Molecular and Cellular Neurobiology
Lecture 1: Ultrastructure of the mammalian synapse
Molecular mechanisms of neuronal communication (By Erik Storkebaum)
Introduction
➢ Synapse: specialized cell-cell junction that allows neurons to communicate with each other
and non-neuronal target cells (e.g. muscle cells). So the synapses are the structures that
mediate neuronal communication.
➢ Synapse (2 types) → chemical: neurotransmitter-based. So use neurotransmitters to
c communicate.
→ electrical: electrically coupled -> way of communicating.
➢ Neural circuit: multiple neurons linked together by a set of synapses in series.
Number and diversity of synapses
➢ A typical neuron in a mammalian brain makes on average of 10.000 synapses with other
neurons.
➢ The adult human brain is estimated to contain > 1014 synapses. Because of this enormity the
human brain is able to do what is can do. For example perceive, interpret the world and be
creative and respond to stimuli and think. In order to do all this you need lots of neurons and
also lots of synapses. Not every synapse in the brain is equivalent and different kind of
synapses can be found. For example the neurotransmitter that is used and the postsynaptic
response.
➢ Diversity based on neurotransmitter use:
o E.g. glutamate, GABA, glycine, dopamine, acetylcholine, (nor)epinephrine, serotonin,
etc.
o Co-transmission may occur (single synapse uses more than one neurotransmitter,
these two can even be accumulated in same synaptic vesicle. For example GABA
(inhibitory) together with glycine (inhibitory))
➢ Diversity bases on postsynaptic response:
o Excitatory (e.g. glutamate): postsynaptic depolarization (activate post-synaptic
neurons). So briefly what happens is that the presynaptic neurons will secrete the in
this case excitatory neurotransmitter. This will bind to its receptor, which is located
at the postsynaptic site. This receptor is actually an ion channels that is gated by the
neurotransmitter. So a ligand gated ion channel. So when neurotransmitter binds the
channel will open and in the case of excitatory neurotransmitter and their receptors
there will be an influx of positively charged ions (cations) and this will lead to
depolarization of the postsynaptic cells.
o Inhibitory (e.g. GABA, glycine): postsynaptic hyperpolarization (reduce possibility
that action potential will happen in postsynaptic neuron). Inhibitory
neurotransmitters induce postsynaptic hyperpolarization. This is because the
postsynaptic receptors that they bind and activate are channels that conduct
negatively charged ions (cations -> chloride) and this will induce hyperpolarization of
postsynaptic cells and reduce likelihood that action potential is generated.
o Neuromodulatory (e.g. dopamine, serotonin): no ionotropic receptors, induce
biochemical changes in postsynaptic neurons. They discriminate themselves from
excitatory and inhibitory synapses, because the receptors for those
neurotransmitters are not ionotropic meaning they are not ligand gated ion channels
, but are so called metabotropic receptors. Usually g protein coupled, which are not
channels but rather induce biochemical changes in postsynaptic neurons.
➢ An additional way in which you could distinguish synapses is based on anatomical location:
central (between neurons in central nervous system) versus peripheral (outside central
nervous system) synapse (e.g. neuro-muscular junction (synapses between motor-neuron),
neuro-endocrine junction (autonomic neurons and glands), autonomous neuro-effector
junctions(autonomous nerves and e.g. vesicle wall))
➢ Start course with talking about: Chemical synapse → structural aspects
→ functional aspects
Overview: The synaptic vesicle cycle
Presynaptic neuron is loaded with neurotransmitter. Postsynaptic neuron has receptors → post
synaptic density (PSD). The active zone is a specialized region of the membrane where the vesicles
are docking, priming and fusion. With fusion the neurotransmitter is released. You also see a calcium
channel. These are voltage gated calcium channels and will open when an action potential comes in
and the voltage over the membrane changes. This will trigger the exocytosis of the neurotransmitter.
After the neurotransmitter is secretes, the synaptic vesicle will collapse and recycled via endocytosis.
