (GAT)

• Three types; GAT1 + GAT3 most abundant in the brain

• Found in neurons (GAT1) and astrocytes (GAT3)*

• Target of clinical drugs (e.g. antiseizure medications)

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• how GABA is moved around

• drugs of epilepsy target GABARs

Controversies

• Some GABA_A receptors do not require GABA to open (spontaneous, ligand-independent)

• Shifts in GABA receptor expression + function during development (perinatal, pubertal, adult period)

  • Early on, GABA may be excitatory

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• have yet to be resolved

• GABA might be excitatory in developing neurosystems

GABA and interneurons

• Interneurons are not sensory or motor cells, but modulate their signals (e.g. reflexes)

• Synapse on many different compartments

• GABA signaling restricts excitability and shapes neuronal oscillations (L07)

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• inhibitory interneurons can play a role in reflexes

• theories that some of these cells contribute to disorders (like SZ)

• oscillations = rhythmic synchrony of neuron firing

• review

Anti-seizure medications

• Drugs that enhance inhibitory transmission via GABA

  • Affect GABA receptors, GABA transporters

• Drugs that reduce excitation via glutamate

• Drugs affecting channels in the action potential

  • Inhibit voltage-gated sodium + calcium channels

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• for treatment in epilepsy → viewed as disorder of imbalance in excitation + inhibition

Role in neurogenesis

• GABA in neurodevelopment

• evidence suggests that GABAR are expressed early on

• GABAergic signaling may be significant in early neurons

• if we activate, neurogenesis should activate

• or KO of GABAR would affect adult neurogenesis

• review

Drugs affecting GABA

• Many effects: Anxiolytic, Amnestic, Anesthetic, Sedative-hypnotic, Antiepileptic, Analgesia

• Included in this category are benzodiazepines, barbiturates, anesthetics, alcohol and anticonvulsants

• Some drugs are more preferred than others for a given effect (e.g. benzodiazepine > barbiturate)

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Synapses are modifiable!

• Changes in synaptic strength might contribute to learning

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• strength of signaling from neuron A > B can change

  • A can become better at activating B = long-term potentiation (synapse b/w A + B gets stronger over time)

Synaptic strengthening

• First compelling evidence obtained by Bliss + Lomo (1973) in the hippocampus

• High-frequency stimulation of synaptic connections persistently strengthened them

  • experience-dependent

• This phenomenon is called long-term potentiation (LTP) is viewed as a neural correlate of learning

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• potential for synapse to get stronger is modified by GABA

What is LTP?

• A signaling to B (done artificially, but in reality A active due to learning)

• A weakly activates B under normal conditions

• stimulating synapse 100x/sec → synapse gets stronger

LTP and Memory

• LTP is seen throughout the nervous system, including the hippocampus, cortex, striatum + spinal cord

• LTP is often correlated with learning and memory

  • Deficits in learning/memory are linked to deficits in LTP

• Activity-dependent variations in synaptic strength such as LTP may be a fundamental mechanism by which we acquire and modify all behaviors

  • Includes pain, motor learning and substance use disorder

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• in animals with less LTP, memory deficits are seen (rough correlation b/w learning + LTP in animals) → invasive to stimulate + record in humans

• although LTP well characterized in HC → plasticity may be seen anywhere (many synapses may get stronger); anything that can change w experience (pain, substance seeking behaviour) may involve LTP

• LTP may not be affected by just GABAergic signaling

GABA_A receptors restrict LTP

• Agonists tend to impair, antagonists increase

• Baseline differences (e.g. DG vs. CA1) might be explained by inhibition

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• GABA agonist → less/no LTP seen

• inhibit GABAergic signaling → increases LTP (picrotoxin, PTX, and bicuculline does this)

  • if you give 1 and 3 to human → inhibitory removal by drugs increases likelihood of seizures

The story so far…

• LTP and the hippocampus are both important for learning/memory

• GABA constrains hippocampal LTP (agonists ↑, antagonists ↓)

• Based on this, you might argue GABA impairs learning and memory

• This does happen in a few cases, which we’ll cover first

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• agonists impaired LTP, antagonists increase LTP

• GAPA impairs learning + memory but not in all

GABA_A receptors and memory

α5KO mice show improved learning in the Morris water and trace fear conditioning tasks

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• α5R = extra synaptic receptor

• both associated with HC, which is an area where these receptors are expressed

A study in contrast…

δKO mice might show enhanced fear conditioning, too!

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• again, delta = presynaptic receptor

• improved learning + memory in some tasks but not others

GABA_A receptors and memory

• Drugs which increase GABAA receptor activity (e.g. anesthetics, benzodiazepines and alcohol) impair memory

  • Many of these memory effects have been linked to activation of α5GABA_ARs and/or δGABA_ARs

• States of increased GABAA receptor expression/activity are often associated with impaired memory

  • Inflammation

  • Traumatic Brain Injury

  • Reproductive cycle

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• more GABA bad for learning

• activate GABAR + interact them w substances → learning impairments

• changes in GABAR expression impairs learning + memory

On reproductive cycles…

• As hormones are metabolized to neuroactive steroids, hormonal levels are a factor in GABA receptor function

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• neuron active steroids → action depends on pathway

• peripherally derived (level-dependent)

  • spike in hormonal levels + rapid drop = significant change in GABAR (specifically gaba delta a receptors to drugs)

Role of reproductive cycle

• Peak in steroid levels during diestrus in females linked to impairments in memory; changes absent in δKO

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• they do worse at fear conditioning likely due to hormonal variations, having consequences for activity + receptors

• WT mostly impaired → KO won’t (deleting receptors → less significant effect)

• also receptors important in postpartum, puberty, etc.

