1. Neurotransmitter Transporters and Flies: Tools to Study Behavior and Disease
David Krantz MD, PhD
Department of Psychiatry & Biobehavrioal Sciences
David Geffen School of Medicine at UCLA
Gonda (Goldschmied) Center for Neuroscience and
Genetics Research

2. Overview What are neurotransmitter transporters?
How do changes in their function affect the nervous system?
Why use flies to study them?

3. Neurotransmitters are chemicals that allow cells to communicate
S y n a p s e
Neurotransmitter
Cell 2
Receptors

4. Neurotransmitter types
Monoamines
Dopamine, Serotonin, Norepinephrine, Epineprine (Adrenalin)
Actetylcholine
GABA
Glutamate
(Peptides, Gases- not today)

5. Plasma membrane and vesicular transporters

6. Why do neurotransmitters need transporters?
Oil and water don’t mix
Transmitters like to be in water
Cell and organelle boundaries are oily
Transporters bridge the water pools

7. Plasma Membrane Neurotransmitter Transporters
-Monoamine transporters
are part of one gene family
-GABA and glutamate
transporters are in other
families
Dopamine
DAT
Noradrenaline
NET
Serotonin
SERT
Bröer S Br J Pharmacol , 256.

8. VMAT2 in all aminergic brain cells
Mammalian Vesicular Transporters
VMAT2 in all aminergic brain cells

9. Focus on Monoamine Neurotransmitters
Dopamine
Reward
Serotonin
Appetite,Mood
Gastrointestinal motility
Noradrenalin
Attention
Blood pressure
Histamine
Gastric acid release
Immune response

10. Monoamine Transporters the Major Drug Targets for Stimulants and Antidepressants
Vesicular monoamine
Transporter
Dopamine transporter
Serotonin “ “
Norepi “ “

11. Blockade of Plasma Membrane Amine Transporter Increases Extracellular Neurotransmitter
Cocaine
Ritalin
Prozac
Wellbutrin
Zoloft
Dopamine transporter
Serotonin “ “
Norepi “ “

12. Amphetamines Reverses the Flow (Mechanism is complex)
Dopamine transporter
Serotonin “ “
Norepi “ “
Amphetamines
Methamphetamines
Adderall- ADHD

13. Vesicular Transporters are Drug Targets
Vesicular monoamine
Transporter (VMAT2)

14. Block VMAT with reserpine: Decreases monoamine storage and thus release -Antihypertensive effects -Depression
Reserpine
Vesicular monoamine
Transporter (VMAT)

15. What would happen if VMAT worked better. Or there was more of it
What would happen if VMAT worked better? Or there was more of it? Could that change behavior? Act as a stimulant? Antidepressant?

16. Increases monoamine release
More VMAT in vitro:
Increases monoamine release
Force cells to make more VMAT
Record amine release
More amines comes out of each vesicle
(Vesicles get a little bigger)
“Normal cells”
Extra VMAT
Sulzer lab
Pothos et al, J. Neurosci. 2000

17. What about more VMAT in vivo?
-No in vivo mammalian models
-Make fly model
-Why use flies?

18. Why use flies? -Cheap! (good for teaching!) -”Conservation” of genes
e.g. VMAT, DAT
-Cool genetic tools
e.g. to make more VMAT
-Short life span
Genetic experiments in a month!

19. How can we tell if there are more extracellular monoamines in flies
How can we tell if there are more extracellular monoamines in flies? -Look at their known functions
Dopamine
Grooming
Locomotion
Octopamine
Egg laying
Locomotion
Serotonin
Aggression

20. How can we tell if there are more extracellular monoamines in flies
How can we tell if there are more extracellular monoamines in flies? -Look at their known functions
Dopamine
Grooming
Locomotion
Octopamine
Egg laying
Locomotion
Serotonin
Aggression
Stimulants as positive control

21. DVMAT overexpression in vivo mimics the effects of stimulants
Locomotion
Grooming
Chang et al 2006

22. A New Way to Increase Extracellular Amines
Wellbutrin
Prozac
Ritalin
Adderall

23. Could we find a drug the would make VMAT work better
Could we find a drug the would make VMAT work better? Or just increase exocytosis of monoamines? Could this be used to treat ADHD? depression? Parkinson’s disease?

24. Antidepressants and stimulants Some mechanisms not exploited
No Current
Drugs:
Activate
VMAT
Increase exocytotic release
Amine agonist
Receptor
VMAT
Aminergic
neuron
Current
Drugs:
Block reuptake.
degradation
Amphetamines cause efflux, not exocytotic release

25. To find drugs that might make VMAT work better… First make it work worse! Use dVMAT mutant
“Sensitized genetic background”
detects drug effects better than wild type

26. Primary Screen Test drugs in dVMAT mutant1 2 4 5 6
Drug: 3
..1042
Mix into food, Allow larvae to feed, record movement 1 2 3 4 5 6

27. Example of Primary Screen Data
-Select drugs that cause 3-4 SDs above the mean
-40 hits out of 1042 drugs
-3 time points, 2 concentrations, 7 undergrads, 5 months
Lawal et al, 2012

28. Dacarbazine was one of the “hits” in the screen
Chemotherapeutic agent
Alkylating agent
Induces emesis via serotonin release
Supports fly data
Toxicity could be a problem
Limits use

29. Parse toxic vs. active bits with derivatives
Dacarbazine
AICA
Methdiazonium
DNA alkylation
AICA: Amino-4-imidazole-carboxamide
Removes the toxic bit

30. AICA also stimulates larval locomotion
Dacarbazine
AICA

31. How does Dacarbazine/AICA Increase Locomotion?
Increase Storage?
Increase Release?
Other mechanisms?
Collaborate with Nigel Maidment’s lab

32. Potential Clinical Relevance
AICA derivatives for ADHD? Depression?
Parkinson’s disease?

33. What are neurotransmitter transporters?
Why use flies to study them?
How do changes in their function affect the nervous system?

34. Questions?