When this happens the vesicle is acidified and neurotransmitter uptake will happen again. (last year)
In the overview you see the schematic representation of a synapse. With on the hand a presynaptic
neuron and on the other hand a postsynaptic cell. Let’s start with the postsynaptic cell, because it is
the least complicated. What you see in the figure is that the postsynaptic cell expresses receptors
which are clustered at the synapse, which is called the post synaptic density (PSD). Those receptors
will bind the neurotransmitter that is secreted from the presynaptic nerve terminal, resulting in
activation of the receptor, meaning that the channel opens for ions, resulting in a postsynaptic
response. Then the presynaptic cell contains neurotransmitter. This neurotransmitter is concentrated
in synaptic vesicles. These are membrane bound organelles and within the synaptic vesicle the
neurotransmitter is secreted by the presynaptic neuron. How does this neurotransmitter get inside
the synaptic vesicle? It is pumped in the synaptic vesicle by a so called neurotransmitter transporter
,(more about this in chapter 7). Essentially there are 2 types of transporters: one on the synaptic
vesicles and the other type in the membrane (pre or post synaptic membrane). The one in the
membrane is responsible for the reuptake of neurotransmitter and the one on the synaptic vesicle
will be responsible for neurotransmitter uptake in the synaptic vesicles. Once the neurotransmitter is
taken up there will be a fraction of the synaptic vesicle that will be docked to the active zone (Zone in
presynaptic nerve ending, opposing PSD, and where all the machinery is present for
neurotransmitter release). There are different steps: first there is the docking of the synaptic vesicle,
then the priming and then you get fusion of the synaptic vesicle with the presynaptic membrane. This
only happens when the presynaptic neuron was activated, which will result in the influx of calcium.
(more about this in next lectures)
But shortly what happens: If the presynaptic neurons is activated, it generates an action potential.
This action potential is conducted over the membrane of the neuron and will arrive over the axon to
the presynaptic nerve terminal. When it gets there, it will activate the voltage gated Ca2+ channels.
With depolarization these channels will open and there will be a local increase in the calcium
concentration at the presynaptic site. This will trigger the exocytosis of the synaptic vesicles, which
will collapse at the presynaptic membrane and release the neurotransmitter they contain, which will
defuse to the postsynaptic membrane and bind and activate the receptors there. Not too long after
the exocytosis (and thus enlarging the membrane), there will be endocytosis, so recycling of synaptic
vesicles. The first step of the recycling is the endocytosis (more detail chapter 5). What you see here
is clathrin indicated endocytosis, which is typical for this. The next thing that happens is that the
vesicle gets acidified. This is done by a proton pump which is present in the synaptic vesicle
membrane. This acidification is essential to allow later on the uptake of neurotransmitter in the
synaptic vesicle. The acidification represents the fuel (chemical gradient) that allows the pumping of
the neurotransmitter into the synaptic vesicle by the neurotransmitter transporter.
Chapter 1: Ultrastructure of the mammalian synapse
- Central synapse: where the axon of a presynaptic neuron contacts the dendrite of a
postsynaptic neuron. Not necessarily dendrite needs to be contacted, can also be cell body
or axon, but mostly dendrites.
- Type I = asymmetric synapse: excitatory; mainly on dendrites and dendritic spines
- Type II = symmetric synapse (no dendritic spine): inhibitory: concentrate on cell soma and
axonal initial segment
Fig. 1/B-> type I (sp = spine), F2 -> type II
synaptic vesicles – mitochondrion – active zone (presynaptic) – postsynaptic density – dense material
in synaptic cleft – spine apparatus – spine neck – dendritic microtubules – “stalk” of axon active zone
= specialized region on the presynaptic plasma membrane, where synaptic vesicles are docked and
primed for release; aligned with postsynaptic density (PSD)
- Active zone (AZ): specialized region on the presynaptic plasma membrane, where synaptic
vesicles are docked and primed for release; the AZ is aligned with the postsynaptic density
(PSD)
, - Molecular composition of PSD (more in chapter 8) (both present in pre and postsynaptic
neuron): neurotransmitter receptors, transsynaptic adhesion molecules, scaffolding
molecules (protein involved in keeping other proteins together e.g. receptors together etc.
for proper functioning), signal transduction molecules
3D reconstruction of axons and dendritic spines
Figure B on the right:
Reconstructing of axons (white) and part of the axons,
red, is where the synapse is located. In green
reconstruction of dendrite and this dendrite has
enlargements, boutons, and there the axons synapse.
What you can distinguish are boutons that have more
than one synaptic input (more than one axon) =
multisynaptic boutons. Boutons with one synapse is
single synaptic bouton. Non synaptic boutons used to be
boutons, but no synapse here. Synapses are
continuously modified in strength and structurally
modulated and new synapses can be formed and
existing synapses be lost → for example basis of learning.
mito: mitochondrion; mvb: multivesicular body; SSB: single synaptic bouton (single postsynaptic
partner); NSB: nonsynaptic bouton (no postsynaptic partner); MSB: multisynaptic bouton (more than
one postsynaptic partner); dcv: dense-core vesicle. vesicles are yellow
Figure A: Dendrite with dendritic spines. Spines are uniform in shape compared to figure B that show
more variation. A are more mushroom spines, more homogenous and bigger. Cerebellar Purkinje cell
dendrite; dendritic spines have relatively uniform shape; sp: spine. Figure B: Hippocampal CA1
dendrite; f: filopodia-like protrusion; st: stubby protrusion; m: mushroom-shaped spine; t: thin-
shaped spine. Red surfaces are PSD areas.
Which of the following are neuromodulatory neurotransmitters?
