Rotarod Apparatus, Standard Operating Procedure

Rotarod Apparatus, Standard Operating Procedure

1. Rota-Rod Apparatus ( Standard Operating Procedure) Dr. Snehashis Singha Department of Pharmacology and Therapeutics King George’s Medical University, Lucknow

2. Learning Objectives: Introduction Aim & Principle Parts of instrument Functioning of instrument Precautions Materials & Method Observation Merits & Demerits Discussion Summary References

3. Introduction The Rotarod Apparatus is used in pharmacological studies to assess motor coordination, balance, and endurance in rodents It features a rotating rod that gradually increases in speed, testing the animal's ability to maintain balance on it The Rotarod test was invented by Dunham and Miya, in 1957

4. Aim To evaluate the effect of certain drugs on the motor coordination and balance of rodents by measuring their ability to stay on a rotating rod under controlled  conditions

5. Principle The Rotarod Apparatus tests motor coordination by measuring how long an animal can balance on a rotating rod. It typically shows shorter latencies to fall, indicating reduced performance

6. Apparatus Horizontal metal rod coated with rubber: 3 cm in diameter for mice & 6 cm in diameter for rat Speed of rotation of rod: 2-6 rotations/minute Length of rod: 75 cm Rod divided into 4 or 6 sections by plastic disc Rod is at height of 50 cm from table top Cut-off time: 2 minutes

7. Digital Display Screen Sensory P lates Metal Rod coated with rubber material Plastic plates to separate the sections Reset Buttons Speed Regulator Knob Pause button All Start Button All Reset Button Start Buttons Parts of Instrument : Front View

8. Reset Buttons Start Buttons Sensory Plates

9. Display Screen and Control Keys

10. ON/OFF Power Button and Main Power Supply Female Connector to connect the Rotarod to a Computer port Parts of Instrument : Back View

11. Placing of Animals on Rod

12. Functioning of Instrument - Preparation Connect the power cord to the main socket and switch on the device using the ON/OFF button at the back Set up the display screen Adjust the initial speed of the rotating rod

13. Precautions Laboratory should be dim lighted and noise free Animal should be marked properly, to avoid mixing in two groups Handle animals gently to minimize the stress and pain Precondition the animals with the procedure Inspect the apparatus regularly for issues or dirt buildup Avoid excessive speed or test durations that may injure animals

14. Materials Animals: Rats Species: Wistar Weight Range/Sex : 200-250 grams / Either sex, But same sex Syringe/needle : 1ml/preferably 23G onwards Drugs (if applicable): Relevant pharmacological agents (e.g., sedatives, stimulants) at species-appropriate doses . e.g.- Diazepam(Mice: 1-2 mg/kg ,i.p injection )

15. Methods Step 1 : Acclimatize the animals to the test environment for at least 15-30 minutes before testing to minimize stress Step 2 : Weigh the animals and mark them properly for identification and divide the animals into groups (Test and Control, n=6 in each group) Group 1 Control Group (n=6), mice are given saline at the equivalent drug dose Group 2 Test Group (n=6), mice are given diazepam at the dose of 3 mg/kg Step 3 : Place the animal gently onto the rotating rod and start the timer

16. Step 4 : Observe the animal for 2 minutes and note the time until it falls off the rod Step 5 : For drug trials, administer the drug according to the study protocol Step 6 : Repeat the procedure for each animal in the experimental group, ensuring consistent conditions across  tests

17. Observations Data Recording : Record the time each animal remains on the rotating rod at different speed of rotation Frequency of Testing : Each animal may undergo multiple trials to obtain average values Data Analysis : Compare the latency to fall times across control and test groups

18. Retention time on Rota Rod (Cut off 120 sec.) Sl . No. Pre. Drug Post Drug 1 2 3 4 5 6

19. Merits Highly effective for screening drugs affecting motor coordination, such as sedatives or stimulants . Allows objective and quantifiable measurement of motor skills . Simple to operate and provides rapid data for motor function assessments.

20. Demerits Habituation effects can occur, where animals improve performance through repeated exposure The test is best suited for drugs affecting motor function and may not reflect effects on other neurological processes Care must be taken not to overuse animals in trials, as repeated exposure may lead to stress or  injury

21. Calculations : (Rats) Dose of Diazepam to give = 2 mg/kg , 2mg in 1000gm F or 200 gm rat = 0.4 mg diazepam Vial = 5mg/ml of Diazepam , 1 ml=5 mg Diazepam We add 9ml (N.S.) with 1 ml of drug 1 ml + 9 ml = 10 ml   Solution 5mg Diazepam in 10 ml solution We need 0.4 mg diazepam in = 0.8 ml Given  injection Intraperitoneal route

22. Discussion Sl.No Body weight (grams) Fall time in seconds Control group A Fall time in seconds Treatment group B 1 200 220 46 2 250 250 5 2 3 300 200 65 4 350 150 6 4 5 400 149 60 6 450 186 73

23. Average fall time for control group A: 192.5 seconds Average fall time for treatment group B: 60 seconds Decrease in fall time: [(A-B)/A]X100 [(192.5-60 )/]X100 = 68.83% Thus, Diazepam decreases the time spent on rotarod by an average of 68.83%

24. Summary Rotarod Apparatus is used to test motor coordination It measures how long an animal can stay on a rotating rod Drugs that affect coordination reduce the time on the rod The test helps assess balance, coordination and muscle strength Animals falling off the rod indicate impaired motor function

25. References Medhi B, Prakash A. Practical Manual of Experimental and Clinical Pharmacology. 2nd ed. New Delhi: Jaypee Brothers Medical Publishers;2017.p 212-214.

26. THANK YOU

27. Questions: Name some drugs which can be screened through these tests Examples of Centrally and Peripherally acting muscle relaxants What is cut off period? What is the usual rotational speed of metallic rod in this apparatus? What are the other tests to study the motor co-ordination in the rodents ?