How are neurotransmitters indentified?

The first criterion by which neurotransmitters are recognized as such is that they must be stored and synthesized in the PRE-SYNAPTIC NEURON. With this in mind we turn to two methods of identifying molecules that are stored and synthesized in pre-synaptic neurons, making them worthy neurotransmitter candinates.

Immunocytochemistry is an interesting method with a number of steps. The molecule of interest (the molecule we want to determine whether or not it is neurotransmitter), is injected into the blood of an animal, causing anti-bodies to be released which bind to the 'foreign' substance. In immunology, anti-bodies are specific to chemicals they are deployed against so we can expect anti-bodies that are released to deal with (bind to) an injected substance, let's say, dopamine, to also bind to dopamine if extracted from blood and applied to brain tissue. And this is what actually occurs in the immunocytochemistry technique: the anti-bodies are extracted via withdrawing blood from the animal, and are then tracer-marked (so their activity can be located), and are then applied to brain tissue. The anti-bodies will bind to concentrations of neurotransmitter, giving us localized visual descpritions of the concentration of neurotransmitter. If a neurotransmitter and its synthesizing enzyme is found in the same axon, this helps satisfy the first criterion for idenfication as neurotransmitter.


A second method is In Situ Hybridization, another very interesting method. Here, artifical mRNA is constructed in a labatory with a specific code for binding to mRNA that synthesizes neurotransmitter synthesizing enzymes. The artifical mRNA is labelled with a radioatctive 'tag' and applied to brain tissue. The point in this method is to review locations where neurotransmitter synthesis takes place. Radioactivity of 'probe' mRNA is viewed in autoradiography where locations of synthesis will be revealed. To identify the neurotransmitter, the specific neurotransmitter that the probe mRNA sequence ultimately synthesizes will be reviewed. We have thus identified the neurotransmitter associated with the located synthesizing enzymes.



These methods of identifying neurotransmitter are currently being replaced with more up to date methods of searching for neurotransmitter via receptors.