A GABA (→ inhibitory)
B glutamate (→ excitatory)
C dopamine (→ neuromodulatory)
D glycine (→ inhibitory)
,Lecture 2
Chapter 2: Synaptic cell adhesion molecules (SCAMs)
Learning outcomes:
➢ Deduce different functions of SCAMs
➢ Describe different stages of synapse formation and function + role of SCAMs in each stage
➢ Give examples of SCAMS and their functional domains
➢ Schematically draw SCAMs to illustrate function (currently not possible → images are shown)
Synaptic cell adhesion
Synaptic cell adhesion molecules (SCAMs) could be defined as transmembrane proteins (or at least
proteins that are linked to the membrane) which are localized at the synapse (either presynaptic or
postsynaptic, or both) and interact with each other across the synapse (transsynaptic).
➢ Synaptic junctions are organized by trans-synaptic cell adhesion molecules bridging the
synaptic cleft.
In the figure (representation of synapse. Synaptic cleft is between the presynaptic and postsynaptic
membrane) a schematic representation of transsynaptic adhesion molecules. The red molecules are
linked to the upper membrane. These are two cell adhesion molecules that are linked to the
presynaptic membrane and in green are SCAMs that are connected with the postsynaptic membrane.
Why do you know the upper one is presynaptic? → above you see synaptic vesicles, one docked and
primed and one not. So if an action potential arrives here, neurotransmitter from his vesicle will
release. What you further see are voltage gated calcium channels, which means that when an action
potential arrives, there is depolarization of the membrane and this will change the conformation of
the channels, which makes them open. Calcium can now flow in the presynaptic nerve terminal,
which triggers the exocytosis of the synaptic vesicle. The neurotransmitter that would then be
released will bind to the receptors on postsynaptic receptors. These receptors are ligand gated,
meaning that when the transmitter binds the receptor will change conformation and act as an ion
channel to conduct ions that now will flow into postsynaptic cell.
, The function of the cell adhesion molecules is on the one hand to link the post and
presynaptic membranes together so that the synaptic cleft has a given width. Because of the
interaction between the molecules is a standardized distance, which is the proper distance
for neurotransmission to occur. The neurotransmitter that is released needs to travel a given
distance to the postsynaptic receptors by diffusion ,which will result in a certain rate of
activation of the postsynaptic receptors. This is one function of the SCAMs → physically link
pre and postsynaptic site together and bridge the cleft:
➢ Synaptic junctions are organized by trans-synaptic cell adhesion molecules bridging the
synaptic cleft.
SCAMs also have other functions and important is to know that they are transmembrane
proteins, meaning they also have a short part of the protein at the intracellular site, where
they can interact with other intracellular proteins. This way they are involved in organizing
synapses. For example they can make sure that the calcium channels are clustered at
presynaptic membrane or synaptic vesicles are docked. In short they would organize the
presynaptic active zone. Similarly the postsynaptic cell adhesion molecules could interact
with neurotransmitter receptors in order to cluster them at postsynaptic site or interact with
signal transducing or scaffolding proteins. Together this would anchor the postsynaptic
density, which is a collection/dense concentration of proteins. :
➢ Synaptic cell adhesion molecules (SCAMs) not only connect pre- and postsynaptic
compartments, but also mediate trans-synaptic recognition and signaling processes that are
essential for the establishment, specification and plasticity of synapses.
➢ Examples of SCAMs:
o Neurexin
o neuroligins
o Ig domain proteins (e.g. synCAMs)
o Receptor phosphotyrosine kinases and phosphatases (first phosphorylates proteins
and second one dephosphorylates proteins)
o Leucine rich repeat proteins
First two implicated in schizophrenia and autism
Synaptic plasticity: the capacity of synapses to adjust their properties in response to previous activity
Four stages of synapse formation and function
1) First stage is establishment of synapse. When synapse is established there will be a
presynaptic axon nerve ending and a postsynaptic partner (e.g. dendritic spine). The pre- and
postsynaptic partners will approximate each other and they see if they fit together. This is
done on molecular level by synaptic cell adhesion molecules. These interact with each other.
This is also a distinction that is made between SCAMs that interact with each other but are
different (e.g. neuroligins and neurexins) → heterophilic interactions. When same molecule
is expressed pre and post → homophylic interaction. So contact establishment involves
recognition of pre- and postsynaptic neurons; this process may require heterophilic and
homophylic (between identical molecules) interactions between SCAMs to recognize
appropriate synaptic partners
2) Second phase is the assembly stage. During this stage the pre and post synaptic part
components are being assembled, such as the synaptic vesicles that are collected and
recruited to the presynaptic site. And in general the presynaptic active zone is recruited,
including the calcium channels. Also postsynaptic, the PSD is recruited including
neurotransmitter receptors and signaling and scaffolding molecules. The intracellular part of
the SCAMs will interact with other proteins that influence for instance the vesicles or calcium
channels or neurotransmitter receptors.
