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NEUROTRANSMITTERSYSTEMS
Mary ET Boyle, Ph. D.Department of Cognitive ScienceUCSD
Chemical Synapses can______
A have variable signaling
B mediate complex information (excitatory/inhibitory)
C amplify neuronal signals
D All of the above are correct
Electrical Synapses ______
A have variable signaling
B have gap junction channels
C amplify neuronal signals
D have significant synaptic delay
Three classes of neurotransmitters Amino acids, amines, and peptides
Many different neurotransmittersDefining particular transmitter systems
molecule, synthetic machinery, packaging, reuptake and degradation, etc.
Acetylcholine (Ach) First identified neurotransmitter (Otto Loewi)
Nomenclature (-ergic) Cholinergic and noradrenergic
Introduction
Ligands
Ligand-binding methods Identify natural receptors using radioactive ligands Can be: agonist, antagonist, or chemical
neurotransmitter
Molecular analysis two receptor protein classes Transmitter-gated ion channels
GABA receptors 5 subunits, each made with 6 different subunit
polypeptidespotential for enormous diversity
G-protein-coupled receptors - metabotropic
Ligand: any chemical compound that binds to a specific site on a receptor
Neurotransmitter Chemistry
Evolution of neurotransmitters Neurotransmitter molecules
Amino acids, amines, and peptides ACh is derived from acetyl CoA
Dale’s Principle One neuron, one neurotransmitter
Co-transmitters Two or more transmitters released from one nerve
terminal An amino acid or amine plus a peptide
Studying Neurotransmitter Systems1. Synthesis and storage in presynaptic neuron
Presynaptic neuron should contain a transmitter and the appropriate enzymes needed to synthesize the neurotransmitter.
2. Released by presynaptic axon terminalOne should be able to isolate the transmitter and
characterize its structure using biochemical or molecular biological techniques.
3. Produces response in postsynaptic cellMimics response produced by release of
neurotransmitter from the presynaptic neuron
4. Removal of the transmitter from the synaptic cleft
Neurotransmitter– four steps
Synthesis in presynaptic neuron
Precursors should be present in the appropriate places
Enzymes involved in the process should be in the active form
Enzymes should be localized to the appropriate compartment
Synthesis of the xmtr in presynaptic neuron
2
3
1
Neurotransmitters must have ____
A Synthesis and storage of putative xmtr
B Released by the terminal under normal conditions
C Have a response in the postsynaptic terminal
D All of the above
Transmitter Localization
Transmitters and Transmitter-Synthesizing EnzymesImmunocytochemistry –localize molecules to cells
Primary antibody: Rabbit anti-laminin a1 (basement membrane marker), 1:400
Primary antibody: Rabbit anti-laminin a1 (basement membrane marker), 1:400
In situ hybridizationLocalize synthesis of protein or peptide to a cell (detect mRNA)
Storage of neurotransmitter in terminal
Classical and Peptide transmitters are stored in vesicles
(to protect them from enzymatic degradation)
In vesicles make them available for quick release
A mechanism must be present to transport the xmtr or peptide into the
vesicles (e.g. vesicular transporter)
Storage of the xmtr and/or precursors in the presynaptic nerve terminal
2
3
1
Release of the neurotransmitter into the synaptic cleft
Vesicle fuses with cell membrane to release contents into cleft
Release into the synaptic cleft
Transmitter Release Transmitter candidate: Synthesized and localized in terminal and
released upon stimulation CNS contains a diverse mixture of synapses that use different
neurotransmitters Brain slice as a model
Kept alive in vitro Stimulate synapses, collect and measure released chemicals
Buchner funnel
Binding and recognition of xmtr by target receptors
Neurotransmitters that are released interact with receptors
Two types of receptors on target cells: xmtr-gated and metabotropic
Autoreceptor on pre-synaptic neuron
Binding and recognition of xmtr by target receptors 2
3
1
Removing transmitter is critical to synaptic transmission
Removing transmitter enables a new signal to get through
The synapse would become refractory because of receptor
desensitization due to exposure
Three removal mechanisms: diffusion, degradation and reuptake
Removal of transmitter from the synaptic cleft terminates synaptic transmission
2
3
1
Synaptic Mimicry
Qualifying condition: Molecules evoking same response as neurotransmitters
Microionophoresis:Assess the postsynaptic actions
Microelectrode: Measures effects on membrane potential
AgonistA drug that facilitates the effects of a
particular neurotransmitter on the postsynaptic cell.
