NTProfile Flashcards
NT types
1) Amino acid neurotransmitter (-COOH)
e. g. Glutamate, GABA, Glycine
2) Amine neurotransmitters (N)
e. g. Ach, Norephinephrine, Dopamine
3) Peptide neurotransmitters (big)
e. g. Substance P, ligand for opioid receptors
Example NT for AA Amines Purines Peptides FA
AA: GABA, Glutamate, Glycine Amines: Ach, DA, Epinephrine, Histamine, NE, 5HT Purines: ATP, Adenosine, GTP Peptides: Substance P, Pain related etc FA: cannabinoid
Technique: How can NT be labelled
- Inject NT candidate
2. Withdraw specific antibodies (Marked with a label)
Antibodies can be raised against
- Specific synthesizing enzymes like GAD65 and 67 for GABA
- The vesicular transporters -> good target as it is relatively big in size
- The high affinity reuptake transporters
- Small neurotransmitters like GABA or Glu themselves -> by beads labelling but it is a little unspecific
Why is identifying type of NT important?
To classify type of neurons
Other (less common) way for labelling NT
1) In situ hybridisation
Radioactively labelled probe with the complementary nucleic acids binds to mRNA of the target
2) Genetic manipulation
Use of GFP either cloned into a NT gene
Only way to experiment while alive allowing it to conduct some electrophysiology
Where are NT synthesised?
Generally, small neurotransmitter are synthesized in the synapse whereas peptide neurotransmitters are synthesized in the soma!
Peptide neurotransmitters:
- Are synthesized in the soma as precursor proteins
- Often one mRNA codes many repetitions of the same molecule
- The precursor proteins are processed in the vesicle while being transported to the axon
How do positively charged NT get stored?
e.g. monoamines, Ach
Exchange: 2H/NT
1. Acidification of vesicle via vATPaseRaising both chemical (pH) and electrical (+) gradient by pumping in H+ using ATP
2. The extra proton provides E for pumping in the NT
3. For positively charged, !electrical gradient! allows the exchange of proton and NT+. Which leads even more acidic in vesicle, leading to exchange of H+ and Cl-
How do negatively charged NT get stored?
e.g. Glutamate
Exchange: 1H/NT
1. Acidification of vesicle via vATPaseRaising both chemical (pH) and electrical (+) gradient by pumping in H+ using ATP
2. The extra proton provides E for pumping in the NT
3. For negatively charged, the !electrical gradient! allows the exchange of proton and NT-
Which NT is neither positive or negatively charged?
GABA
What does a sudden increase in error bars mean?
In general, when a research shows a graph which includes a sudden increase/difference in error bars, it could be explained by change in sample size (some died etc)
Q. What is the effect of acidification during vesicle loading?
Assist to pack the NT molecule more densely, by making it more neutral
Q. Is the synaptic cleft also acidified? What are the consequences?
Yes, it changes the neurotransmission!
e.g. 1) @ retina
In the dark, Na+ channels are constantly open leading to “dark” current -> increase in vesicle release
e.g. 2) @ photoreceptor (retina)
Acidification of cleft suppressed the current
e.g. 3) @hippocampus
ASIC1a currents are depolarizing and activate inhibitory interneurons
How is acidification of synaptic cleft related to epilepsy?
Acidosis evoked EEG seizures were NOT weakened for mice deficient in ASIC1a (pH sensitive cation channel). pH drop mediates a cationic current, leading to depol of neurons. This allows inhibition of epileptic symptoms as
1) depol neurons are inhibitory (@ hippo: inhib > exit, opposite of amyg)
2) Inhibitory interneurons have larger acidosis induced currents than pyramidal cells
3) Interneurons in wt respond to acidosis with increased activity
=> ASIC1a currents are depolarising and activate inhibitory interneurons
What are the players for controlling acidification in neurotransmission
Not only proton but also chloride
What drugs are used to regulate NT transport/loading into vesicles?
