The document summarizes a presentation on the preclinical evaluation of antidepressants. It discusses the objectives of reviewing information on depression and antidepressants, surveying standard antidepressant drugs on the market, and studying the pharmacokinetics, pharmacodynamics, adverse drug reactions, and uses of antidepressants. It also explains in vivo and in vitro models for preclinical evaluation, including behavioral tests, activity-based tests, and screening methods involving neurotransmitter uptake inhibition and receptor binding assays. Common tests discussed include the forced swim test in rats or mice, tail suspension in mice, and learned helplessness in rats.

1. 1
““A REVIEW ON PRECLINICALA REVIEW ON PRECLINICAL
EVALUATION OF ANTIDEPRESSANTS”EVALUATION OF ANTIDEPRESSANTS”
PRESENTING BY,PRESENTING BY,
Ms. SONALI B. DIWATEMs. SONALI B. DIWATE
GUIDED BY,GUIDED BY,
PROF. R. CHANSHETTIPROF. R. CHANSHETTI
MODERN C.O.P.,MOSHI, PUNE-412105MODERN C.O.P.,MOSHI, PUNE-412105

2. To Review and Study the Preclinical Evaluation of AntidepressantsTo Review and Study the Preclinical Evaluation of Antidepressants
OBJECTIVEOBJECTIVE
1. COLLECT THE INFORMATION ABOUT DEPRESSION & ANTIDEPRESSANTS.1. COLLECT THE INFORMATION ABOUT DEPRESSION & ANTIDEPRESSANTS.
(LITERATURE SURVEY).(LITERATURE SURVEY).
2. SURVEY THE STATUS AND STANDARD ANTIDEPRESSANT DRUGS AVAILABLE IN2. SURVEY THE STATUS AND STANDARD ANTIDEPRESSANT DRUGS AVAILABLE IN
MARKET.MARKET.
3. STUDY THE COMMON ADR & USES OF ANTIDEPRESSANTS.3. STUDY THE COMMON ADR & USES OF ANTIDEPRESSANTS.
4. STUDY THE PHRMACOKINETICS,PHARMACODYNAMICS OF ANTIDEPRESSANTS.4. STUDY THE PHRMACOKINETICS,PHARMACODYNAMICS OF ANTIDEPRESSANTS.
5. EXPLAIN INVIVO & INVTRO MODELS FOR PRECLINICAL EVALUATION OF5. EXPLAIN INVIVO & INVTRO MODELS FOR PRECLINICAL EVALUATION OF
ANTIDEPRESSANTS.ANTIDEPRESSANTS.
6. ANALYZE ALL THE PARAMETERS RELATED TO ANTIDEPRESSANTS6. ANALYZE ALL THE PARAMETERS RELATED TO ANTIDEPRESSANTS
7.INVESTIGATE RESEARCH SCOPE IN THIS AREA.7.INVESTIGATE RESEARCH SCOPE IN THIS AREA.
8. AWARE PEOPLE ABOUT DEPRESSION & ANTIDEPRESSANTS.8. AWARE PEOPLE ABOUT DEPRESSION & ANTIDEPRESSANTS.
9.SUMMERIZATION OF THE INFORMATION.9.SUMMERIZATION OF THE INFORMATION. 2

3. CONTENTS…..CONTENTS…..
1. AIM AND OBJECTIVE1. AIM AND OBJECTIVE
2.INTRODUCTION TO DEPRESSION & ANTIDEPRESSANTS2.INTRODUCTION TO DEPRESSION & ANTIDEPRESSANTS
3.STANDARD ANTIDEPRESSANTS AVAILABLE IN MARKET3.STANDARD ANTIDEPRESSANTS AVAILABLE IN MARKET
4. COMMON ADR & USES OF ANTIDEPRESSANTS4. COMMON ADR & USES OF ANTIDEPRESSANTS
5. PHARMACOKINETICS & PHARMACODYNAMICS5. PHARMACOKINETICS & PHARMACODYNAMICS
6. PRECLINICAL SCREENING METHODS6. PRECLINICAL SCREENING METHODS
7. SUMMERY7. SUMMERY
8. CONCLUSION8. CONCLUSION
9. BIBLIOGRAPHY9. BIBLIOGRAPHY
3

