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Zebrafish: An emerging model in drug abuse research
Kevin S. Jones, Ph.D.Assistant. Professor PharmacologyThe College of Osteopathic Medicine of the PacificWestern University of Health Sciences
Charles Drew University 4th Annual Drug Abuse Research Symposium September 26,2008
What is a “zebrafish”?
The zebrafish, Danio rerio, is:
• Tropical, fresh water fish native of India.
•Member of the teleost class (bony fish).
•Well established model for vertebrate development.
•Readily accessible to transgenesis.
Zebrafish have 3 developmental stages
~30 hpf (hours post fertilization)
1 mm
Embryonic
> 3 months
Adult
5 dpf
1 mm
Larval
Zebrafish as a model organism
Advantages Disadvantages1. Small size
2. High fecundity
3. Sequenced genome
4. Optical transparency
5. External Development
6. Genetic manipulation
7. Automated analysis
Evolutionarily distant from humans
Limited behavioral repertoire
Larval zebrafish (10dpf)
96 well plateU.S. Penny
How can zebrafish be used to study drug abuse?
pcp
PCP (phencyclidine) blocks NMDA receptors
PCP
PCP
kd < 5 nM
MK-801
MK-801 is a potent analogue of PCP
Psychostimulants cause movement disorders
Actions of psychostimulantsDrug-treated
HUMANDrug-treated
RodentsHallucinations YES ???
Delusions YES ???
Thought Disorder YES YES
Movement Disorders YES YES
Zebrafish have NMDA receptors
• zNMDAR is ~90 % IDENTICAL to human NMDAR
•Expressed as early as 24 hpf
Developmental Dynamics 2005 234: 756-766
Objective:
Characterize the behavioral effects of NMDA receptor blockade in zebrafish
• Individual zebrafish larvae are placed into single wells of a multi well plate
• Multi-plate illuminated from below with a 15w lightbox
• Larvae given 5 minutes to acclimate
• Subsequent activity recorded for 5 minutes on digital camera
Data Acquisition
Methods
MethodsData Analysis
• Video recordings analyzed off-line with particle tracking software
Vehicle-treated MK-801-treated
Prototypical Data
Dose Response of MK-801-induced locomotion
EC50 ~ 10.3 uM
Locomotor kinetics
t (h)
Spe
ed
(mm
/sec
)
Developmental Sensitivity
The antipsychotic, ziprasidone,attenuate the locomotor effects of MK-801
Ziprasidone targets several receptors
Target Receptor D2 D3 5HT2A 5HT2C 5HT1A 5HT1D α1
Ki 4.8 7.2 0.4 1.3 3.4 2 10
Dopamine antagonists do not disrupt MK-801-indcued locomotion
MPTP pre-treatment diminishes locomotor effects of MK-801
Δ S
PE
ED
(mm
/sec
)
VEH 500 uM MPTP 1 mM MPTP
Summary• Zebrafish retain the most reliable symptom of
NMDA blockade… increased locomotion.
• Antipsychotic meds block behavioral effects in zebrafish.
• Zebrafish may prove useful for elucidating importantmolecular pathways of drugs of abuse
Future Directions
• Elucidate molecular mechanism(s).• Examine developmental sensitivity.• Probe for behavioral sensitization.• Molecular neurotoxicology.
Acknowledgments
Mr. John Chen (Pomona College)
Ms. Roshni Patel (Pomona College)
Biology Dept Pomona College
Pomona College—Dean’s office