• review

Simple principle of “lower GABA, better memory” doesn’t always hold.

Some types of memory might require GABA signaling and be impaired by its removal.

Consider the following task… → Distinguishing experiences

• X and X’ might each have neural representations

• Large differences (d = X-X’) might be useful in discriminating them

• To maximize discrimination, the brain might transform similar inputs (X and X’) into different outputs (X and Y)

• This ability is termed pattern separation

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• discrimination tasks may be better with GABA inhibitory signaling

• brain takes similar stimuli (visually) and stores is diff in brain (to maximizing differences for late)

• orthogonalization = making inputs different = pattern separation

Pattern separation

Thought to involve the dentate gyrus (DG) of the hippocampus

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• DG key in pattern separation - ideal for orthogonalization in this brain area

GABA and memory discrimination

• Inhibition is thought to be critical to pattern separation; loss of it is associated with impaired discrimination

• Hyperactivity in the DG in humans is associated with reduced pattern separation and may contribute to cognitive impairment in dementia

• α5KO and δKO mice show poor pattern separation and treat different environments as being similar

  • In both cases, receptors in the DG are implicated

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• information coding could never be selected w absence of inhibition

• hyperactive DG = poor performance in discrimination in humans → same effect on KO mice as well

• better fear conditioning but poor pattern separation → leads to concept that inhibition may be important but may also impair

Role of GABA depends on…

• The receptor subtype involved (α5 and δ receptors are different)

• The learning task (e.g. FC/NOR vs. pattern separation)

• Sex of animal (males and females, linked to hormones)

• Physiological state (e.g. puberty, stress, reproductive, again due to hormones)

Anxiety disorders

• Generalized Anxiety Disorder

• Panic Disorder

• Phobias

• Related conditions, but now independent in the DSM-5, are obsessivecompulsive disorder (OCD) and post-traumatic stress disorder (PTSD)

Role of the amygdala

Limbic system structures (amygdala, hypothalamus, orbitofrontal cortex, cingulate gyrus + hippocampus)

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• emotional processing, fear-conditioning → amygdala

• inhibition of such cells → expect less fear and anxiety

The Amygdala and Anxiety

• Activated during arousing states (e.g. emotion) and other situations

• Lesion can reduce fear/anxiety (e.g. Patient SM)

• Selectivity inhibiting the amygdala (e.g. with drugs) reduces anxiety

• Many anxiolytics increase GABAA receptor activity (see L01) and perhaps work by affecting the amygdala

• Is it the case that more GABA = better mood?

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• short term, benzodiazepine

• long term, SSRIs → better tolerated + fewer risks

GABA and mood

• Correlative evidence in humans and experimental evidence in animals

• In humans, there may be lower expression of GABA_A receptors in psychological disorders

  • GABA levels may also be reduced (e.g. depression)

• Reduced levels of δ and γ2 subunit-containing GABA_A receptors are linked with anxiety

• GAD65-KO mice also have elevated anxiety levels

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• variations in GABA signaling may lead to anxiety?

δKO + post-partum depression

• Deletion of the δGABAA receptor subunit is associated with depressionlike behavior* post-partum

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• reproductive cycle → associated w changes in GABAergic signaling due to changes in steroids…?

• deletion of GABAR = post-partum like phenotype

• pups survive less + die at higher rates + build lower quality nests

Other cases of variation

• receptor sensitive to hormonal levels →

Summary

• Natural variations in GABA signaling might contribute to natural variations in mood (puberty, post-partum…)

• Increased GABA signaling is associated with reduced anxiety (in general)

• Decreased GABA signaling is associated with increased anxiety and depression

• Modulating GABAA receptors w/drug affects anxiety*

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• treat anxiety thru substances that affect receptors

Drugs affecting GABA

• increasing dosages → different effects to GABAergic signaling

When it comes to anxiety…

…there are many drugs possible, not just GABA drugs

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What is pain? + Types of pain

• Private experience (sensory and emotional) associated with actual or potential tissue damage (belief of tissue damage)

• Adaptive, allows us to identify danger + withdraw

• Types of pain:

  • Acute pain is brief, and overlaps with the healing process (associated w injury)

    • Normal and more manageable

  • Chronic pain is persistent (>3 months) beyond healing period (knee if fine but still experience pain)

    • Many disorders include chronic pain as a symptom (e.g. diabetic neuropathy, migraine, fibromyalgia, arthritis…)

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• useful signal bc it is unbearable

Why do we get chronic pain?

• Following injury, changes in nociceptive signaling system can occur

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• on at wrong times, and too on/active at right times

Pain pathways

• The primary afferent – second order sensory synapse in the dorsal horn is key + early

• The activity of dorsal horn neurons is modulated by other cells (esp. inhibitory interneurons)

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• primary afferent into SC

• circled - may be interneurons present

Changes in inhibitory balance

• Death of interneurons could increase pain (due to regulation in pathway)

• With changes in transport across the membrane, inhibitory signals could “lose their value”, becoming insufficiently inhibitory or even excitatory

• Even if inhibition remains stable, increases in excitation could occur that amplify pain-related signals (central sensitization)

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• inhibitory signals could stop being inhibitory (removing gradients bc ions are no longer being conducted)

• weaker ones may be stronger (review)

Central sensitization

Signaling at this synapse may increase in strength after injury or other forms of stimulation.

DNM diagram

Drugs for pain

• GABA signaling may regulate pain and GABA receptors are a potential target for analgesic drug

• Pharmacological activation of δGAB_AA receptors, for example, can have analgesic effects

• Drugs well-known for their analgesic effects, including gabapentin, might work in part by increasing δGAB_AA receptor expression

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• certain drugs could be analgesic effects through GABAergic signaling indirectly