, So recruitment of synaptic
vesicles, AZs and PSDs; during this
stage, SCAMs regulate physical
cell-cell adhesion and serve as
anchor proteins to cluster or
recruit receptors or components
of the pre- and postsynaptic
machinery
3) Third stage is called the
specification stage. Once all the
components of the pre and
postsynaptic synapse have been
recruited, the synapse can start
functioning. Meaning when an
action potential reaches the
presynaptic nerve terminal, there
will be secretion of
neurotransmitters by synaptic
vesicle exocytosis, resulting in
activation of postsynaptic
receptors, which will result in
activation of postsynaptic cells.
The cell adhesion molecules are
during this stage responsible for
organizing the molecular
components of the synapse
(similar to stage 2 for SCAMs).
With all this the synapse now
functioning.
So functional specification stage:
organization of the molecular
components of the synapse, resulting in functionality of the synapse.
4) The fourth stage is the plasticity stage. Synaptic plasticity is essentially the adaptation of the
strength of the synapse based on previous activity. This adaptation of the synaptic efficacy
can involve either structural or functional changes. The cell adhesion molecules are also
involved in this stage. For instance if synaptic strength wishes to be increased, more vesicles
are recruited at synaptic site so more neurotransmitter release. Or they recruit more
postsynaptic receptors, which results in a higher chance that postsynaptic cells gets activated
in response to the neurotransmitter.
So synaptic plasticity: SCAMs may contribute to structural and functional synaptic plasticity in
activity-dependent adaptive events
Note: Once synapses have been established, it are not static structures. Synapses continue to be
formed and eliminated during the lifetime of an organism.
It is thought that learning and memory is based on this plastic capacity of our brain.
Functional domains of SCAMs
How do SCAMs function? On the one hand they need to interact with other cell adhesion molecules
in order to form trans-synaptic connections. The SCAMs do this through well defined protein
functional domains. Functional domains of a protein are domains that mediate a certain function of
the protein. In this case there is a limited number of functional domains that is found in different
synaptic cell adhesion molecules that mediate the adhesive function of those proteins. These
functional domains are often assembled in repeat units:
, ➢ The adhesive function of SCAMs is based on a limited number of extracellular domains, often
assemble intro repeat units:
o Examples of such domains are:
▪ Immunoglobulin (Ig) domains: most frequently bind to other lg domains;
most SCAMs containing lg domains also contain fibronectin III (FNIII)
domains.
▪ Cadherin domains: always occur in multiple copies connected by a linker that
bind 2-3 calcium ions. Therefore, cadherin mediated interaction are calcium
dependent and often homophylic.
▪ Lamin A, neurexin, and sex hormone binding protein (LNS -> mediate
interaction between SCAMs)) domains (aka laminin G-Like (LG) domains)
▪ Leucine rich repeats (LRRs)
Example neurexin-neuroligin interactions
ɑ-Neurexin has an LNS domain that interacts with
the Neuroligin molecule. The fact that those two
molecules form a dimer is mediated by an
esterase-domain of neuroligin. So that’s how the
two partners in the dimer recognize and interact
with each other.
(last year) Synaptic vesicle docked to presynaptic
membrane. There are cell adhesion molecules and
usually have a intracellular portion (short!) and an
extracellular portion. There are pre and post
synaptic adhesion molecules. They are to keep the
pre and post synaptic membrane together. This
for faster transmission. The molecules also are anchors to which other molecules can attach. Even
though both are different molecules they both have a protein domain at which they recognize each
other. The intracellular portions also connect to other proteins to activate them and can cause a
cascade
➢ Neurexins:
o Presynaptic single-pass membrane proteins with a large extracellular sequence and a
short cytoplasmic tail.
o Vertebrates have 3 neurexin genes (Nrx1-3); each gene produces from independent
promoters a larger ɑ- and a shorter β-neurexin isoform. (also seen in figure above: ɑ
larger than β)
o Extracellularly, ɑ-neurexins contain 6 LNS domain with 3 interspersed EGF-like
domains; β-neurexins only contain a single LNS domain.
o Neurexins are extensively alternatively spliced at 5 conserved sites in their
extracellular region, creating potentially thousands of isoforms (lots of forms of the
protein). This may constitute a recognition code (allows the cell to recognize the
appropriate post synaptic partner).
o Intracellularly, the short cytoplasmic tails of neurexins contain PDZ-domain (protein
interaction domain, which interacts with other proteins) binding sequences
(sequence in protein that binds other proteins that contain a PDZ domain) that bind
to intracellular proteins, e.g. CASK and protein 4.1
➢ Neuroligins
o Postsynaptic single-pass membrane proteins
Lecture 1: Ultrastructure of the mammalian synapse
Molecular mechanisms of neuronal communication (By Erik Storkebaum)
Introduction
➢ Synapse: specialized cell-cell junction that allows neurons to communicate with each other
and non-neuronal target cells (e.g. muscle cells). So the synapses are the structures that
mediate neuronal communication.