4. INTRODUCTION TO DEPRESSIONINTRODUCTION TO DEPRESSION[3]
DEPRESSION is a word used in many different contexts and can cover many different
meanings. in a medical context the term depression is used to describe a syndrome,
that is a cluster of symptoms which often occur together, but where the underlying
causes may vary.
The Symptoms of Depression
1. Decreased energy, fatigue, feeling “slowed down”
2. Depressed mood
3. Difficulty in concentrating or making decisions
4 .Feeling restless.
5. Feelings of worthlessness or inappropriate guilt
6. Insomnia, early-morning awakening, or oversleeping
7. Loss of interest or pleasure in hobbies, work
8. Recurrent thoughts of death or suicide; suicide attempts
5
TYPES OF DEPRESSION
UNIPOLAR (EMOTIONAL SYMPTOMS)
BIPOLAR (BIOLOGICAL SYMPTOMS)

5. HISTORY OF ANTIDEPRESSANTSHISTORY OF ANTIDEPRESSANTS
The drugs or agents which are used in treatment of depression is called as
antidepressants
Antidepressants acts by balancing brain neurotransmitters level to ease
depression.
CLASSIFICATION OF ANTIDEPRESSANTS
6
INTRODUCTION TO ANTIDEPRESSANTSINTRODUCTION TO ANTIDEPRESSANTS[1]
1. TCAs: Amitriptyline, Imipramine, Clomipramine, Nortriptyline, Desipramine.
2. Tetracyclic antidepressants: Mianserin, Maprotiline.
3. SSRIs: Fluoxetine, Paroxetine, Sertraline, Fluvoxamine, Citalopram, Escitalopram.
4. SNRIs: Venlafaxine/ Venlafaxine XR, Duloxetine.
5. MAOIs: Moclobemide, Phenelzine, Isocarboxazid, Tranylcypromine.
6. Lithium salts: Lithium carbonate.

6. STANDARD DRUGS AVAILABLE IN MARKETSTANDARD DRUGS AVAILABLE IN MARKET[5]
7

7. COMMON ADVERSE DRUG REACTIONCOMMON ADVERSE DRUG REACTION[5]
8

8. DRUG INTERACTIONS [2]

9. CONTRAINDICATIONS [2]

10. The mechanism of action of tricyclic antidepressants
Tricyclic antidepressants (TCAs) are a group of drugs used to treat affective, or
‘mood’, disorders. Mood disorders are associated with reduced levels of
monoamines in the brain. TCAs binding to 5-HT and noradrenalin re-uptake
transporters prevents the re-uptake of these monoamines from the synaptic cleft
and their subsequent degradation. This re-uptake blockade leads to the
accumulation of 5-HT and noradrenalin in the synaptic cleft and the concentration
returns to within the normal range.
The mechanism of action of SSRI antidepressants
• SSRIs ease depression by affecting naturally occurring chemical messengers
(neurotransmitters), which are used to communicate between brain cells. SSRIs block
the reabsorption (reuptake) of the neurotransmitter serotonin in the brain. Changing
the balance of serotonin seems to help brain cells send and receive chemical
messages, which in turn boosts mood.
• Most antidepressants work by changing the levels of one or more of these
neurotransmitters. SSRIs are called selective because they seem to primarily affect
serotonin, not other neurotransmitters. 10
PHARMACOKINETICS & PHARMACODYNAMICS OF ANTIDEPRESSANTSPHARMACOKINETICS & PHARMACODYNAMICS OF ANTIDEPRESSANTS[
3]
1) GIVEN BY ORAL ROUTE
2) METABOLISM BY LIVER
3) EXCREATION THROUGH URINE

