Download Neurotransmitter Systems

Neurotransmitter Systems
Jianhong Luo, Ph.D.
Department of Neurobiology
Zhejiang University School of Medicine
Main Reference: Neuroscience Exploring the Brain, 3rd Ed.
By M.F. Bear, B.W. Connors, and M.A. Paradiso
Introduction
Studying Neurotransmitter Systems
Localization of transmitters and synthesizing enzymes
Studying transmitter release / synaptic mimicry / receptors
Neurotransmitter Chemistry
Cholinergic/Catecholaminergic/Serotonergic/mino acidergic neurons
Other neurotransmitter candidates and intercellular messengers
Transmitter-gated Channels
The basic structure of transmitter-gated channels
Amini acid-gated channels
G-Protein-Coupled Receptors And Effectors
The basic structure of G-protein-coupled receptors
The ubiquitous G-protein
G-protein-coupled effector systems
Divergence and Convergence in Neurotransmitter Systems
Introduction
Neurotransmitters (amino acids, amines, and peptides)
Plus
The molecular machinery (for transmitter synthesis, vesicular
packaging, reuptake and degradation, and transmitter action)
Acetylcholine (ACh), the first NT, identified in the 1920s.
The neurons producing and releasing Ach given the term
cholinergic by British pharmacologist Henry Dale (shared the
1936 Nobel Prize with Loewi).
The suffix –ergic: noradrenergic, glutamatergic, GABAergic,
peptidergic, and so on, for the various synapses, neurons and
neurotransmitter systems.
Introduction
Elements of neurotransmitter systems
Studying Neurotransmitter Systems
Criteria to identify a neurotransmitter:
1. The molecule must be synthesized and stored in the
presynaptic neuron.
2. The molecule must be released by the presynaptic axon
terminal upon stimulation.
3. The molecule, when experimentally applied, must produce a
response in the postsynaptic cell that mimics the response
produced by the release of neurotransmitter from the
presynaptic neuron.
Studying Neurotransmitter Systems
Localization of Transmitters and their Synthesis Enzymes
Whatever the inspiration, the first step in confirming the
hypothesis on a new neurotransmitter is to show that the molecule
is, in fact, localized in, and synthesized by, particular neurons.
Many methods have been used to satisfy this criterion for
different neurotransmitters. Two of the most important techniques
used today are immunocytochemistry（免疫细胞化学）and in
situ hybridization （原位杂交）.
Studying Neurotransmitter Systems
Immunocytochemistry. This method uses labeled antibodies
to identify the location of molecules within cells.
Studying Neurotransmitter Systems
 Immunocytochemistry can be used to localize any molecule
for which a specific antibody can be generated, including the
neurotransmitters themselves and the synthesizing enzymes
for transmitter candidates.
Immunocytochemical localization of
a peptide neurotransmitter in neurons
Studying Neurotransmitter Systems
In situ hybridization
Strands of mRNA consist of
nucleotides arranged in a specific
sequence. Each nucleotide will
stick to one other complementary
nucleotide. In the method of in situ
hybridization, a synthetic probe is
constructed containing a sequence
of complementary nucleotides that
will allow it to stick to the mRNA.
If the probe is labeled, the location
of cells containing the mRNA will
be revealed.
Studying Neurotransmitter Systems
In situ hybridization of the mRNA for a neuropeptide
transmitter in hippocampus of mice (A. wild type and B. the
peptide Knockout). Only neurons with the proper mRNA are
labeled, with clusters of white dots.
Studying Neurotransmitter Systems
Studying Transmitter Release
 To show that a neurotransmitter candidate is actually released
upon stimulation.
Axon stimulation → test biological activity → chemical analysis
(as Loewi and Dale did in identification of ACh as a transmitter at
many peripheral synapses)
 A diverse mixture synapses in CNS makes it impossible to
stimulate a single population of synapses. Researchers have to
collect and measure all the chemical mixture (e.g. using brain
slices in a solution containing a high K+ concentration).
