1.0 PURPOSE
To design a Standard Operating
Procedure that describes the procedure for operation and maintenance of Rotarod.
2.0
SCOPE
This Standard Operating Procedure
(SOP) is applicable for the operation, maintenance, and calibration of the tail
flick for Pharmacology studies.
3.0
RESPONSIBILITY
3.1 All personnel in the Pharmacology Department shall be
responsible for the implementation and compliance with the SOP.
3.2 Head/Designees/Study Directors of the Pharmacology
department shall be responsible to ensure the implementation and compliance with
the SOP.
4.0
DISTRIBUTION
4.1 The Quality Assurance department is responsible to keep
SOP ‘Master Copy’ approved through Quality Assurance.
4.2
The copy of the ‘Control Copy’ of SOP is being distributed
in the Pharmacology department and placed near related Equipment/ Instrument as Display
Copy.
5.0
DEFINITION(S)
5.1
Definition(s)
5.1.1 The Rotarod performance test is a
performance test based on a rotating rod with forced motor activity being
applied, usually by a rodent. The test measures parameters such as riding time
(seconds) or endurance.
6.0 ABBREVIATION(S)
6.1 Abbreviation(s)
6.1.1
SOP : Standard Operating Procedure
6.1.2 PPEs : Personal Protective Equipment
7.0
PROCEDURE
7.1
Introduction
7.1.1 The ROTAROD units facilitate easy testing of motor activity in rodents. Drug administration,
central nervous system damage, and disease effects on motor coordination or
fatigue can be assessed by measuring the time during which the animal continues
walking in a rotating rod.
6.1.2 A
central rod (made of Perspex, and knurled to provide adequate grip) divided into
sections, rotates at either a constant speed or a steadily accelerating rate
(from 4 to 40 rpm in an interval of time ranging from 30 seconds to 10 minutes).
The use of acceleration has been reported to produce less variable data.
Figure 1: Rotarod Apparatus
6.2
Instrument
description
6.2.1
Front panel
7.2.1.1
Start/Stop button: to start or stop the experiment.
7.2.1.2
Working mode selector: to choose the desired working
mode, constant or acceleration.
7.2.1.3
Rod speed indicators: show the current and set rod
rotation speed.
7.2.1.4
Lane counters: depending on the rotarod model,
this will be from two to five columns
7.2.1.4.1
The first row displays the time that the animals have been on the
rod.
7.2.1.5
The second row displays the rod speed.
Figure 2: Front Panel
7.2.1.6
Hold ramp button: functional during the Acceleration working mode:
7.2.1.6.1
First press of this button starts the rod acceleration and begins
the active lane counters recording.
7.2.1.6.2
second press maintains the current rod speed constant
7.2.1.7
Settings screen: Displays the settings screen.
7.2.1.8
Reset button: Clears the lane counters indicators to
zero.
7.2.1.9
Speed increase/decrease button: Increments/decrements by one
unit the rod speed. Applies only to constant speed working mode.
7.2.1.10
Speed screen: Displays the Set Speed screen. Applies
only to constant speed working mode.
7.2.1.11
Ramp screen: Displays the Set Ramp screen. Applies
only to acceleration working mode.
6.2.2
Rear panel
7.2.1.12
USB: USB B-type female connector is used to
connect the Rotarod to a computer USB port. Data is sent to the Sedacom
software through this connector.
7.2.1.13
Power: Male panel jack used to connect the
Rotarod to the AC-DC adapter.
7.2.1.14
Switch: The main switch is used to turn on and off the Rotarod.
Fig 3: Rear panel
6.3
Equipment
connection
The connections and necessary
cables are listed in the following table
Figure 6: Equipment
connection
6.4
Operation
Fig 4: Main Part
6.4.1
Once
the equipment is turned on, the start-up screen appears after a few seconds:
Figure 7: Start-Up Screen
6.4.2
Changing settings
7.2.1.15
The following options can be modified
through the Settings screen.
7.2.1.15.1
Motor direction: Establishes the rod
rolling direction: forward and reversed.
Figure 8: Setting Option
7.2.1.15.2
Display backlight: Establishes the percentage
of the backlight of the display.
Figure 9: Backlight Settings
Screen
7.2.1.15.3
Upgrade software: Used to update the
firmware to the latest version.
7.2.1.15.4
System Information: Displays the Rotarod model type, the firmware
version number, and the serial number of the device.
6.4.3
Selection of the working mode
7.2.1.16
Constant mode
7.2.1.16.1 The motor works to provide
a speed from 4 to 40 rpm, depending on the needs of the user, who can select
the speed either before starting the experiment or when it is already underway.
The speed is the same for each lane, as there is a single motor driving the
single rod on which all the subjects are moving.
7.2.1.16.2 The starting point of the timers is determined when the relevant lever is lifted; each timer works independently from the others. The endpoint of each TIMER is determined when the relevant lever is lowered.
Figure 10: Screen in Constant mode
7.2.1.17
Acceleration mode
7.2.1.17.1 The motor increases speed
automatically and linearly from 4 up to 40 rpm, thus producing a constant
acceleration of the rod. The speed is the same for each lane, as there is a
single motor driving the single rod on which all the subjects are moving.
7.2.1.17.2 It is possible to determine
the acceleration TIME that the rod will take to achieve the maximum speed. The
time slope may range from 30 seconds up to 10 minutes.
