EVALUATION OF THE EFFICACY OF TEST FORMULATIONS IN MOUSE MODEL OF MPTP-INDUCED PARKINSON’S DISEASE

EVALUATION OF THE EFFICACY OF TEST FORMULATIONS IN MOUSE MODEL OF MPTP-INDUCED PARKINSON’S DISEASE

1.0    TEST SYSTEM DETAILS:

Species                  :Mus musculus(Mouse)

Strain                    :C57BL/6

Age                       : 6-8 weeks

Body Weight         : 22- 25g

Sex                        : Male/ Female

No. of animals      : 10 animals /Group

Total animals        : 70

Study duration      :40 days

2.0   ALLOCATION OF GROUPS:

Groups	Treatment	Dose; ROA	No. of Animals
G1	Normal Control	0.5% MC, p.o.	10
G2	Disease Control	0.5% MC, p.o.	10
G3	Reference Control – Selegiline	3 mpk, p.o.	10
G4	Test Formulation 1	X1 mpk; p.o.	10
G5	Test Formulation 1	X2 mpk; p.o.	10
G6	Test Formulation 1	X3 mpk; p.o.	10
G7	Test Formulation 1	X4 mpk; p.o.	10

No extra animals will be taken; X1, X2, X3, and X4 are defined as the incremental doses of the Test formulation. MC-Methyl Cellulose; ROA-route of administration; mpk-mg/kg, p.o.-per os

3.0  METHOD:

·       Animals will be procured, randomized based on body weight and allocated into 7 different groups consisting of 10 animals each.

·       Animals of Group G1 will serve as Normal control and will receive 0.5% MC, p.o.

·       Animals of Group G2 will serve as disease control group will receive 0.5% MC, p.o.

·       Animals of Group G3 will serve as reference control and treated with 3 mg/kg selegiline p.o.

·       Animals of Group G4 to G7 will be treated with test compounds at respective dose levels.

·       Basal neurobehavioral readings will be assessed, prior to MPTP injection for all groups.

·       Three days before MPTP treatment, mice will be given the drug treatment orally. Drug treatment will be continued for a total of 18 days.

·       To construct the model, mice will be injected with MPTP (dissolved in 0.9% saline, 30mg/kg, intraperitoneally) for 5 consecutive days, while the normal control group will be given an equal volume of normal saline, intraperitoneally.

·       Two days after the last MPTP injection, behavioural assessments will be performed using the open field test, pole test and rotarod test.

Collection and Measurement of Specimens:

·       On Day 19, blood will be collected from the retro-orbital sinus and subsequently the animals will be euthanized with an overdose of thiopentone followed by collection of brain for histopathological, biochemical and molecular evaluation.

 

4.0 END POINT PARAMETER(S):

·       Clinical observation

·       Body weight

·       Open field test

·       Rotarod test

·       Pole test

·       Histopathology and immunohistochemistry of the brain

·       High-performance liquid chromatography (HPLC) for the estimation of neurotransmitters in the brain

·       Western blot analysis of the brain

·       Measurement of Cytokines in the brain

·       qPCR in the brain

5.0 REFERENCES:

1.    Zhang, Qiu-shuang, et al. "Reassessment of subacute MPTP-treated mice as animal model of Parkinson's disease." Acta Pharmacologica Sinica 38.10 (2017): 1317-1328.

2.    Lee, Eunju, et al. "MPTP-driven NLRP3 inflammasome activation in microglia plays a central role in dopaminergic neurodegeneration." Cell Death & Differentiation 26.2 (2019): 213-228.

                                                                          END THE DOCUMENT

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