➢ Synapse (2 types) → chemical: neurotransmitter-based. So use neurotransmitters to
c communicate.
→ electrical: electrically coupled -> way of communicating.
➢ Neural circuit: multiple neurons linked together by a set of synapses in series.
Number and diversity of synapses
➢ A typical neuron in a mammalian brain makes on average of 10.000 synapses with other
neurons.
➢ The adult human brain is estimated to contain > 1014 synapses. Because of this enormity the
human brain is able to do what is can do. For example perceive, interpret the world and be
creative and respond to stimuli and think. In order to do all this you need lots of neurons and
also lots of synapses. Not every synapse in the brain is equivalent and different kind of
synapses can be found. For example the neurotransmitter that is used and the postsynaptic
response.
➢ Diversity based on neurotransmitter use:
o E.g. glutamate, GABA, glycine, dopamine, acetylcholine, (nor)epinephrine, serotonin,
etc.
o Co-transmission may occur (single synapse uses more than one neurotransmitter,
these two can even be accumulated in same synaptic vesicle. For example GABA
(inhibitory) together with glycine (inhibitory))
➢ Diversity bases on postsynaptic response:
o Excitatory (e.g. glutamate): postsynaptic depolarization (activate post-synaptic
neurons). So briefly what happens is that the presynaptic neurons will secrete the in
this case excitatory neurotransmitter. This will bind to its receptor, which is located
at the postsynaptic site. This receptor is actually an ion channels that is gated by the
neurotransmitter. So a ligand gated ion channel. So when neurotransmitter binds the
channel will open and in the case of excitatory neurotransmitter and their receptors
there will be an influx of positively charged ions (cations) and this will lead to
depolarization of the postsynaptic cells.
o Inhibitory (e.g. GABA, glycine): postsynaptic hyperpolarization (reduce possibility
that action potential will happen in postsynaptic neuron). Inhibitory
neurotransmitters induce postsynaptic hyperpolarization. This is because the
postsynaptic receptors that they bind and activate are channels that conduct
negatively charged ions (cations -> chloride) and this will induce hyperpolarization of
postsynaptic cells and reduce likelihood that action potential is generated.
o Neuromodulatory (e.g. dopamine, serotonin): no ionotropic receptors, induce
biochemical changes in postsynaptic neurons. They discriminate themselves from
excitatory and inhibitory synapses, because the receptors for those
neurotransmitters are not ionotropic meaning they are not ligand gated ion channels
, but are so called metabotropic receptors. Usually g protein coupled, which are not
channels but rather induce biochemical changes in postsynaptic neurons.
➢ An additional way in which you could distinguish synapses is based on anatomical location:
central (between neurons in central nervous system) versus peripheral (outside central
nervous system) synapse (e.g. neuro-muscular junction (synapses between motor-neuron),
neuro-endocrine junction (autonomic neurons and glands), autonomous neuro-effector
junctions(autonomous nerves and e.g. vesicle wall))
➢ Start course with talking about: Chemical synapse → structural aspects
→ functional aspects
Overview: The synaptic vesicle cycle
Presynaptic neuron is loaded with neurotransmitter. Postsynaptic neuron has receptors → post
synaptic density (PSD). The active zone is a specialized region of the membrane where the vesicles
are docking, priming and fusion. With fusion the neurotransmitter is released. You also see a calcium
channel. These are voltage gated calcium channels and will open when an action potential comes in
and the voltage over the membrane changes. This will trigger the exocytosis of the neurotransmitter.
After the neurotransmitter is secretes, the synaptic vesicle will collapse and recycled via endocytosis.
When this happens the vesicle is acidified and neurotransmitter uptake will happen again. (last year)
In the overview you see the schematic representation of a synapse. With on the hand a presynaptic
neuron and on the other hand a postsynaptic cell. Let’s start with the postsynaptic cell, because it is
the least complicated. What you see in the figure is that the postsynaptic cell expresses receptors
which are clustered at the synapse, which is called the post synaptic density (PSD). Those receptors
will bind the neurotransmitter that is secreted from the presynaptic nerve terminal, resulting in
activation of the receptor, meaning that the channel opens for ions, resulting in a postsynaptic
response. Then the presynaptic cell contains neurotransmitter. This neurotransmitter is concentrated
in synaptic vesicles. These are membrane bound organelles and within the synaptic vesicle the
neurotransmitter is secreted by the presynaptic neuron. How does this neurotransmitter get inside
the synaptic vesicle? It is pumped in the synaptic vesicle by a so called neurotransmitter transporter
,(more about this in chapter 7). Essentially there are 2 types of transporters: one on the synaptic
vesicles and the other type in the membrane (pre or post synaptic membrane). The one in the
membrane is responsible for the reuptake of neurotransmitter and the one on the synaptic vesicle
will be responsible for neurotransmitter uptake in the synaptic vesicles. Once the neurotransmitter is
taken up there will be a fraction of the synaptic vesicle that will be docked to the active zone (Zone in
presynaptic nerve ending, opposing PSD, and where all the machinery is present for
neurotransmitter release). There are different steps: first there is the docking of the synaptic vesicle,
then the priming and then you get fusion of the synaptic vesicle with the presynaptic membrane. This
only happens when the presynaptic neuron was activated, which will result in the influx of calcium.