 Also have to show Ca2+ dependency of release, and from the
presynaptic axon terminal upon
Studying Neurotransmitter Systems
Studying Synaptic Mimicry
To meet the third criterion, microionophoresis (离子微电泳) is
often use to assess the postsynaptic actions of a transmitter
candidate.
The candidates in solutions in a glass pipette is ejected on in very
small amounts by passing electrical current through the surface of
neurons, and the membrane potential can be measured.
If it mimics the effects of transmitter released at the synapse, and if
the other criteria of localization, synthesis, and release have been
met, then the molecule and the transmitter usually are considered to
be the same chemical.
Studying Neurotransmitter Systems
Microionophoresis
Studying Neurotransmitter Systems
Studying Receptors
As a rule, no two neurotransmitters bind to the same receptor;
however, one neurotransmitter can bind to many different receptors,
receptor subtype. e.g. two different cholinergic receptor subtypes.
Three approaches to study the different receptor subtypes:
Neuropharmacological Analysis.
For instance, cholinergic receptor subtypes respond differently to
various drugs.
Nicotine (烟碱), a receptor agonist in skeletal muscle, nicotinic ACh
receptors (channels). Curare (筒剑毒) is its selective antagonist.
Muscarine (毒蕈碱), a receptor agonist in the heart, muscarinic
receptors (GPCR). Atropine is its selective antagonist
Nicotinic and muscarinic receptors also exist in the brain.
Studying Neurotransmitter Systems
Three subtypes of glutamate receptors at the synaptic excitation in
the CNS: AMPA receptors, NMDA receptors, and kainate receptors,
each named for a different chemical agonist.
The neurotransmitter glutamate activates all the subtypes, but
AMPA acts only at the AMPA receptor…..
Two subtypesthes of NE receptors, α and β, and of GABA receptors,
GABAA and GABAB.
Thus, selective drugs have been extremely useful for categorizing
receptor subclasses. In addition, neuropharmacological analysis has
been invaluable for assessing the contributions of neurotransmitter
systems to brain function.
Studying Neurotransmitter Systems
The neuropharmacology of cholinergic synaptic transmission.
Sites on transmitter receptors can bind either the transmitter itself
(ACh), an agonist that mimics the transmitter, or an antagonist
that blocks the effects of the transmitter and agonists.
Studying Neurotransmitter Systems
The neuropharmacology of glutamatergic synaptic transmission
Three subtypes of glutamate receptors, each of which binds glutamate,
and each of which is activated selectively by a different agonist.
Studying Neurotransmitter Systems
Studying Neurotransmitter Systems
Ligand-Binding Methods.
Selective drugs provide an opportunity to analyze
receptors directly, even before the neurotransmitter
itself had been identified.
Story of discovery of Opiate receptors
Solomon Snyder (1938-) at Johns Hopkins University
Opiates effects on the brain
Hypothesis: opiates are agonists for specific receptors in CNS.
Radioactively labeled opiate compounds
labeled specific sites on some neurons in the brain.
Led to the discovery of opiate receptors, and identification of
endogenous opiates, or endorphins (内啡肽), e.g. enkephalin
Opiate neurotransmitter systems eventually proved .
Studying Neurotransmitter Systems
Opiate receptor binding to a slice of rat brain. Special film was
exposed to a brain section that had radioactive opiate receptor
ligands bound to it. The dark regions contain more receptors.
Studying Neurotransmitter Systems
Molecular Analysis
Enable us to divide the neurotransmitter receptor proteins into
two groups: transmitter-gated ion channels and G-proteincoupled (metabotropic) receptors
The structure of receptor subunits by molecular analysis
presented a broad extent of the diversity in subunit composition
e.g. Each GABA receptor channel requires five subunits, from
five major classes, α, β, γ, δ and ρ, α 1-6 isoforms, β1-4, γ1-4.