11. PHARMACODYNAMICS OF ANTIDEPRESSANTSPHARMACODYNAMICS OF ANTIDEPRESSANTS
The mechanism of action of SNRI antidepressants
• Serotonin (ser-o-TOE-nin) and norepinephrine (nor-ep-ih-NEF-rin) reuptake
inhibitors ease depression by affecting chemical messengers (neurotransmitters)
used to communicate between brain cells. Like most antidepressants, SNRIs work
by changing the levels of one or more of these naturally occurring brain chemicals.
• SNRIs block the absorption (reuptake) of the neurotransmitters serotonin and
norepinephrine in the brain. They also affect certain other neurotransmitters.
Changing the balance of these chemicals seems to help brain cells send and
receive messages, which in turn boosts mood. Medications in this group of
antidepressants are sometimes called dual-action antidepressants.
The mechanism of action of Lithium Salt
• Lithium may also increase the release of serotonin by neurons in the brain.
• The excitatory neurotransmitter glutamate could be involved in the effect of
lithium.
• The other mechanisms by which lithium might help to regulate mood include the
alteration of gene expression.

12. SCREENING METHODSSCREENING METHODS [11][11]
11
IN VIVO METHODS IN VITRO METHODS
BEHAVIORAL
TESTS
ACTIVITY DEPENDS
UPON MOA

13. CLASSIFICATION OF SCREENING METHODSCLASSIFICATION OF SCREENING METHODS
IN VIVOIN VIVO METHODSMETHODS
BEHAVIORAL TESTS
1) Catalepsy antagonism in
chicken
2) Despair swim test
3) Tail suspension in mice
4) Learned helplessness in rats
5) Muricide behavior in rats
6) Behavioral changes after
neonatal Clomipramine
treatment
ACTIVITY DEPENDS ON MOA
1) Potenciation of NE toxicity in rats
2) Compulsive gnawing in mice
3) Reserpine induced hypothermia
4) Tetrabenazine antagonism
5) Yohimbin toxicity enhancement
6) Apo morphine induced hypothermia
7) 5HTpotentiation in rats
8) Tryptamine potentiation in rats
9) Syrotonine syndrome in rats
10) Sexual behavior in rats
13

14. CLASSIFICATION OF SCREENING METHODSCLASSIFICATION OF SCREENING METHODS
IN VITROIN VITRO METHODSMETHODS
1) Inhibition of NE uptake in rat brain
2) Inhibition of Dopamine uptake in rat striatal
3) Inhibition of Serotonin uptake in synapsome
4) Binding to monoamine transporters
5) Antagonism of p-chloramphetamine toxicity by inhibition of
seronine uptake
6) Measurement of B-adrenoreceptor stimulated adenyl
cyclase
7) Tests for anticholinergic properties in rat brain
8) Moa inhibitors in rat synaptosomes
Type-a
Type-b 14

15. CATALEPSY ANTAGONISM IN CHICKENCATALEPSY ANTAGONISM IN CHICKEN
 Adult white leghorn chickens are used
 Grasped the animal & turn it on its back & hold it for 1 min
 Cataleptic numbness occurs
 Remove the hand carefully
 It remains in cataleptic state
 Clapping of hands above the head arouses the chicken which
jumps up & runs away
 Pretesting is done to ensure about cataleptic behavior
 Control studies shows that, in untreated animals this
phenomenon could be elicited 6 times (every 30 min for 5 days)
 Inject test compound I.P.
 The test is performed for 4 times (every 30 min during 2 hrs)
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PURPOSE
This method is used
to screen the antidepressant
Activity in chickens
EVALUATION
1. If cataleptic rigor does not occur
after treat.=Test +ve
OR
it is interrupted within 1 Min at
least twice during 2 hrs Period.
2. Arousal after hand clapping Or
pulling on the wings is recorded to
register central Sedative effect.
3. To obtained DRC 12 Animals/GRP
are treated.
4. ED50 is calculated.

