1. DESCRIBING NEUROPIL REGIONS IN THE ZEBRAFISH BRAIN
COLLETTE GOODE
MENTOR: ALEX TALLAFUSS
WASHBOURNE LAB, INSTITUTE OF NEUROSCIENCE
UNIVERSITY OF OREGON
OURS 2016

2. axon
cell body
dendrite

3. NEUROPIL REGIONS ARE DENSE WITH DENDRITES, AXONS, AND SYNAPSES.
Whole Brain Catalog, UCSD , 2010
axon
cell body
dendrite

4. A SYNAPSE IS THE JUNCTION BETWEEN TWO NEURONS.
PRE-SYNAPTIC TERMINAL
POST-SYNAPTIC TERMINAL
THERE ARE EXCITATORY AND INHIBITORY SYNAPSES.
THOUSANDS OF PROTEINS FORM A SYNAPSE.

5. ZEBRAFISH ARE EXCELLENT MODELS FOR STUDYING SYNAPSE FORMATION

6. PURPOSE
TO DESCRIBE NEUROPIL REGIONS IN THE DEVELOPING ZEBRAFISH BRAIN:
WHAT IS THE DISTRIBUTION OF SYNAPTOTAGMIN 2B?
SYNAPTOTAGMIN: PRE-SYNAPTIC CALCIUM SENSOR
WHAT OTHER PRE-SYNAPTIC PROTEINS DOES SYNAPTOTAGMIN ASSOCIATE WITH?
WHAT IS THE DISTRIBUTION OF EXCITATORY AND INHIBITORY SYNAPSES?

7. 2. IMMUNOHISTOCHEMISTRY
1. HISTOLOGY
2. IMMUNOHISTOCHEMISTRY
FLUORESCENT TAG
SECONDARY ANTIBODY
PRIMARY ANTIBODY
SYNAPTIC PROTEIN
3. CONFOCAL MICROSCOPY

8. olfactory bulb
optic tectum
telencephalon
diancephalon
cerebellum
medulla

9. LABELED SYNAPTOTAGMIN 2B AND PRE-SYNAPTIC PROTEINS, SV2 AND SYNAPSIN.
SYNAPTOTAGMIN 2B WAS STRONGLY EXPRESSED IN MIDBRAIN AND HINDBRAIN.
LABELED SYNAPTOTAGMIN 2B AND PRE-SYNAPTIC PROTEINS, SV2 AND SYNAPSIN.
SYNAPTOTAGMIN 2b
SYNAPSIN
SV2

10. LABELED SYNAPTOTAGMIN 2B AND PRE-SYNAPTIC PROTEINS, SV2 AND SYNAPSIN.
SYNAPTOTAGMIN 2B WAS STRONGLY EXPRESSED IN MIDBRAIN AND HINDBRAIN.
LABELED SYNAPTOTAGMIN 2B AND PRE-SYNAPTIC PROTEINS, SV2 AND SYNAPSIN.
OPTIC TECTUM
CEREBELLUM
SYNAPTOTAGMIN 2b
SYNAPSIN
SV2
TELENCEPHALON
OPTIC TECTUM
UPPER CEREBELLUM
LOWER CEREBELLUM
MIDBRAIN

11. SYNAPTOTAGMIN 2B STRONGLY ASSOCIATED WITH EXCITATORY AND INHIBITORY SYNAPSES IN OPTIC TECTUM
GLUTAMATE RECEPTOR:
EXCITATORY POST-SYNAPTIC PROTEIN
GEPHYRIN:
INHIBITORY POST-SYNAPTIC PROTEIN
_ _ _ _ _ _ _

12. SYNAPTOTAGMIN 2B STRONGLY ASSOCIATED WITH INHIBITORY AND EXCITATORY SYNAPSES IN OPTIC TECTUM
GLUTAMATE RECEPTOR:
EXCITATORY POST-SYNAPTIC PROTEIN
GEPHYRIN:
INHIBITORY POST-SYNAPTIC PROTEIN
_ _ _ _ _ _ _
GEPHYRIN
SYNAPTOTAGMIN 2b
GLUTAMATE RECEPTOR

13. NEUROPIL REGION IN CEREBELLUM
GLUTAMATE RECEPTOR:
EXCITATORY POST-SYNAPTIC PROTEIN
GEPHYRIN:
INHIBITORY POST-SYNAPTIC PROTEIN
_ _ _ _ _ _ _
SYNAPTOTAGMIN 2b
GEPHYRIN
GLUTAMATE RECEPTOR
UPPER
CEREBELLUM
LOWER
CEREBELLUM

14. SYNAPTOTAGMIN 2B ASSOCIATES WITH BOTH GEPHYRIN AND GLUTAMATE RECEPTORS IN MIDBRAIN NEUROPIL REGION
_ _ _ _ _ _ _
SYNAPTOTAGMIN 2b
GEPHYRIN
GLUTAMATE RECEPTOR

15. SYNAPTOTAGMIN 2B STRONGLY ASSOCIATED WITH GEPHYRIN IN MEDIAL NEUROPIL REGION
_ _ _ _ _ _ _
SYNAPTOTAGMIN 2b
GEPHYRIN
GLUTAMATE REEPTOR

16. CONCLUSIONS SYNAPTOTAGMIN 2B WAS NOT EXPRESSED IN TELENCEPHALON.
SYNAPTOTAGMIN STRONGLY ASSOCIATED WITH SV2 ANTIBODY.
SYNAPTOTAGMIN STRONGLY ASSOCIATED WITH BOTH GEPHYRIN AND GLUTAMATE RECEPTOR IN OPTIC TECTUM, LOWER CEREBELLUM AND MIDBRAIN.
SYNAPTOTAGMIN ASSOCIATED WITH GEHPYRIN PROTEIN IN MIDBRAIN NEUROPIL REGION.

17. FUTURE DIRECTIONS
USE TRANSGENIC VGLUT:GFP LINE TO VERIFY ANTIBODY LABELING OF EXCITATORY SYNAPSES
IDENTIFY NEURON TYPES: USE TRANSGENIC LINE TO IMAGE INDIVIDUAL NEURONS
NEUROPIL DEVELOPMENT : APPLY IMAGING METHODS TO JUVENILE ZEBRAFISH

18. ACKNOWLEDGEMENTS
EISEN LAB UNIVERSITY OF OREGON FISH FACILITY HISTOLOGY LAB, POH KHENG LOI SPUR: PETER O’DAY AND MARILYN DRENNAN FUNDING: NIH UNDERGRADUATE RESEARCH PROGRAM AT THE UNIVERSITY OF OREGON NIH-R25HD07081
PI: PHILIP WASHBOURNE MENTOR: ALEX TALLAFUSS WASHBOURNE LAB: MELISSA POSTLER JAVIER FIERRO SARAH STEDNITZ AND THE REST!