Biology 463 - Neurobiology
Topic 6
An Introduction to
Neurotransmitter Systems
Lange
Introduction
There are three classes of neurotransmitters (as discussed in Chapter 5)
– Amino acids, amines, and peptides
There are a variety of different ways of defining particular transmitter
systems:
–
–
–
–
–
By the molecule
synthetic machinery
packaging
reuptake and degradation
as well as others
It is important to note that all neurotransmitter systems can be described by
the above 5 ways.
Neurotransmitter
systems have five
different ways they
can be described.
Henry Dale – received the Nobel Prize for Medicine in 1926
for his work on synaptic transmission.
He is also well known for his coining of the term “cholinergic.”
System #1: Cholinergic System
• acetylcholine (Ach)
System #2 : Noadrenergic System (adrenergic derived from “adrenalin” the
origin of NE
• norepinephrine (NE)
System #3: Glutamatergic System
• glutamate
System #4: GABAergic System
• gamma-aminobutryic acid (GABA)
System #5: Peptidergic System
• various peptide neurotransmitters
With the myriad of different chemicals produced in neurons, how do we define a
“neurotransmitter”?
• the chemical must elicit change in another neuron or cell
Experimental Design Associated With
Identifying Neurotransmitters
However, because the chemicals are in minutely small quantities and because it is challenging to discern
which chemical is actually affecting a receptor, the following criteria are used to assess experimentally if
something is a neurotransmitter:
•
•
•
the chemical must be synthesized and stored in a presynaptic neuron (Criteria A)
the chemical must be released by the presynaptic neuron upon stimulation (Criteria B)
exogenous application of the chemical will elicit a response that is similar to endogenous release
(Criteria C)
In the next several slides, we will look at a variety of ways to establish the above
criteria.
Studying Transmitter Localization
I. Immunocytochemistry (ICC)– a technique used to localize
production of chemicals (molecules) to particular cells
Steps in this technique:
1.
2.
3.
4.
Purified molecule of interest is injected into an animal to provoke an immune response (keep in
mind these are protein-based molecules, or if smaller, they are coupled to larger proteins).
The antibodies that the host animal produces can be collected, purified and tagged with a marker
(radioactive, fluorescent. or another type).
These tagged antibodies (designed by the host to attack the “invader” injection, will now bind to
any sample of nervous system tissue with the same molecule when they are applied.
This will then allow for identification of brain, spinal cord, and other nervous system tissues that
have the same suspected neurotransmitter compounds.
This technique helps to show localization of production and storage (Criteria A)
Please note that in this slide of the rat brain, ICC
has been used to identify vasopressin containing
neurons and therefore identify vasopressin
producing regions of the brain.
II. In situ hybridization – a technique where the identification and
localization of a synthesized protein or peptide occurs (at the cell level).
This technique is essentially detecting mRNA.
1.
2.
3.
4.
5.
A complementary strand to a known mRNA is made in the lab (this is called a probe).
When the probe binds to mRNA, this is the process of hybridization.
The probe is labeled in some fashion (like with ICC, this can be radioactivity, fluorescent dye,
or something else)
The probe will “stick” to the mRNA of particular neurons containing that mRNA sequence and
therefore will identify it.
Radioactive isotopes are still most commonly used in this process and are detected via
autoradiography (basically with X-ray film).
I. and II. in the preceding detect potential neurotransmitters via TWO different routes.
Can you identify these routes?
A variant of traditional In-Situ Hybridization that uses fluorescent labeled probes
and allows the use of a light microscope to view.
Studying Transmitter Release
Studying transmitter release is fairly effective in the PNS, but it is much more difficult in the CNS
because the range of different chemicals in the numerous different regions of the brain and spinal cord
make isolation very difficult.
One technique that can be used to approximate the study of CNS transmitter release to a REGION (not
an individual neuron) of the brain or spinal cord involves in vitro techniques where slices of brain or
spinal cord are kept alive in physiological saline (specifically neurophysiological saline).
With this technique, and then by bathing the tissue with a different solution very high in potassium,
membrane depolarization can be stimulated.
A challenge remains however… just because a neurotransmitter candidate is RELEASED,
does not mean it is a neurotransmitter… it could be simply a secondary chemical that is
released or some other form of artifact. This ambiguity makes it difficult to establish
Criteria B with certainty in the CNS.
Studying Synaptic Mimicry
Microionophoresis – a technique that helps to establish
Criteria C, that responses are able to be ellicited by
exogenous application of the neurotransmitter candidate.
1.
2.
3.
The neurotransmitter candidate is dissolved in solution
and acquires an electrical charge.
A pulled micropipette is used to insert into a neuron the
solution of interest.
If the response of the exogenous injection is the same
as the endogenous effect, it has been shown that the
molecule of interest is a neurotransmitter according to
Criteria C.
A typical
pipette
puller like
this can
make a
glass tube
with
openings
only a few
micons in
diameter.
Three Crucial Terms Used for Studying
Neurotransmitters:
Agonist - a chemical that binds to a receptor of a cell and
triggers a response by that cell. Agonists often mimic the
action of a naturally occurring substance.
Antagonist - whereas an agonist causes an action, an
antagonist blocks the action of the agonist.
Inverse Agonist – compound that causes an action
opposite to that of the agonist.
Studying Receptors
Neuropharmacological Analaysis – the discovery of TWO different
cholinergic receptor types for acetylcholine (Ach) that act differently was
made through this technique.
The first type has been called the MUSCARINIC receptor.
Muscarine is an agonist and its effect is to stop the
heart. It works in a much similar way to Ach on
cardiac muscle.
Ach Receptor Subtype 2 is the receptor type found
on cardiac cells.
Muscarine – a type of
poison found in some
mushrooms.
Moto-Taxi Driver (1999) at a coffee shop in Bangalore, India
He is stimulating his nicotinic receptors.
The second type has been called the NICOTINIC
receptor.
Nicotine is an agonist and its effect is to
affect the Ach receptors on skelletal
muscle.
Ach Receptor Subtype 1 is the receptor
type found on skelletal muscle cells.
END.