Parkinson Disease Model
Parkinson’s Disease (PD) is a neurodegenerative disorder characterized by the loss of dopaminergic neurons in the brain.
Exposure to the neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) induces selective loss of zebrafish dopaminergic neurons also, eliciting symptoms characteristic of PD. This model allows for identifying compounds that prevent the loss of dopaminergic neurons and the subsequent behavioral alterations.
Applications
- Study dopaminergic neuronal population.
- Study the molecular and physiological mechanisms of PD.
- High throughput screening of new therapeutic compounds for PD.
Advantages
High conservation of zebrafish dopaminergic system with vertebrates and humans.
Mammal MPTP-induced neurotoxicity mechanism is conserved in zebrafish.
Automated locomotion monitoring allows reproducibility testing and unbiased results.
Behavioral endpoints, which entails complex connections, in an in vivo model with high experimental throughput.
Transparent embryos permit in vivo characterization of specific neuronal populations.
Method description
The assay consists of co-incubating the candidate drug together with the neurotoxin MPTP and evaluating the locomotor behavior of the treated larvae. The experimental protocol for MPTP-induced Parkinson's phenotype consists of 25min dark/light alternating environments (5 min each). Optionally, the startling behavior in response to mechanical stimuli (tapping) and the diencephalic dopaminergic neuron population can be assessed in treated larvae. This evaluation is focused on the tuberculum area by the in situ hybridization technique enabling visualization of dopaminergic neurons.
Figure 1. Behavioral protocol scheme. Blue arrows depict dark phases, whereas white arrows depict light phases.
Readouts
- Locomotion activity: total distance moved under light/dark conditions and symptomatic movement patterns are evaluated.
- (Optional) Startle response: percentage of animals responding to physical stimulus (tapping).
- (Optional) Quantification of dopaminergic neurons in the diencephalic region by anti-tyrosine hydroxylase (anti-TH) immunolabeling.
Figure 2. Light/dark larvae locomotion per minute. MPTP-treated larvae show a strong decrease in locomotion during dark and light phases. In this example, the test compound induces a partial recovery of the control phenotype, increasing larval locomotion during the dark phase. Means and SD are plotted.
Figure 3. Startle behavior in response to mechanical stimuli (tapping). Dots depict the average larval response to each tap. All the groups show similar startle responses, which confirms that differences in locomotion between groups are not due to mortality or extreme toxicity but to MPTP-induced neurotoxicity and locomotor problems. Means and SD are plotted.
Figure 4. Tyrosine hydroxylase expression in the tuberculum area. Representative images of zebrafish larvae treated with MPTP (right) or not treated (left) after performing in situ protocol labeling the tyrosine hydroxylase (TH) expression. Arrows depict the loss of the TH expression in the tuberculum region after MPTP treatment.
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References
- McKinley ET, Baranowski TC, Blavo DO, Cato C, Doan TN, Rubinstein AL. Neuroprotection of MPTP-induced toxicity in zebrafish dopaminergic neurons. Brain Res Mol Brain Res. 2005 Nov 30;141(2):128-37.