Anxiolytics & Screening Models Made By - Adarsh Gaur M.Pharm Research Scholar (Pharmacology) NIET, Pharmacy Institute, Greater Noida

1. SCREENING MODELS &
METHODS OF ANXIOLYTICS
BY – ADARSH GAUR
NIET, PHARMACY INSTITUTE
M. PHARM (I-SEMESTER), PHARMACOLOGY
GUIDED BY- DR. SAUMYA DAS
PROFESSOR, H.O.D (PHARMACOLOGY)

2. ANXIETY ?
• Anxiety is a normal emotional response in animals.
• It is the body's way of responding to stressors or potential threats.
• In animals, anxiety can manifest in various ways, such as changes in
behavior, physiological responses, and changes in hormone levels.
• Some common signs of anxiety in animals include:- Restlessness,
agitation, panting, trembling, and hiding.

3. ANXIOLYTICS?
- Anxiolytics are minor tranquilizers and a class of medications that
are used to treat anxiety and anxiety-related disorders.
- They work by altering the levels of certain chemicals in the brain that
are involved in regulating mood and emotions.
- Common examples of anxiolytics include benzodiazepines (such as
diazepam, alprazolam, lorazepam), buspirone and some
antidepressants such as SSRI’s, i.e. Selective serotonin reuptake
inhibitors.

4. PARAMETERS ?
When performing in vivo (animal) tests to evaluate the potential efficacy of
new compounds as anxiolytic agents, several parameters should be
observed:
- Behavioral measures - Physiological measures
- Dosage - Timing of treatment
- Ethical considerations - Face validity
- Predictive validity

5. EPM (Elevated Plus Maze) test
- The EPM consists of two open
arms and two closed arms,
arranged in a plus shape and
elevated off the ground.
- The setup of the EPM is as follows:
. The two open arms are typically
illuminated and have no walls or
barriers, allowing the animal to see
the surrounding environment.
. The two closed arms are typically
dark and have walls or barriers,
preventing the animal from seeing the
surrounding environment.

6. PROCEDURE
- The animal is placed in the center of the maze, and its behavior is observed
and recorded. The animal's behavior is typically monitored and recorded for a
specific period of time, typically 5-10 minutes.
- The parameters typically measured are:
1. Time spent in open arms: Animals with high levels of anxiety will spend less
time in the open arms
2. Time spent in closed arms: Animals with high levels of anxiety will spend
more time in the closed arms
3. Number of entries into open arms: Animals with high levels of anxiety will
make fewer entries into open arms.
4. Number of entries into closed arms: Animals with high levels of anxiety will
make more entries into closed arms.

7. PREPARATIONAL MEASURES
- SD Rats (200-250 gm) are housed in pairs for 10 days prior to testing in the
apparatus. (Acclimatization).
- 6 rats are taken for each group.
- 30 mins after administration of the test drug or standard, the rat is placed in
the center of the maze facing one of the open arms.
- Duration of the test is 5 mins where all data is collected.
- Keep in mind for scenarios to avoid variations like test should be performed
in sound proof room and the maze should be cleaned by wet tissue paper
(10% alcohol solution).
- Anxiolytics specifically increase the no. of entries in the open arm entries and
the time spent there.

8. Light Dark Exploration Model
-The light-dark exploration test is a behavioral test used to
measure anxiety-like behavior in animals.The test consists of
a light compartment and a dark compartment, separated by
a small opening.
-The setup of the test is as follows:
.The light compartment is typically brightly lit and has a
white or light-colored floor.
.The dark compartment is typically dark and has a black or
dark-colored floor, while the walls and ceiling are also black.
A small opening (typically around 7cm x 7cm) connects the
light and dark compartments.
. Both chambers/cages made of polypropylene equipped
with photocells, carries an animex activity monitor, that
monitors the locomotor activity count.

9. PROCEDURE
- Crawley and Goodwin (1980)
- Mice and rats have a tendency to explore novel environment.
- A 2 chambered system, where the animal can freely move between bright
and dark areas, they show more crossings between 2 chambers and more
locomotor activity treatment with anxiolytics.
- Animals are treated 30 mins before the experiment with test or vehicle
intraperitoneally (I.P) and are observed for 10 mins.
- Group of 6-8 are used for each dose.
-Evaluation – No. of crossings through the partition between light and dark
chamber are compared with total activity counts during 10 mins.

10. PREPARATIONAL MEASURES
The animal is placed in the light compartment and its behavior is observed
and recorded.
- The animal's behavior is typically monitored and recorded for a specific
period of time, typically 5-10 minutes.
- The parameters typically measured are:
- Time spent in light compartment: Animals with high levels of anxiety will
spend less time in the light compartment
- Time spent in dark compartment: Animals with high levels of anxiety will
spend more time in the dark compartment
- Number of transitions between compartments: Animals with high levels of
anxiety will make fewer transitions between compartments.

11. VogelWater Lick ConflictTest
-TheVogel conflict test, also known as the Vogel et al water lick test, is a
behavioral test used to measure anxiety-like behavior in animals.The test
is based on the principle that animals will work to obtain a reward (such as
water) even in the presence of an aversive stimulus (such as an electric
shock).
-The setup which is also known as anxiometer of the test is as follows:
-The animal is placed in a cage and has access to a water spout.
- An electric shock is delivered to the cage floor at random intervals, and
the animal is required to lick the spout to obtain water.
-Thirsty animals gain water reward through a water spout, but at the
expense of receiving a mild electric shock delivered to the tongue.

12. PROCEDURE
- The parameters typically measured are:
- Number of licks: Animals with high levels of anxiety will make fewer licks
- Latency to lick: Animals with high levels of anxiety will have a longer latency to lick
- Lick rate: Animals with high levels of anxiety will have a lower lick rate
- Number of punished licks: Animals with high levels of anxiety will make fewer
punished licks (licks made during the shock).
- Licking in untreated/controls is suppressed due to the conflict.
- Diazepam and pentobarbital produces a significant anti conflict effect, which
means that, these drugs increases the no. of electric shocks the mice received during
the test session.

13. PREPARATIONAL MEASURES
- Shock intensity:The intensity of the electric shock should be determined based on previous
studies, and the effects of different intensities should be evaluated.
Shock frequency:The frequency of the electric shock should be determined based on previous
studies, and the effects of different frequencies should be evaluated.
Control groups: Control groups should be used to compare the effects of the compound to a
placebo or a standard anxiolytic drug.
Sample size: Adequate sample size should be used to ensure that the results are statistically
significant and representative.
Ethical considerations: Animal welfare should be taken into account and the study should be
approved by an ethical review board before starting.
Lick parameters: Number of licks, latency to lick, lick rate, number of punished licks should be
measured and analyzed.

14. REFERENCES
H. Gerhard Vogel (Ed.) (2002) “Drug Discovery and Evaluation – Pharmacological Assays”. 2nd Edition.
Springer-Verlag Berlin Heidelberg Publishers, NewYork. PP – 430-441
Michel Bourin. MD, PhD. (September 17, 2015). Animal models for screening anxiolytic-like drugs: a
perspective. Retrieved from NLH. https://www.ncbi.nlm.nih.gov/pmc/articles/PMC4610614/