16. DESPAIR SWIM TESTDESPAIR SWIM TEST
 ADULT MALE SPRAGUE-DAWLEY RATS (WT=160-180gm)ARE USED.
 NAÏVE RATS ARE INDIVIDUALLY FORCED TO SWIM INSIDE A
VERTICAL PLEXIGLAS CYLINDER (HIGHT=40 cm , DIA.=18cm)
 RATS ARE PLACED IN CYLINDER VIGOROUSLY SWIMMING IN
CIRCLE (TEMP.=25’C)
(water level=15 cm)
 TRYING TO CLIMB ON WALLS OF CYLINDER & AFTER 2-3
MIN,ACTIVITY BEGINS TO SUBSIDE & FLOTING OF INCREASING
LENGTH
 AFTER 5-6 MIN IMMOBILITY REACHES A PLATEAU (RATS REMAINS
IMMOBILE)
 AFTER 15 MIN ,REMOVE RATS.ALLOW TO DRY THEM (32’C)
 RATS ARE PLACED IN CYLINDER & IMMOBILITY IS CHECKED DURING
5 MIN IT HAS BEEN FOUND TO BE REPRODUCIBLE IN DIFF.GRP
 IMMOBILE ANIMAL=WHICH REMAINS FLOATING PASSIVELY IN
WATER IN A SLIGHTLY HUNCHED BUT UPRIGHT POSITION
 INJECT TEST TEST/STD DRUG BEFORE 1 hr TESTING i.p.
PURPOSE
1. It was suggested that mice/rats Forced
to swim in restricted space From which
they can not escape are Induced to
characteristic behavior Of immobility.
2. This behavior reflects a state of
Despair which can reduced by several
Agents which are therapeutically
Effective in human depression.
EVALUATION
1. Duration of immobility is measured in
Controls & treated animals with
Various Doses of test drug & standard
2. Antidepressants acts like stimulants
Like amphetamine which reduces
Duration of immobility
3. Dose response can be evaluated.

17. TAIL SUSPENSION IN MICETAIL SUSPENSION IN MICE
 MALE BALB/CJ MICE(WT=20-25gm)ARE USED.
 ANIMALS ARE TRANSFER INTO TESTING AREA TO
ADAPT NEW ENVIRONMENT 1 hr BEFORE TESTING
 GROUP OF 10 ANIMALS TREATED WITH TEST
COMP./VEHICLE BEFORE 30 MIN OF TESTING(i.p)
 FOR THE TEST MICE ARE SUSPENDED ON THE EDGE
OF SHELF 58cm ABOVE A TABLE TOP BY ADHESIVE
TAPE PLACED 1cm FROM TIP OF THE TAIL.
 THE DURATION OF IMMOBILITY IS RECORDED FOR
PERIOD OF 5 MIN.
 MICE ARE CONSIDERED AS IMMOBILE WHEN THEY
HANG PASSIVELY & COMPLETELY MOTIONLESS FOR
ATLEAST 1 MIN.
PURPOSE :
1. The immobility displayed by rodents , when
Subjected to unavoidable & inescapable
Stress has been hypothesized to reflect
Behavioral distress which in turn may
Depressive disorder in humans.
2. Clinically effective antidepressants reduces
The immobility that mice display after active
& Unsuccessful attempts to escape when
Suspended by the tail.
EVALUATION :
1. % of animals showing passive Behavior is
counted & compared with Vehicle treated
control.
2. Using various doses ed50 value can
Be calculated.