(more about this in next lectures)
But shortly what happens: If the presynaptic neurons is activated, it generates an action potential.
This action potential is conducted over the membrane of the neuron and will arrive over the axon to
the presynaptic nerve terminal. When it gets there, it will activate the voltage gated Ca2+ channels.
With depolarization these channels will open and there will be a local increase in the calcium
concentration at the presynaptic site. This will trigger the exocytosis of the synaptic vesicles, which
will collapse at the presynaptic membrane and release the neurotransmitter they contain, which will
defuse to the postsynaptic membrane and bind and activate the receptors there. Not too long after
the exocytosis (and thus enlarging the membrane), there will be endocytosis, so recycling of synaptic
vesicles. The first step of the recycling is the endocytosis (more detail chapter 5). What you see here
is clathrin indicated endocytosis, which is typical for this. The next thing that happens is that the
vesicle gets acidified. This is done by a proton pump which is present in the synaptic vesicle
membrane. This acidification is essential to allow later on the uptake of neurotransmitter in the
synaptic vesicle. The acidification represents the fuel (chemical gradient) that allows the pumping of
the neurotransmitter into the synaptic vesicle by the neurotransmitter transporter.
Chapter 1: Ultrastructure of the mammalian synapse
- Central synapse: where the axon of a presynaptic neuron contacts the dendrite of a
postsynaptic neuron. Not necessarily dendrite needs to be contacted, can also be cell body
or axon, but mostly dendrites.
- Type I = asymmetric synapse: excitatory; mainly on dendrites and dendritic spines
- Type II = symmetric synapse (no dendritic spine): inhibitory: concentrate on cell soma and
axonal initial segment
Fig. 1/B-> type I (sp = spine), F2 -> type II
synaptic vesicles – mitochondrion – active zone (presynaptic) – postsynaptic density – dense material
in synaptic cleft – spine apparatus – spine neck – dendritic microtubules – “stalk” of axon active zone
= specialized region on the presynaptic plasma membrane, where synaptic vesicles are docked and
primed for release; aligned with postsynaptic density (PSD)
- Active zone (AZ): specialized region on the presynaptic plasma membrane, where synaptic
vesicles are docked and primed for release; the AZ is aligned with the postsynaptic density
(PSD)
, - Molecular composition of PSD (more in chapter 8) (both present in pre and postsynaptic
neuron): neurotransmitter receptors, transsynaptic adhesion molecules, scaffolding
molecules (protein involved in keeping other proteins together e.g. receptors together etc.
for proper functioning), signal transduction molecules
3D reconstruction of axons and dendritic spines
Figure B on the right:
Reconstructing of axons (white) and part of the axons,
red, is where the synapse is located. In green
reconstruction of dendrite and this dendrite has
enlargements, boutons, and there the axons synapse.
What you can distinguish are boutons that have more
than one synaptic input (more than one axon) =
multisynaptic boutons. Boutons with one synapse is
single synaptic bouton. Non synaptic boutons used to be
boutons, but no synapse here. Synapses are
continuously modified in strength and structurally
modulated and new synapses can be formed and
existing synapses be lost → for example basis of learning.
mito: mitochondrion; mvb: multivesicular body; SSB: single synaptic bouton (single postsynaptic
partner); NSB: nonsynaptic bouton (no postsynaptic partner); MSB: multisynaptic bouton (more than
one postsynaptic partner); dcv: dense-core vesicle. vesicles are yellow
Figure A: Dendrite with dendritic spines. Spines are uniform in shape compared to figure B that show
more variation. A are more mushroom spines, more homogenous and bigger. Cerebellar Purkinje cell
dendrite; dendritic spines have relatively uniform shape; sp: spine. Figure B: Hippocampal CA1
dendrite; f: filopodia-like protrusion; st: stubby protrusion; m: mushroom-shaped spine; t: thin-
shaped spine. Red surfaces are PSD areas.
Which of the following are neuromodulatory neurotransmitters?