1) Reserpine inhibits vesicular monoamine transport and leads to MA depletion, leading to clinical depression
2) Amphetamines lead to the release of vesicular MA stores into cytoplasm and synapse are psychostimulants and can induce psychosis
What are the general steps for neurotransmission?
1) precursors travel along the axon
2) precursor get altered to NT around axon terminal
3) NT get loaded into vesicle via vesicular transporter
4) release
5) receptor action
6) inactivation (reuptake, glia, diffusion or degradation)
What are the example of inactivation
- degradation of the Neurotransmitter (ACh)
- by reuptake (Glu, Gaba) and passively by diffusion
Glu(tamate)/Glutamic acid Profile
!Source/metabolic origin: Kreb cycle or glia
Precursor: glutamine α-Ketoglutarate
!Enzyme: PAG & Transaminase
!Storage/Transporter name: vGluT
E provider: electrochemical (& H+ antiport)
!Inactivation/Transporter name: no enzymatic/EAAT
E provider: Na+ gradient
Localisation: Glia and neuron
!Recycling: Reuptake and glial Gln Shuttle
Glutamate synthesis & enzyme()
1) α-Ketoglutarate (mostly from KREB, some from glia)
- (Glu-aminotransferase/aka transaminase)-> Glu
2) Glutamine
- (Phosphate Activated Glutaminase)->Glu
Glutamate Storage
- via vGluT
- Highly specific; charged by ELECTROchemical gradient of proton
- 3 isoforms (1&2 neuronal, 3 neuronal & astrocyte)
- also act as a phosphate carrier/transporter, on presynaptic cell, which activate PAG to synthesise more Glu
Glutamate inactivation
- via Exitatory AA Transporter
- E used for plasma membrane: Na+
- E used for vesicle memb: proton and ATP
- co-transport of K+, which restores the original configuration of the transporter (Glu binding site open to the outside)
- Cl-conductance is independent of Glu binding and transport
- 2 modes of operation:1) transient and fast buffering in which the availability of Glu is reduced but no real transport takes place (just binding)2) actual transport of Glu molecules
EAAT localisation
Inactivation transporter for Glu
@hippo
-EAAT1&2 on glia
-EAAT3&4 on neurons
-Glia cells in this region only marginally/loosely surround synapses-> allows Glu to accumulate in cleft-> allows Glu to activate nearby cleft
-Findings with TBOA (EAAT inhibitor)
Inhibition of Glu (excitatory AA) uptake seems to affect synaptic transmission only at high frequencies
@ cerebellum
-Glia cells (Bergmann cells) in this region only tightly surround synapses-> efficient uptake
-Findings with TBOA
Inhibition of Glu (excitatory AA) uptake seems to affect synaptic transmission both at low and high frequencies
What is TBOA
EAAT inhibitor
Glutamate receptors
NMDA: Mg blocked
AMPA: important role in LTP
metabotropicGluR
GABA profile
!Source/metabolic origin: α-Ketoglutarate (Kreb cycle)
Precursor: Glutamate
!Enzyme: GAD 65 & 67
!Storage/Transporter name: VIAAT
E provider: H+ antiport & electrochemical
!Inactivation/Transporter name: no enzymatic/GAT
E provider: Na+ gradient
Localisation: Glia & neuron
!Recycling: reuptake
Glutamate synthesis & enzyme()
Storage
Inactivation
Receptor/recycling
!Source/metabolic origin: Precursor: !Enzyme: !Storage/Transporter name: E provider: !Inactivation/Transporter name: E provider: Localisation: !Recycling:
GABA synthesis & enzyme()
de-carboxylating (CO2) glutamate
GABA = Glutamate with COOH removed
Glu
- (glutamic acid decarboxylase (GAD, 2 isoforms GAD 65 and GAD 67))
- > GABA
Where is GABA located?
Only in neural tissue (Glu & Gln is everywhere)