Theoretically, there are 151,887 possible combinations and
arrangements of subunits.
What this means?
Neurotransmitter Chemistry
 Evolution is conservative and opportunistic, and it often puts
common and familiar things to new uses.
 Amino acids are essential to life. Most of the known
neurotransmitter molecules are either (1) amino acids, (2)
amines derived from amino acids, or (3) peptides constructed
from amino acids.
 ACh is an exception; but it is derived from acetyl CoA, a
ubiquitous product of cellular respiration in mitochondria, and
choline, important for fat metabolism.
 Amino acid and amine transmitters are generally each stored
in and released by separate sets of neurons, so-called Dale’s
principle. However, many peptide-containing neurons violate
Dale’s principle. Co-transmitters: peptide + amino acid or
peptide + amine
Neurotransmitter Chemistry
Cholinergic Neurons Acetylcholine (ACh)
 The neurotransmitter at NMJ, synthesized by all the motor
neurons in the spinal cord and brain stem. Other cholinergic
cells contribute to the functions of specific circuits in the PNS
and CNS.
 ACh synthesis needs an enzyme, choline acetyltransferase
(ChAT). Only cholinergic neurons contain ChAT, so this
enzyme is a good marker, e.g. antibody-ICC. ChAT transfers
an acetyl group from acetyl CoA to choline .
 Transport of choline into the neuron is the rate-limiting step
in ACh synthesis.
 Acetylcholinesterase (AChE) secreted from Cholinergic and
noncholinergic neurons to degrades ACh. Inhibition of AChE
disrupts transmission at cholinergic synapses on skeletal
muscle and heart muscle.
Neurotransmitter Chemistry
The life cycle of ACh
rate-limiting step
low micromolar
concentrations
with the fastest catalytic
rate. the target of nerve
gases and insecticides
Neurotransmitter Chemistry
Acetylcholine. (a) ACh synthesis. (b) ACh degradation.
Neurotransmitter Chemistry
Catecholaminergic Neurons
The amino acid tyrosine is the precursor for three different
amine neurotransmitters that contain a chemical structure
catechol, collectively called catecholamines (儿茶酚胺).
Include dopamine (DA), norepinephrine (NE), and
epinephrine. Catecholaminergic neurons are found in regions of
the nervous system for regulation of movement, mood,
attention, and visceral function.
Catechol
group
Dopamine
norepinephrine
(nonadrenaline)
epinephrine
adrenaline
Neurotransmitter Chemistry
 All such neurons contain tyrosine hydroxylase (TH), which
catalyzes the first step in catecholamine synthesis, the
conversion of tyrosine to a compound called dopa (Ldihydroxyphenylalanine 二羟苯基丙氨酸).
 The activity of TH is rate limiting for catecholamine
synthesis, regulated by various signals in the cytosol of the
axon terminal (end-product inhibition, increase when [Ca2+]i
elevated by a high rate release).
 Dopa is converted into the neurotransmitter dopamine by the
enzyme dopa decarboxylase. Parkinson’s disease and dopa
supplement therapy.
Neurotransmitter Chemistry
酪氨酸
酪氨酸羟化酶
rate limiting enzyme
二羟苯基丙氨酸
（多巴）
多巴脱羧基酶
多巴胺
多巴胺β羟化酶
located within the
synaptic vesicles
去甲肾上腺素
苯基乙醇胺N-甲基转移酶
肾上腺素
located in the
cytosol
Neurotransmitter Chemistry
 Neurons that use NE as a neurotransmitter contain, in
addition to TH and dopa decarboxylase, the enzyme
dopamine β-hydroxylase (DBH), which converts dopamine
to norepinephrine. DA is transported from the cytosol to the
synaptic vesicles, and there it is made into NE.
 The last in the line of catecholamine neurotransmitters is
epinephrine (adrenaline). Adrenergic neurons contain the
enzyme phentolamine Nmethyltransferase (PNMT), which
converts NE to epinephrine.