A GABA (→ inhibitory)
B glutamate (→ excitatory)
C dopamine (→ neuromodulatory)
D glycine (→ inhibitory)
,Lecture 2
Chapter 2: Synaptic cell adhesion molecules (SCAMs)
Learning outcomes:
➢ Deduce different functions of SCAMs
➢ Describe different stages of synapse formation and function + role of SCAMs in each stage
➢ Give examples of SCAMS and their functional domains
➢ Schematically draw SCAMs to illustrate function (currently not possible → images are shown)
Synaptic cell adhesion
Synaptic cell adhesion molecules (SCAMs) could be defined as transmembrane proteins (or at least
proteins that are linked to the membrane) which are localized at the synapse (either presynaptic or
postsynaptic, or both) and interact with each other across the synapse (transsynaptic).
➢ Synaptic junctions are organized by trans-synaptic cell adhesion molecules bridging the
synaptic cleft.
In the figure (representation of synapse. Synaptic cleft is between the presynaptic and postsynaptic
membrane) a schematic representation of transsynaptic adhesion molecules. The red molecules are
linked to the upper membrane. These are two cell adhesion molecules that are linked to the
presynaptic membrane and in green are SCAMs that are connected with the postsynaptic membrane.
Why do you know the upper one is presynaptic? → above you see synaptic vesicles, one docked and
primed and one not. So if an action potential arrives here, neurotransmitter from his vesicle will
release. What you further see are voltage gated calcium channels, which means that when an action
potential arrives, there is depolarization of the membrane and this will change the conformation of
the channels, which makes them open. Calcium can now flow in the presynaptic nerve terminal,
which triggers the exocytosis of the synaptic vesicle. The neurotransmitter that would then be
released will bind to the receptors on postsynaptic receptors. These receptors are ligand gated,
meaning that when the transmitter binds the receptor will change conformation and act as an ion
channel to conduct ions that now will flow into postsynaptic cell.
, The function of the cell adhesion molecules is on the one hand to link the post and
presynaptic membranes together so that the synaptic cleft has a given width. Because of the
interaction between the molecules is a standardized distance, which is the proper distance
for neurotransmission to occur. The neurotransmitter that is released needs to travel a given
distance to the postsynaptic receptors by diffusion ,which will result in a certain rate of
activation of the postsynaptic receptors. This is one function of the SCAMs → physically link
pre and postsynaptic site together and bridge the cleft:
➢ Synaptic junctions are organized by trans-synaptic cell adhesion molecules bridging the
synaptic cleft.
SCAMs also have other functions and important is to know that they are transmembrane
proteins, meaning they also have a short part of the protein at the intracellular site, where
they can interact with other intracellular proteins. This way they are involved in organizing
synapses. For example they can make sure that the calcium channels are clustered at
presynaptic membrane or synaptic vesicles are docked. In short they would organize the
presynaptic active zone. Similarly the postsynaptic cell adhesion molecules could interact
with neurotransmitter receptors in order to cluster them at postsynaptic site or interact with
signal transducing or scaffolding proteins. Together this would anchor the postsynaptic
density, which is a collection/dense concentration of proteins. :
➢ Synaptic cell adhesion molecules (SCAMs) not only connect pre- and postsynaptic
compartments, but also mediate trans-synaptic recognition and signaling processes that are
essential for the establishment, specification and plasticity of synapses.
➢ Examples of SCAMs:
o Neurexin
o neuroligins
o Ig domain proteins (e.g. synCAMs)
o Receptor phosphotyrosine kinases and phosphatases (first phosphorylates proteins
and second one dephosphorylates proteins)
o Leucine rich repeat proteins
First two implicated in schizophrenia and autism
Synaptic plasticity: the capacity of synapses to adjust their properties in response to previous activity
Four stages of synapse formation and function
1) First stage is establishment of synapse. When synapse is established there will be a
presynaptic axon nerve ending and a postsynaptic partner (e.g. dendritic spine). The pre- and
postsynaptic partners will approximate each other and they see if they fit together. This is
done on molecular level by synaptic cell adhesion molecules. These interact with each other.
This is also a distinction that is made between SCAMs that interact with each other but are
different (e.g. neuroligins and neurexins) → heterophilic interactions. When same molecule
is expressed pre and post → homophylic interaction. So contact establishment involves
recognition of pre- and postsynaptic neurons; this process may require heterophilic and
homophylic (between identical molecules) interactions between SCAMs to recognize
appropriate synaptic partners
2) Second phase is the assembly stage. During this stage the pre and post synaptic part
components are being assembled, such as the synaptic vesicles that are collected and
recruited to the presynaptic site. And in general the presynaptic active zone is recruited,
including the calcium channels. Also postsynaptic, the PSD is recruited including
neurotransmitter receptors and signaling and scaffolding molecules. The intracellular part of
the SCAMs will interact with other proteins that influence for instance the vesicles or calcium
channels or neurotransmitter receptors.