7.2.1.17.3 Once the maximum has been
achieved, the rod continues at this speed. When working in this mode, two states
and one optional state exist:
Figure 11: Screen in Acceleration Mode Waiting
7.2.1.17.4 This is the first state
after pressing the Start button in this mode.
7.2.1.17.5 This is the preparation
state of the instrument in order to place the subjects in their respective slot
on the rod. The speed of the rod for this phase of preparation is a minimum, of 4 rpm.
7.2.1.17.6 Levers must be lifted to be
ready to begin the experiment.
6.4.4
Acceleration
Figure 12: Screen in Acceleration
Mode Running
7.2.1.18
This is the second state after pressing
the Hold Ramp button in this mode.
7.2.1.19
The motor ramps from 4 to 40 rpm,
following the predetermined SLOPE TIME once the Hold Ramp button has been
pressed.
7.2.1.20
After the ramp is initiated, the
ACCELERATION TIME cannot be changed.
7.2.1.21
The TIMER counters corresponding to the
raised LEVERS begin recording the time after the Hold Ramp button is pressed.
7.2.1.22
Once the relevant LEVER is depressed,
the TIMER will show the total rotating time as the end point for that subject
on that particular trial.
7.2.1.23
When the last subject falls from the
rod, the instrument automatically enters the STOP state
6.4.5
Preparing the instrument
6.4.5.1
Holding the animal by its tail, swing
them up from a position lower than the rod so that they find itself
walking inside.
6.4.5.2
It is advisable to set the cylinder in
motion before placing the animals on the rod. Otherwise, by the time the last
animal has been seated in position, the first animal may be facing the wrong
direction.
6.4.5.3
It is also advisable to let the animal
familiarize themselves with the rod by having them walk at minimum speed for a few
moments.
Figure 13: Placing The Animal In
The Roll
6.4.6
Conducting experiments
6.4.6.1
Turn on the instrument using the POWER
switch.
6.4.6.2
Lower the LEVERS, this will prevent the timer
counters from the beginning.
6.4.6.3
Select the desired working mode.
6.4.7
Constant
6.4.7.1
Establish the desired initial rod speed
using the “+”, “-” or “Speed” buttons. The rod will not begin rotating until the
“Start” button is pressed.
Figure 14: Set rod speed screen
6.4.7.2
Press the “Start” button and this will
initiate the rod rotation at the set speed in step 1.
6.4.7.3
If any LEVER remains in the raised
position, the message “Place all levers down” will be shown on the display
until all the LEVERS are in a depressed position.
6.4.7.4
Place the animals in their respective
lanes on the rod
6.4.7.5
Manually lift the LEVERS to activate the
relevant counters for the lanes that will be used in that trial.
6.4.7.6
Use the “+”, “-”or “Speed” buttons to
modify the desired rod speed.
Figure 15: Display running in
Constant mode
6.4.8
Acceleration
6.4.8.1
The DISPLAY will indicate that the unit
remains in WAITING mode. Now select the RAMP TIME using the “Ramp Time” (the
acceleration is automatically calculated from this time to reach max rotation
speed).
6.4.8.2
Press the “Start” button to initiate the
rod rotating at the initial constant speed, 4 rpm.
6.4.8.3
Place the animals in their respective
lanes on the rod.
6.4.8.4
Manually lift the LEVERS for the
respective lanes that will be used. If there are no LEVER lifted when the “Hold
Ramp” button is pressed in the next step, the display will show the message
“Raise levers to activate lanes” until at least one LEVER is lifted.
Figure 16: Set Ramp Time Screen
Figure 17: Display waiting for
activating levers in accelerated mode
6.4.8.5
Press the “Hold Ramp” button. The lifted
lane counters start counting time and the ramp starts accelerating the rod
rotation.
Figure 18: Display running in
accelerated mode
6.4.8.6
When the last subject falls from the
rod, the instrument automatically enters the STOP state.
6.4.8.7
START/STOP BUTTON is pressed again, it
will interrupt the experiment, halting the acceleration process. The display
will continue to show the last time at any of the subjects fell from the rod.
6.5
Maintenance
Figure
19: Maintenance
6.5.1
Cleaning the unit
6.5.1.1
Levers force adjustment
6.5.1.1.1 On
the back of the levers there is a screw that is used for adjusting the strength
of the levers. If screwed (clockwise), the separation between the lever and the
magnet that holds the lever will increase. This will decrease the attraction
and the lever will be more sensitive to the detection of falls. If on the contrary,
it is unscrewed (counterclockwise) the distance to the magnet that holds it
will decrease, which will result in an increase in the strength of the lever
and less sensitivity to detect a fall from the rod.
7.0
PRECAUTIONS
7.1 All PPEs like aprons, gloves, masks, head caps, and shoe
covers should be worn before using the instrument.
7.2
Press Keys softly.
7.3
Clean the equipment with a dry piece of robe or a
lightly moistened one, do not use organic solutions.
7.4
Never use abrasive products or dissolvents.
7.5
Never pour water or liquids on the equipment.
7.6 Do not use organic solvents to clean the unit, as they
may damage the Perspex sheets or crack the acrylic front panel of the liquid
crystal display.
7.7
A dampen cloth and water can be used to clean the
equipment and then a dry cloth should be used to dry it.
8.0
REFERENCES
8.1
Reference
8.1.1
Refer to the manufacturer’s manual for additional information.
END OF THE DOCUMENT
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