 In addition to serving as a neurotransmitter in the brain,
epinephrine is released by the adrenal gland into the
bloodstream. Circulating epinephrine acts at receptors
throughout the body to coordinate visceral response.
Neurotransmitter Chemistry
 The actions of catecholamines in the synaptic cleft are
terminated by selective uptake of the neurotransmitters back
into the axon terminal via Na+-dependent transporters.
 This step is sensitive to a number of different drugs. For
example, amphetamine and cocaine block catecholamine
uptake.
 Once inside, the axon terminal, the catecholamines may be
reloaded into synaptic vesicles for reuse, or they may be
enzymatically destroyed by the action of monoamine oxidase
(MAO), an enzyme found on the outer membrane of
mitochondria.
Neurotransmitter Chemistry
Serotonergic Neurons
 The amine neurotransmitter serotonin, also called 5hydroxytryptamine and abbreviated 5-HT, is derived from the
amino acid tryptophan. Serotonergic neurons are relatively few
in number, but they appear to play an important role in the brain
systems that regulate mood, emotional behavior, and sleep.
 Serotonin synthesis appears to be limited by the availability of
tryptophan in the extracellular fluid bathing neurons. The source
of brain tryptophan is the blood, and the source of blood
tryptophan is the diet.
 5-HT is removed from the synaptic cleft by the action of a
specific transporter, which is sensitive to a number of different
drugs. e.g. antidepressant drugs like fluoxetine. Once back in the
cytosol, 5-HT is either reloaded to SVs or degraded by MAO.
Neurotransmitter Chemistry
色氨酸
色氨酸羟化酶
5-羟色氨酸
5-羟基色氨酸
脱羧酶
5-羟色胺
The synthesis of serotonin from tryptophan.
Neurotransmitter Chemistry
Amino Acidergic Neurons
glutamate (Glu)
Glycine (Gly)
GABA
 Amino acid neurotransmitters Glu, Gly, and GABA serve as
neurotransmitters at most CNS synapses
 Glutamate and glycine are synthesized from glucose and other
precursors by enzymes existing in all cells. Differences among
neurons are quantitative rather than qualitative.
 e.g. the glutamatergic terminals have about 20 mM Glu, only
2-3 times higher than nonglutamatergic cells. Importantly, in
glutamatergic terminalss, but not in other’s, the glutamate
transporter concentrates Glu in SVs to reach about 50 mM.
Neurotransmitter Chemistry
GAD
 GABA is not one of the 20 amino acids used to construct
proteins, it is synthesized in large quantities only by the neurons
that use it as a neurotransmitter.
 The precursor for GABA is glutamate. The key synthesizing
enzyme is glutamic acid decarboxylase (GAD), a good marker
for GABAergic neurons. One chemical step to convert the major
excitatory into the major inhibitory neurotransmitter in the brain!
 The synaptic actions of GABA are terminated by selective
uptake into the terminals and glia via specific Na+-dependent
transporters. Inside the cytosol, GABA is metabolized by the
enzyme GABA transaminase.
Neurotransmitter Chemistry
Other Neurotransmitter Candidates and Intercellular
Messengers
 ATP is concentrated in vesicles at many synapses in the CNS
and PNS, released into the cleft by presynaptic spikes in a Ca2+dependent manner.
 ATP is often packaged in vesicles along with another classic
transmitter (e.g. catecholamine) which means they are probably
co-transmitters.
 ATP directly excites some neurons by gating a cation channel.
ATP binds to purinergic receptors, both transmitter-gated ion
channels and a large class of G-protein coupled purinergic
receptors.
Neurotransmitter Chemistry
 The interesting discovery in the past few
years is that small lipid molecules,
endocannabinoids (大麻酚,endogenous
cannabinoids), can be released from
postsynaptic neurons and act on
presynaptic terminals, called retrograde
signaling; thus, endocannabinoids are
retrograde messengers, a kind of
feedback regulation.
