Christine Yen [鄧思婷] Taiwan Tech Trek Academic Conference August 14, 2015

Modeling Neurodegenerative

Diseases in Mice

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Parkinson’s Disease (PD)

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• Characteristics

- Loss dopaminergic neurons in substantia nigra pars compacta (SNpc)

• Causes: unknown, sporadic

- Genetic, environmental factors

• Prevalence

- Primarily among elderly

- 1-3% of population over 55 yrs

http://what-when-how.com/wp-content/uploads/2012/04/tmp3675_thumb_thumb.jpg

Parkinson’s Disease

• Pathophysiology

-SNpc releases dopamine-> receptors in striatum

- Dopamine deficiency

- Striatum cannot signal

motor thalamus, basal ganglia

• Symptoms - Tremors, rigidity, bradykinesia

difficulty walking

- Dementia

http://file.scirp.org/Html/2-2620020/24f49151-d68f-4540-bd71-c1b45e35562d.jpg http://discovermagazine.com/~/media/Images/Issues/2014/March/dopamine-chart.jpg

Striatum

SNpc

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6-OHDA Model

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• What is 6-OHDA?

‒ Neurotoxin induces SNpc neurodegeneration

• Dopaminergic selectivity - 6-OHDA structural analog:

dopamine -Taken into SNpc via dopamine transporters (DAT)

http://openi.nlm.nih.gov/imgs/512/250/2957224/2957224_PD2010-427810.002.png

Before

After

• Mechanism of neurotoxicity

- Degradation by MAO

- Generates H2O2, free radicals

- Inhibits mitochondrial complex I

• Why model PD?

- No cure, limited treatments

- Better understand pathology

- Develop drugs: pre-clinical trials

6-OHDA Model of PD

http://www.sciencedirect.com/science/article/pii/S030100820100003

Method of implementation

• Stereotaxic surgery

– Direct injections striatum

– Unilateral vs bilateral injections

• Retrograde degeneration of SNpc

– Mimics PD: humans

• Subjects

– 9 mice injected w/ 3 mg/kg 6-OHDA

– 3 controls injected with saline

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Motor Outputs

Left Brain Right Brain

Controls left side of body

Controls right side of body

6-OHDA lesion

Motor impairments develop

Motor function unaffected

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Onset: PD Phenotypes

• Successful model: replicates phenotypes

- Symptoms typically emerge within weeks

• Conduct behavioral assays

- Assess degree motor deficits develop

• Establish SOP’s: behavioral tests

-Ensure protocols run smoothly

http://www.nature.com/nrn/journal/v10/n7/fig_tab/nrn2652_F1.html 8

Behavioral Assays: Overview

Rotarod Test

Adhesive Removal Test

Forelimb Rearing Test

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Rotarod Test

• Assesses coordination, balancing abilities

• Rod acceleration: 4-40 rpm

• Walked 300s, rested: 15 min intervals

• Record time spent continuously walking

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(3 weeks post injection)

Rotarod in Action

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Data Collection/Results

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Rotarod Performance: 6-OHDA groups vs control (training period)

180

190

200

210

220

230

240

250

260

270

280

7/29 7/30 7/31

6-OHDAgroups

Control group

Ave

rage

Tim

e w

alke

d (

s)

Dates

14

215

220

225

230

235

240

245

250

255

260

265

7/29 8/5

6-OHDA groups

Control group

Rotarod Performance: 6-OHDA groups vs control A

vera

ge T

ime

wal

ked

(s)

Dates

Adhesive Removal Test

• Measure sensorimotor deficits

• Adhesives: alternately applied

• Left, right forepaw

• Measure time: sense, remove adhesives

• Mice affected by 6-OHDA

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Sticker

Forelimb Rearing Test

• Assess forelimb motor impairment

• Placed glass cylinder

• Note forelimb favored: when rearing

• Record frequency use:

• Left, right or both forelimbs

• Mice affected by 6-OHDA:

• Ipsilateral forelimb dependence

Forelimb Rearing Test

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Modeling Alzheimer’s Disease (AD)

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Morris Water Maze

• Alzheimer’s disease behavioral assay

• Tests spatial learning/memory

• Developed SOP’s

• Protocol runs smoothly

• Establish appropriate environment

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MWM Procedure

▪ Mice swim in tub: opaque water

▪ Find submerged platform

▪ Visual cues posted: walls

▪ Released: four different directions

▪ Factors of importance

NW

W SE

SW

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Conclusions

▪ Early stages: developing PD phenotypes

▪ SOP’s: identify, correct interfering factors

▪ Lack training

▪ Timing: trials

▪ Mouse behaviors

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Acknowledgements

▪ My appreciation to:

Dr. Huang for her guidance

National Animal Laboratory Center, NARLabs

The mouse subjects

▪ Thank you all!

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