EXPERIMENT NO.
08
OBJECT: To study muscle relaxant property of diazepam in rat using Rota rod apparatus.
REFERENCE: Kulkarni, S.K., “Hand book of experimental pharmacology”, Published
by Vallabh Prakashan, Page no. 122.
REQUIREMENTS: Animal – Rat
Drug - diazepam
Apparatus – Rota Rod
PRINCIPLE: One of the important pharmacological actions of antianxiety agent of
benzodiazepines class is muscle relaxant property. These agents reduce anxiety and
tension. The loss of muscle grip is an indication of muscle relaxation. These agents can
easily study in Rotarod apparatus.
Rota rod
The Rota rod apparatus consists of a metal rod (3 cm diameter) coated with rubber
attached to a motor with the speed adjusted to 2 rotations per minute. The rod is 75 cm in
length and is divided into 3sections by metallic discs, allowing the simultaneous testing of
3 mice. The rod is in a height of about 50 cm above the tabletop in order to discourage the
animals from jumping off the roller. Cages below the section serve to restrict the
movements of the animals when they fall from the roller.
PROCEDURE
o Weighed the animals.
o Turn on the Rota rod and select an appropriate speed (20-25 rpm ).
o Placed the animals on rata rod and noted the fall down time when the mice falls
from rotating rod.
o Inject Diazepam to the animal, after 30 min repeated the experiment and noted the
fall down time.
o Compare the falloff time of animals before and after diazepam treatment
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Dose Calculation:
Observation Table:
S.No. Body wt. Dose Fall down time % in time
Before After
1
2
3
4
5
Comments:
RESULT:
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EXPERIMENT NO. 09
OBJECT: To study the analgesic effect of Diclofenac sodium in rat using Tail Flick
method.
REFERENCE: Kulkarni S.K. “Hand book of experimental pharmacology” 3rd Edition,
2005. Published by Vallabh Prakashan, Page No-123-124.
REQUIREMENTS:
Animal Rat
Equipment Analgesiometer
Drug Diclofenac sodium (Dose 10mg/kg).
PRINCIPLE:
Analgesia is defined as a state of reduced awareness to pain. Analgesics are substance
which decreases pain sensation by increasing threshold to painful stimuli. The
commonly used analgesic is aspirin, paracetamol (non-narcotic) and morphine (narcotic
type).
Painful reaction in experimental animals can be produced by applying noxious stimuli
such as;
Thermal
Chemical
Physical pressure
But the major procedure used in tail flick method using analgesiometer, hot plate and
acetic acid induced writhing method
PROCEDURE:
• Weigh and number the animals.
• Take basal reaction time to radiant heat by placing the trip of tail on radiant heat
source, the tail withdrawal from the heat (flicking response) is taken as the end
point.
• Normally rat withdrawals its tail within 3-5sec. A cut off period of 10-12 sec is
observed to prevent damage to the tail.
• Any animal falling to withdrawals its tail within 3-5sec is rejected from the
study.
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• Take a basal 3-5 basal reaction time for each mouse at gap of 5 min to conform
normal behavior of the animal.
• Injected Diclofenac sodium and noted the time at 5, 10, 15 and 30 min after the
drug. As the reaction time reaches to 1o sec .it is considered maximum analgesia
and the tail is removed from the source of heat to avoid tissue damage.
• Calculate the percentage the increase in reaction time.
Dose Calculation:
Observation Table:
S. No. Body wt. Dose Analgesic activity Average percentage increase
Before After
1
2
3
Comments:
RESULT:
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EXPERIMENT NO. 10
OBJECT: To study the anxiolytic (antianxiety) effect of diazepam in rat using elevated
Plus Maze Apparatus.
REFERENCE: Kulkarni S.K. “Hand book of experimental pharmacology” 3rd edition,
2005. Published by Vallabh Prakashan, page no-135-136.
REQUIREMENTS:
Animal Rat (150-200gm).
Drug Diazepam (2mg/kg).
Equipment Plus-maze apparatus (2 open & 2 closed arm)
PRINCIPLE:
Anxiety is the major CNS disorder, different class of benzodiazepines used to treat the
anxiety disorder. In experimental way, the elevated plus maize is most simple apparatus
to study anxiolytic effect of almost all type of anti-anxiety agents. Exposure to animal to
novel maze alley evokes an approach-avoidance conflict which is stronger in open arm
as compared to enclosed arm. Rodents have aversion for high and open space and prefer
enclosed arm, and therefore spend the greater amount of time in enclosed arm. When
animal enter in open arm, they freeze and become immobile, defecate and show fear like
movements. The plasma cortisol level is also reported to be increased, as a true
reflection of anxiety.
PROCEDURE:
1) Weighed & no the animals, divide them into 2 groups each consisting of 6 mice, one
group is used as control & other for drug treatment.
2) Placed animal individually in center of maze , head facing towards open arm and start
the stop watch and note following parameter for five minute
a) First presence of mouse to open and enclosed arm.
b) Number of entries in open & enclosed arm (an arm entry defined as entry of four
paws into the arm)
c) Average time each animal spend in each arm. (average time=total duration in
arm/no. of entries
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3) Injected diazepam to test gp .After 30 min place the animal individually in the centre of
maze & note all parameters.
4) Compare the preference of the animal to open and enclosed arm. Average time spent in
open arm & no of entries in open arm in each group.
Comments:
RESULT:
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EXPERIMENT NO. 11
OBJECT: To study the analgesic effect of Diclofenac Sodium in rat by Hot Plate.
analgesiometer.
REFERENCE: Kulkarni, S.K., “Hand book of experimental pharmacology”, Published
by Vallabh Prakashan, Page no. 125.
REQUIREMENTS: Animal - Rat
Drug- Diclofenac-sodium
Equipment – EDDY’S Hot plate
PRINCIPLE: Analgesia is defined as a state of reduced awareness of pain. In this
method heat is used as a source of pain. Animals are individually placed on a hot plate
maintained at constant temperature (550C) and the reaction of animals, such as paw
licking or jump response is taken as the end point. Analgesics increase the reaction-time.
PROCEDURE:
Weigh and number the rat.
Take the basal reaction-time by observing hind paw licking or jump
response (whichever appears first) in animals when placed on the hot plate
maintained at constant temperature (550C).
Normally animals show such response in 6-8 sec. A cut off period of 15 sec
is observed to avoid damage to the paws.
Inject morphine to animals and note the reaction time of animals on the hot
plate at 15, 30, 60 and 120 min after the drug administration.
As the reaction time increases with morphine, 15 sec is taken as maximum
analgesia and the animals are removed from the hot plate to avoid injury to
me paw.
Calculate percent increase in reaction -time (as index of analgesia) at each
time interval.
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Dose Calculation:
Observation Table:
S. No. Body Dose Basal Reaction Time Basal Reaction Time
Weight
Paw Paw Paw Paw
Licking jumping Licking jumping
1
2
3
4
Comments:
RESULT:
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