 Vigorous firing in the postsynaptic
neuron → Ca2+ influx throughvoltagegated calcium channels of postsynaptic
neurons→ [Ca2+ ]i increase→ stimulates
the synthesis of endocannabinoid
molecules from membrane lipids.
Retrograde signaling
with endocannabinoids
Neurotransmitter Chemistry
 There are several unusual qualities about endocannabinoids:
1. They are not packaged in vesicles like other neurotransmitters;
instead, they are produced rapidly and on-demand.
2. They are small and membrane permeable; once synthesized,
they can diffuse rapidly across the membrane to contact
neighboring cells.
3. They bind selectively to the CB1 type of cannabinoid
receptor, mainly located on certain presynaptic terminals.
Cannabis
Marijuana
大麻
THC (Δ9-tetrahydrocannabinol)
Short-term effects on brain?
Neurotransmitter Chemistry
 CB1 receptors are GPCRs, and their main effect is often to
reduce the opening of presynaptic calcium channels, and
inhibit release of its neurotransmitter.
 Gaseous molecule, nitric oxide (NO). Carbon monoxide (CO)
has also been suggested as a messenger, being extensively
studied and hotly debated.
 Many of the chemicals we call neurotransmitters may also be
present and function in non-neural parts of the body. (Amino
acids, ATP, Nitric oxide, Ach, serotonin)
Transmitter-gated Channels
 The transmitter-gated ion channels are magnificent tiny
machines.
 A single channel can be a sensitive detector of chemicals and
voltage, it can regulate the flow of surprisingly large currents
with great precision, it can sift and select between very
similar ions, and it can be regulated by other receptor systems.
 Yet each channel is only about 11 nm long, just barely
visible with the best computer-enhanced electron microscopic
methods.
Transmitter-gated Channels
The Basic Structure of Transmitter-Gated Channels
 The most thoroughly studied transmitter-gated ion channel is
the nicotinic ACh receptor at NMJ. Five subunits arrange like a
barrel to form a single pore. Four different subunits α, β, γ, δ
are used. There is one ACh binding site on each of the α
subunits.
 The nicotinic ACh receptor on neurons is also a pentamer, but,
unlike the muscle receptor, most of them are comprised of α
and β subunits only.
 The subunits of GABA- and Glycine-gated channels have a
similar primary structure to the nicotinic ACh receptor, with
four hydrophobic segments to span the membrane, also thought
to be pentameric complexes.
Transmitter-gated Channels
The subunit arrangement of the nicotinic ACh receptor
(a) Side view showing how the four α-helices of each subunit packed together.
(b) Top view showing the location of the two ACh binding sites.
Transmitter-gated Channels
?
Similarities in subunit structure for different transmitter-gated ion channels
(a) They have in common the four regions called M1–M4, which are segments
where the polypeptides will coil into alpha helices to span the membrane. Kainate
receptors are subtypes of glutamate receptors. (b) M1–M4 regions of the ACh
subunit, as they are believed to be threaded through the membrane.
Transmitter-gated Channels
 The glutamate-gated channels are most likely tetramers
structure. The M2 region does not span the membrane, but
instead forms a hairpin that both enters and exits from the
inside of the membrane, resembling potassium channels.
 The purinergic (ATP) receptors also have an unusual structure.
Each subunit has only two membrane-spanning segments, and
the number of subunits of a complete receptor is not known.
 Different transmitter binding sites let one channel respond to
distinct transmitters; certain amino acids around the narrow ion
pore allow only Na+ and K+ to flow through some channels,
Ca2+ through others, and only Cl- through yet others.
Transmitter-gated Channels
Amino Acid-Gated Channels
Amino acid-gated channels mediate most of the fast synaptic
transmission in the CNS.
Several properties of these channels distinguish them from
one another and define their functions within the brain.
 pharmacology
 kinetics
 selectivity
 conductance
All these properties are a direct result of the molecular
structure of the channels.