, So recruitment of synaptic
vesicles, AZs and PSDs; during this
stage, SCAMs regulate physical
cell-cell adhesion and serve as
anchor proteins to cluster or
recruit receptors or components
of the pre- and postsynaptic
machinery
3) Third stage is called the
specification stage. Once all the
components of the pre and
postsynaptic synapse have been
recruited, the synapse can start
functioning. Meaning when an
action potential reaches the
presynaptic nerve terminal, there
will be secretion of
neurotransmitters by synaptic
vesicle exocytosis, resulting in
activation of postsynaptic
receptors, which will result in
activation of postsynaptic cells.
The cell adhesion molecules are
during this stage responsible for
organizing the molecular
components of the synapse
(similar to stage 2 for SCAMs).
With all this the synapse now
functioning.
So functional specification stage:
organization of the molecular
components of the synapse, resulting in functionality of the synapse.
4) The fourth stage is the plasticity stage. Synaptic plasticity is essentially the adaptation of the
strength of the synapse based on previous activity. This adaptation of the synaptic efficacy
can involve either structural or functional changes. The cell adhesion molecules are also
involved in this stage. For instance if synaptic strength wishes to be increased, more vesicles
are recruited at synaptic site so more neurotransmitter release. Or they recruit more
postsynaptic receptors, which results in a higher chance that postsynaptic cells gets activated
in response to the neurotransmitter.
So synaptic plasticity: SCAMs may contribute to structural and functional synaptic plasticity in
activity-dependent adaptive events
Note: Once synapses have been established, it are not static structures. Synapses continue to be
formed and eliminated during the lifetime of an organism.
It is thought that learning and memory is based on this plastic capacity of our brain.
Functional domains of SCAMs
How do SCAMs function? On the one hand they need to interact with other cell adhesion molecules
in order to form trans-synaptic connections. The SCAMs do this through well defined protein
functional domains. Functional domains of a protein are domains that mediate a certain function of
the protein. In this case there is a limited number of functional domains that is found in different
synaptic cell adhesion molecules that mediate the adhesive function of those proteins. These
functional domains are often assembled in repeat units:
, ➢ The adhesive function of SCAMs is based on a limited number of extracellular domains, often
assemble intro repeat units:
o Examples of such domains are:
▪ Immunoglobulin (Ig) domains: most frequently bind to other lg domains;
most SCAMs containing lg domains also contain fibronectin III (FNIII)
domains.
▪ Cadherin domains: always occur in multiple copies connected by a linker that
bind 2-3 calcium ions. Therefore, cadherin mediated interaction are calcium
dependent and often homophylic.
▪ Lamin A, neurexin, and sex hormone binding protein (LNS -> mediate
interaction between SCAMs)) domains (aka laminin G-Like (LG) domains)
▪ Leucine rich repeats (LRRs)
Example neurexin-neuroligin interactions
ɑ-Neurexin has an LNS domain that interacts with
the Neuroligin molecule. The fact that those two
molecules form a dimer is mediated by an
esterase-domain of neuroligin. So that’s how the
two partners in the dimer recognize and interact
with each other.
(last year) Synaptic vesicle docked to presynaptic
membrane. There are cell adhesion molecules and
usually have a intracellular portion (short!) and an
extracellular portion. There are pre and post
synaptic adhesion molecules. They are to keep the
pre and post synaptic membrane together. This
for faster transmission. The molecules also are anchors to which other molecules can attach. Even
though both are different molecules they both have a protein domain at which they recognize each
other. The intracellular portions also connect to other proteins to activate them and can cause a
cascade
➢ Neurexins:
o Presynaptic single-pass membrane proteins with a large extracellular sequence and a
short cytoplasmic tail.
o Vertebrates have 3 neurexin genes (Nrx1-3); each gene produces from independent
promoters a larger ɑ- and a shorter β-neurexin isoform. (also seen in figure above: ɑ
larger than β)
o Extracellularly, ɑ-neurexins contain 6 LNS domain with 3 interspersed EGF-like
domains; β-neurexins only contain a single LNS domain.
o Neurexins are extensively alternatively spliced at 5 conserved sites in their
extracellular region, creating potentially thousands of isoforms (lots of forms of the
protein). This may constitute a recognition code (allows the cell to recognize the
appropriate post synaptic partner).
o Intracellularly, the short cytoplasmic tails of neurexins contain PDZ-domain (protein
interaction domain, which interacts with other proteins) binding sequences
(sequence in protein that binds other proteins that contain a PDZ domain) that bind
to intracellular proteins, e.g. CASK and protein 4.1
➢ Neuroligins
o Postsynaptic single-pass membrane proteins