AntagonistA drug that opposes or inhibits the effects of a particular neurotransmitter on the postsynaptic cell.
An antagonist drug
A Facilitates the effects of a neurotransmitter
B Opposes or inhibits the effects of a neurotransmitter
C All of the above
D None of the above
Direct agonists and antagonists act directly on the neurotransmitter binding site
Indirect agonists and antagonists act on an alternative binding site and modify the effects of the neurotransmitter
on opening of the ion channel.
Sites on transmitter receptors can bind:• Transmitter• Agonist• Antagonist
Agonist-1 Agonist-2
transmitteragonist-2
antagonist-2
One neurotransmitter can bind to many different receptors.No two neurotransmitters bind to the same receptor
Receptor subtypes can be distinguished by the action of different drugs.
One neurotransmitter can bind to many receptors?
A True
B False
C Don’t know
Cholinergic receptor subtypes -
Responds to Nicotine Tobacco plant derivative Agonist – (means that it mimics ACh)
Blocked by Curare (poison darts) Antagonist – (means that it blocks ACh and Nicotine)
Skeletal muscle & CNS
ACh
nicotinic
• Responds to Muscarine• Mushroom (e.g. Amanita muscaria)• Agonist – (means that it mimics ACh)
• Blocked by Atropine (belladona plant)• Antagonist – (means that it blocks ACh & Muscarine)
• Cardiac muscle & CNS
ACh
muscarinic
The tale of two receptors:
nicotine vs. muscarine
Which type of cholinergic receptor is at the endplate?
A Muscarinic
B Nicotinic
C Both Muscarinic and Nicotinic
D There is no difference they both use ACh
“Black widow spider venom stimulates abnormal release of acetylcholine
The bite of the black widow spider initially produces an increase in neuromuscular activity that leads to painful skeletal muscle spasms and rigidity.
This phase of hyperexcitability is rapidly followed by progressive failure of neuromuscular transmission and paralysis.
The venom in ACh followed by a progressive decline and failure of both spontaneous and induced transmitter release and depletion of presynaptic vesicles.”
Barchi RL. (1999) Defects in Neuromuscular Transmission Can Interrupt Normal Muscle Function
ACh Biosynthesis
Acetyl coenzyme A
choline Choline acetyltransferase
(ChAT)
coenzyme A
Acetylcholine
Which enzyme is used to synthesize ACh?
A AChE
B ChAT
C AChM
D None of the above
Cholinergic (ACh) Neurons
Botulium and the venom of the black widow spider, affect the release of acetylcholine.
Botulinum toxin is produced by clostridium botulinum, a bacterium.
Black widow spider venom has the opposite effect: it stimulates the release of ACh.
ACh Deactivation
AcetylcholinesteraseAChE
Choline gets recycled back to the presynaptic
terminal.
Agonists:
Receptors:Rece
ptor
sub
type
s
Each named for a different chemical agonist
Catecholaminergic Neurons Involved in movement,
mood, attention, and visceral function
Tyrosine: Precursor for three amine neurotransmitters that contain catechol group
Dopamine (DA) Norepinephrine (NE) Epinephrine (E, adrenaline)
Dopamine
Produces both excitatory and inhibitory PSPs
Movement, attention, learning, rewards
AMPT inactivates
tyrosine hydroxylase
Parkinson
Disease
SubstantiaNigra
Ventral Tegmental Area
Norepinephrine
Also in ANS –adrenalin
epinephrine noradrenalin
Final synthesis step is in the
vesicle
Fusaric acid inhibits dopamine-
B-hydroxylase
Excess NE in terminal is
destroyed by MAO-A
Serotonergic (5-HT) Neurons
Amine neurotransmitter
Derived from tryptophan
Regulates mood, emotional behavior, sleep Selective
serotonin reuptake inhibitors (SSRIs) - Antidepressants
Synthesis of serotonin
Serotonin
Mood regulation
Released from varicosities rather
than terminal boutons (like NE)
At least 9 different types of
receptors and autorecptors
SSRI – block reuptake receptors
4. Drug stimulates release of NT (e.g. black widow venom – ACh)
2. Drug inactivates synthetic enzyme; inhibits synthesis of NT(e.g. PCPA – serotonin)3. Drug prevents storage of NT in
vesicles(e.g. reserpine – monoamines)
1. Drug serves as a precursor (e.g. L-Dopa – dopamine dopamine)
5. Drug inhibits release of NT (e.g. botunlinum toxin – ACh )
6. Drug stimulates postsynaptic receptors
(e.g. nicotine, muscarine – ACh )
7. Drug blocks post synaptic receptors (e.g. curare, atropine – ACh )
8. Drug stimulates autoreceptors; inhibits synthesis/release of NT(e.g. apomorphine – dopamine)
9. Drug blocks autoreceptors; increases synthesis/release of NT
(e.g. idazoxan – norepinephrine)
10. Drug blocks reuptake(e.g. cocaine – dopamine)
11. Drug inactivates acetylcholinesterase
(e.g. physostigmine – ACh)
They each have their own receptors and do not interact with each other.