Transmitter-gated Channels
Glutamate-Gated Channels.
 Three glutamate receptor subtypes: AMPA, NMDA, and
kainate. The AMPA- and NMDA-gated channels mediate the
bulk of fast excitatory synaptic transmission in the brain.
 AMPA-gated channels are permeable to both Na+ and K +, and
most of them are not permeable to Ca2+. The net effect is to
admit Na+ ions into the cell, causing a rapid and large
depolarization.
 NMDA-gated channels differ from AMPA receptors in two
very important ways: (1) NMDA-gated channels are permeable
to Ca2+, and (2) inward ionic current through NMDA-gated
channels is voltage dependent.
Transmitter-gated Channels
The coexistence of NMDA and AMPA receptors in the postsynaptic membrane
of a CNS synapse. (a) An impulse arriving in the presynaptic terminal causes the
release of glutamate. (b) Glutamate binds to AMPA-gated and NMDA-gated
channels in the postsynaptic membrane. (c) The entry of Na through the AMPA
channels, and Na and Ca2 through the NMDA channels, causes an EPSP.
Transmitter-gated Channels
Inward ionic current through the
NMDA-gated channel.
(a) Glutamate alone causes the channel
to open, but at the resting membrane
potential, the pore becomes blocked by
Mg2+ ions.
(b) Depolarization of the membrane
relieves the Mg2+ block and allows Na+
and Ca2+ to enter.
Transmitter-gated Channels
 It is hard to overstate the importance of intracellular Ca2+ to cell
functions. presynaptical and postsynaptical; physiological and
pathological. Thus, activation of NMDA receptors can, in
principle, cause widespread and lasting changes in the
postsynaptic neuron.
 The magnitude of this inward Ca2+ and Na+ through NMDAgated channels depends on the postsynaptic membrane potential
in an unusual way, for an unusual reason. “magnesium block”,
voltage dependent release. Both glutamate and depolarization
must coincide before the channel will pass current. This
property has a significant impact on synaptic integration at
many locations in the CNS.
Transmitter-gated Channels
GABA-Gated and Glycine-Gated Channels.
GABA mediates most synaptic inhibition in the CNS, and
glycine mediates most of the rest. Both are chloride channels.
Synaptic inhibition must be tightly regulated in the brain. Too
much causes a loss of consciousness and coma; too little leads to
a seizure.
 It is why the GABAA receptor has several other sites where
chemicals can dramatically modulate its function. e.g.
Benzodiazepines（苯二氮卓）and barbiturates （巴比妥）.
When GABA is present, increase the frequency, or the duration
of channel openings, respectively, thus, more Cl- current, stronger
IPSPs, enhanced behavior inhibition. And selective for GABAA
receptor, no effect on glycine receptor.
Transmitter-gated Channels
 Ethanol, another popular drug, strongly enhances GABAA
receptor function in a way of dependence on the receptor
specific structure. Ethanol has also complex effects on
NMDA, glycine, nicotinic ACh, and serotonin receptors.
 What is the endogenous ligands for these drug binding sites?
They may serve as regulators of inhibition. Substantial
evidence indicates that natural benzodiazepine-like ligands
exist. Other good candidates as natural modulators of
GABAA receptors are the neurosteroids (神经甾体), natural
metabolites of steroid hormones, but also in glial cells of the
brain. Some neurosteroids enhance inhibitory function while
others suppress it, and they seem to do so by binding to their
own site on the GABAA receptor.
Transmitter-gated Channels
The binding of drugs to the GABAA receptor
The drugs by themselves do not open the channel, but change the
effect when GABA binds to the channel at the same time as the drug.
G-Protein Coupled Receptors and Effectors
Transmission at GPCRs involves three steps: (1) binding of the
neurotransmitter to the receptor protein, (2) activation of Gproteins, and (3) activation of effector systems.
