Question

You are a graduate student testing a new drug (drug G) that reduces cocaine addiction in mice. You want to find out what the pharmacological action of drug G is, and you know that it involves monoaminergic signaling, but you don’t know more about it than that. You have found a monoaminergic synapse whose signaling is affected by drug G. You can stimulate the presynaptic neuron at this synapse and record the resulting EPSPs in the postsynaptic neuron. This synapse only releases one neurotransmitter (it is a neurotransmitter we have discussed in class), and at this synapse there is only one type of receptor for this particular neurotransmitter.

You decide to first identify the neurotransmitter released at the synapse you are studying, so you apply an antibody to detect aromatic amino acid decarboxylase (AAAD). You find that the presynaptic neuron at this synapse does express AAAD.

Which of the following types of neurotransmitter COULD be released by this neuron? MORE THAN ONE ANSWER MAY BE CORRECT. CHOOSE ALL THAT APPLY.

	a.	Dopamine
	b.	Epinephrine
	c.	Norepinephrine
	d.	5-HT

Based on the results of the experiment you did in the previous question, you realize that you still haven’t definitively narrowed it down to a single type of neurotransmitter. So, you do another experiment.

Which of the following experiments would help you be more specific? Note that the correct answer wouldn’t necessarily give you the final answer (i.e. narrow it down to a single neurotransmitter), but it would help narrow it down somewhat.

	a.	Reduce the intracellular concentration of BH4
	b.	Stain for VMAT2
	c.	Block the function of monoamine oxidase
	d.	Label the tissue for PNMT

Finally, an undergrad in the lab suggests you stain for DAT, and low and behold, you determine that DAT is expressed in the presynaptic neuron. Now have your answer about which type of neurotransmitter is actually released at this synapse.

Now, you start trying to figure out how drug G affects signaling at this synapse. The next two questions indicate ways drug G might affect signaling. Choose the answer for each question that accurately describes the effect drug G would have at the synapse if the mechanism indicated in each question was correct . For each question, assume drug G has a single action at this synapse (i.e. only directly affects a single type of receptor, enzyme, etc., as relevant).

What would occur if drug G blocked DAT?

	a.	EPSP magnitude would increase
	b.	EPSP magnitude would decrease
	c.	EPSP magnitude did not change

What would occur if drug G increased the efficiency of COMT?

	a.	EPSP magnitude would increase
	b.	EPSP magnitude would decrease
	c.	EPSP magnitude did not change

Answer 1

1. As per the given data, the most possible neurotransmitters which could be released are dopamine, epinephrine and nor-epinephrine. As the antibody is tested for AAAD (aromatic amino acid decarboxylase) which is the common enzyme for synthesis of all the three neurotransmitters.

2. To narrow down the possibility of neurotransmitter secreted, option b and d could be performed. Dopamine released is sequestered in synaptic vesicles or particles following uptake by the relatively nonspecific ATP-dependent vesicular membrane transporter 2 (VMAT-2), so staining for VMAT-2 could be done. But in complementary labeling the tissue for PNMT is essential experiment to check further conversion of DOPA. Absence of PNMT in tissue would confirm the neurotransmitter to be DOPA and narrow down the correct neurotransmitter released.

3. Its well known that cocaine addiction is reduced by lowering dopamine level. As the selected drug G is known to reduce cocaine addiction its mechanism would be to block synthesis of dopamine. Hence EPSP magnitude would decrease.

4.A catechol-O-methyltransferase (COMT) inhibitor is a drug that inhibits the enzyme catechol-O-methyl transferase. This enzyme methylates catecholamines such as dopamine, norepinephrine and epinephrine. Use of drug to increase efficiency of COMT inhibitors would increase magnitude of EPSP.