amino acids have an amino group and a carboxyl group in
their chemical structures
Amino Acidergic NeuronsDifferences among amino acidergic neurons quantitative NOT
qualitative Glutamic acid decarboxylase (GAD)Key enzyme in GABA synthesisGood marker for GABAergic neuronsGABAergic neurons are major of synaptic inhibition in the CNS
Synthesized from glucose and other precursors.Exist in all cells.Major excitatory transmitter!
Synthesized only by neurons that use it.Major source of synaptic inhibition in the brain.
ATP: Excites neurons; Binds to purinergic receptors
Both transmitter and G-protein coupled receptors
Concentrated in vesicles at many synapses (CNS + PNS) Ca++ dependent release Co-localized with catecholamines
EndocannabinoidsEndogenous
cannabinoidsAnandamide
(“internal bliss”)Arachidonylglycerol
(2-AG)
High Ca++ concentration
Synthesis of endocannabinoidmolecules from membrane
lipids
Small and membrane permeable
Once synthesized they can diffuse to neighboring cells
Bind selectively to the CB1 type of cannabinoid
receptor
CB1 is G-protein coupled receptors
Main effect raise VGCC threshold
Amino Acid-Gated ChannelsGlutamate-Gated ChannelsAMPA, NMDA, kainite
one neuron stimulates another by releasing glutamate into the
synapse
1
the glutamate binds to AMPA receptors on the postsynaptic
neuron
2
receptors open ion channels that allow sodium into the
postsynaptic neuron
3
The sodium influx depolarizes the cell to some degree.
4
Depolarization affects postsynaptic glutamate NMDA
receptors which control ion channels for calcium
5
The depolarization pops the magnesium gatekeepers out of the ion channels associated with the NMDA receptors, allowing calcium ions to flow into the postsynaptic neuron.
6
Amino Acid-Gated ChannelsGlutamate-Gated ChannelsAMPA, NMDA, kainite
Voltage dependent NMDA channels
How does ionic current flow
through the NMDA-gated channel?
GABA-Gated and Glycine-Gated Channels
GABA mediates inhibitory
transmission
Glycine mediates non-GABA inhibitory
transmission
There are two classes of GABA
receptors: GABAAand GABAB
GABAA are ligand gated ion channels
GABAB are G-protein coupled
receptors
This is interesting because a single molecule binds to
receptors which function in completely different ways
G-Protein-Coupled Receptors and Effectors
Three steps Binding of the
neurotransmitter to the receptor protein
Activation of G-proteins Activation of effector systems
The Basic Structure of G-Protein-Coupled Receptors (GPCRs) Single polypeptide with seven
membrane-spanning alpha-helices
• The Ubiquitous G-Proteins • GTP-binding (G-)
protein• Signal from
receptor to effector proteins
Neurotransmitters Transmit information between neurons Essential link between neurons and effector cells
Signaling pathways Signaling network within a neuron somewhat
resembles brain’s neural network Inputs vary temporally and spatially to increase
and/or decrease drive Delicately balanced Signals regulate signals- drugs can shift the balance
of signaling power
Concluding Remarks
“Dr. Snyder pioneered the labeling of receptors by reversible ligand binding in the identification of opiate receptors and extended this technique to all the major neurotransmitter receptors in the brain.”
He also worked out intracellular messenger systems and established gases (nitric oxide and carbon monoxide) as a new class of neurotransmitters.
http://www.ohsu.edu