The Basic Structure of GPCRs
 A family members (about 100), a single polypeptide, seven
membrane-spanning α-helices, two extracellular loops
(binding sites). Two intracellular loops (binding to and
activating G-proteins)
 Structural variations at these two sites determine which
agonist binding and which G-proteins and effector systems
activated in response to transmitter binding.
Transmitter-gated Channels
Transmitter-gated Channels
The basic structure of a G-protein-coupled receptor.
Most metabotropic receptors have seven membrane-spanning αhelices, a transmitter binding site on the extracellular side, and a Gprotein binding site on the intracellular side.
G-Protein Coupled Receptors and Effectors
The Ubiquitous G-Proteins
 G-proteins are the common link signaling pathways; GTP
binding protein, about 20 family members; Less than
transmitter receptors.
 The same basic mode of operation:
1. Each has three subunits, α, β, and γ. In the resting state, GDP
is bound to the Gα.
2. If this G-protein hits the proper receptor with a transmitter
bound , then releases its GDP and exchanges it for a GTP.
3. The activated G-protein splits into 2 parts: the Gα plus GTP,
and the Gβγ complex. Both can influence various effectors.
4. The Gα is itself an enzyme that eventually breaks down GTP
into GDP, and terminates its own activity.
5. The Gα and Gβγ subunits come back together, allowing the
cycle to begin again.
G-Protein Coupled Receptors and Effectors
The basic mode of operation of G-proteins
(a) In its inactive state, the α subunit of the G-protein binds GDP. (b) When
activated by a G-protein-coupled receptor, the GDP is exchanged for GTP. (c) The
activated G-protein splits, and both the Gα (GTP) subunit and the Gβγ subunit
become available to activate effector proteins. (d) The G subunit slowly removes
phosphate (PO4) from GTP, converting GTP to GDP and terminating its own activity.
G-Protein Coupled Receptors and Effectors
G-Protein-Coupled Effector Systems
Activated G-proteins exert their effects by binding to either of:
G-protein-gated ion channels and G-protein-activated enzymes.
The Shortcut Pathway:
 A variety of neurotransmitters use the shortcut pathway, from
receptor to G-protein to ion channel. e.g. ① the muscarinic
receptors in the heart to explain why ACh slows the heart rate.
② neuronal GABAB receptors.
 The fastest of the G-protein-coupled systems (30–100 msec)
since no intermediary between receptor and channel. And also
very localized since it is within the membrane and cannot
move very far.
G-Protein Coupled Receptors and Effectors
The shortcut pathway.
(a) G-proteins in heart
muscle are activated by
ACh binding to
muscarinic receptors.
(b) The activated G
subunit directly gates a
potassium channel.
G-Protein Coupled Receptors and Effectors
Second Messenger Cascades.:
 G-proteins can also directly activate certain enzymes and the
laters trigger an elaborate series of biochemical reactions, a
cascade that often ends in the activation of other “downstream”
enzymes that alter neuronal function.
 Between the first enzyme and the last are several second
messengers. The whole process that couples the
neurotransmitter, via multiple steps, to activation of a
downstream enzyme is called a second messenger cascade.
 the cAMP second messenger cascade initiated by the
activation of the NE β receptor.
G-Protein Coupled Receptors and Effectors
The components of a second messenger cascade
G-Protein Coupled Receptors and Effectors
 Many biochemical processes are regulated with a push-pull
method, one to stimulate them and one to inhibit them, and
cAMP production is no exception.
 The activation of NE α2 receptor leads to the activation of Gi,
which suppresses the activity of adenylyl cyclase.
 Some messenger cascades can branch. e.g. G-proteins →
PLC → PIP2 → DAG + IP3. DAG (within the membrane) →
PKC; IP3 (water-soluble) → IP3 receptor (IP3-gated calcium
channels) on the smooth ER and other organelles → discharge
of stored Ca2+. Elevation in cytosolic Ca2+ can trigger
widespread and long-lasting effects. One effect is activation
of CaMK, an kinase implicated in memory.
G-Protein Coupled Receptors and Effectors
The stimulation and inhibition of adenylyl cyclase by different
G-proteins. (a) Binding of NE to the β receptor activates Gs, which
in turn activates adenylyl cyclase. Adenylyl cyclase generates cAMP,
which activates the downstream enzyme protein kinase A. (b)
Binding of NE to the α2 receptor activates Gi, which inhibits
adenylyl cyclase.
G-Protein Coupled Receptors and Effectors
Second messengers generated by the breakdown of PIP2, a
membrane phospholipid. ➀ Activated G-proteins stimulate
the enzyme phospholipase C (PLC). ➁ PLC splits PIP2 into DAG
and IP3. ➂ DAG stimulates the downstream enzyme protein kinase
C (PKC). ➃ IP3 stimulates the release of Ca2+ from intracellular
stores. The Ca2+ can go on to stimulate various downstream enzymes.
G-Protein Coupled Receptors and Effectors
Phosphorylation and Dephosphorylation:
 key downstream enzymes in many of the second messenger
cascades are protein kinases (PKA, PKC, CaMK), that
transfer phosphate from ATP to proteins (phosphorylation), It
changes the protein’s conformation slightly, thereby changing
its biological activity. e.g. ion channels.
 NE β receptors on cardiac muscle cells → rise in cAMP
activates PKA → phosphorylation of voltage-gated calcium
channels → more Ca2+ influx → heart beats more strongly.
 By contrast, the β-adrenergic receptors in neurons inhibits
certain potassium channels. Reduced K+ conductance causes a
slight depolarization, making the neuron more excitable.
G-Protein Coupled Receptors and Effectors
 Protein phosphatases act rapidly to remove phosphate groups.
The degree of channel phosphorylation depends on the
dynamic balance of phosphorylation by kinases and
dephosphorylation by phosphatases.
Protein phosphorylation and dephosphorylation.
G-Protein Coupled Receptors and Effectors
The Function of Signal Cascades.
 Transmission using transmitter-gated channels is simple and
fast, while that involving GPCRs is complex and slow.
 The advantages of having such long chains of command:
1. one is signal amplification: The activation of one GPCR can
lead to the activation of not one, but many, ion channels.
2. the use of small messengers that can diffuse quickly (such as
cAMP) allows signaling at a longer distance.
3. provide any sites for further regulation, as well as interaction
between cascades.
4. Finally, generate very long-lasting chemical changes in cells,
which may form the basis for a lifetime of memories.
G-Protein Coupled Receptors and Effectors
Signal amplification by
G-protein coupled second
messenger cascades
When a transmitter activates
a GPCR, there can be
amplification of the
messengers at several stages
of the cascade, so that
ultimately many channels
are affected.
Divergence and Convergence in Neurotransmitter Systems
 The ability of one transmitter to activate more than one subtype
of receptor, and cause more than one type of postsynaptic
response, is called divergence (幅散). e.g. Glutamate and
GABA. Divergence is the rule among neurotransmitter systems.
It even occurs at points beyond the receptor level, depending on
which G-proteins and which effector systems are activated.
 Neurotransmitters can also exhibit convergence (会聚) of
effects. Multiple transmitters can converge to affect the same
effector systems at the level of the G-protein, the second
messenger cascade, or the type of ion channel.
 Neurons integrate divergent and convergent signaling systems,
resulting in a complex map of chemical effects.
Divergence and Convergence in Neurotransmitter Systems
Divergence and convergence in
neurotransmitter signaling
systems.
(a) Divergence.
(b) Convergence.
(c) Integrated divergence and
convergence.
扩展阅读及章后思考题
Box 6.1 Pumping Ions and Transmitters
Box 6.2 This is your brain on Endocannabinoids.
Box 6.3 Deciphering the language of neurons.
Box 6.4 The Brain’s exciting Poisons
谢谢！