18. Learned Helplessness in RatsLearned Helplessness in Rats
 MALE SPRAGUE RATS (WT=300gm)ARE USED.
 LEARNED HELPLESSNESS IS PRODUCED BY EXPOSURE TO ELECTRIC SHOCK
FOR 1 hr (0.7mA) ON SCHEDULE OF 10 S OF SHOCK/MIN
 APPARATUS IS A 30*45*30 cm BOX WITH A GRID FLOOR
(FLOOR=7.5*7.5 cm) (HIGHT=20 cm)
 THE PLATFORM CAN BE INSERTED THROUGH ONE SIDE WALL TO ALLOW A
JUMP-UP ESCAPE RESPONSE.
 AFTER APPROPRIATE TREATMENT,THE ANIMALS ARE TESTED FOR
ACQUISITION OF JUMP-UP ESCAPE IN THE SAME APPARATUS.
 AT THE BEGINNING OF TRIAL ,PLATFORM IS PUSHED INTO THE BOX
 0.4mA SHOCK IS INITIATED & TERMINTED IN 10 S.IF ANIMAL HAS NOT
ESCAPED ONTO THE PLATFORM BY THIS TIME.
 ESCAPE RESPONSE OCCURED.ANIMAL ALLOWED TO REMAIN ON
PLATFORM(10SEC).RETURN IT TO GRID FLOOR.
 10 TRIAL WITH TIME INTERVAL OF 20SEC ARE GIVEN
 IN NAÏVE CONTROL GRP OF RATS,THIS TRAINING RESULTED IN 80% OF
AQUIRING LEARNES HELPLESSNESS BEHAVIOUR. & DRUGS ARE GIVEN
BEFORE TRAINING & THE TEST PERIOD.
PURPOSE
1. Animals are exposed to inescapable
& Unavoidable electronic shocks in
One situation later fail to escape
Shock in a different situations when
Escape is possible.
2. This phenomenon was evaluated as
A potential animal model of
Depression.
Evaluation
1. Drug is considered to be effective if
Learned helplessness is reduced & the
no. of failures to escape is decreased.
2. Imipramine is found to be active only
After repeated application.
3. Benzodiazepines=effective
Chlorpromazine HCl= ineffective

19. MURICIDE BEHAVIOUR IN RATS
 MALE SPRAGUE-DAWLEY RATS (WT=300-350gm)ARE USED.
 ONE MOUSE IS PLACED INTO THE RAT CAGE
 ABOUT 10-30% OF RATS KILL THE MOUSE BY BITING THE ANIMAL
THROUGH THE CERVICAL CORD
 ONLY RATS CONSISTANTLY KILLING MICE WITHIN 5 MIN .
 THE MICE ARE REMOVED AFTER 15-45 SEC TO AVOID EATING BY RATS
 DRUGS ARE INJECTED (i.p.) TO THE RATS BEFORE TEST
 MICE ARE PRESENTED FOR 30,60,120 MIN AFTER DRUG
ADMINISTRATION.
 ED50 VALUE IS CALCULATED.
PURPOSE :
1.THIS METHOD CAN BE USED FOR
EVALUATION OF ANTIDEPRESSANTS
LIKE TRICYCLIC & MAO INHIBITORS.
EVALUATION :
1. Failure to kill mouse within 5
Min is considered as inhibition of
Muricide behaviour.
2. Performing dose response
Experiment ed50 is defined as
the
Dose which inhibits mouse
Killing in 50% of the rats.

20. IN VITROIN VITRO METHODMETHOD
INHIBITION OF [3H]- NOREPINEPHRINE UPTAKE IN RAT BRAININHIBITION OF [3H]- NOREPINEPHRINE UPTAKE IN RAT BRAIN
SYNAPTOSOMESSYNAPTOSOMES
 TISSUE PREPARATION :
 MALE WISTAR RATS ARE DECAPITATED
 THE BRAINS RAPIDLY REMOVED
 THE HYPOTHALAMIC REGION IS PREPARED, WEIGHED AND HOMOGENIZED
[IN 9 VOLUMES OF ICE-COLD 0.32 M SUCROSE SOLUTION USING A
POTTER- ELVEJHEM HOMOGENIZER]
 THE HOMOGENATE IS CENTRIFUGED AT 1000 G AT 0-4 oC
 THE SUPERNATENT IS DECANTED &USED FOR EXPERIMENT
16
PURPOSE :
1. THE NEURONAL REUPTAKE OF NE IS
MOST IMPORTANT PHYSIOLOGICAL
PROCESS FOR REMOVING &
INACTIVATING NE IN SYNAPTIC CLEFT.
2. THIS UPTAKE IS INHIBITED BY
ANTIDEPRESSANTS.
3. IN THE BRAIN THE HYPOTHALAMUS
SHOWS GREATEST UPTAKE OF
NA.THEREFORE,THIS REGION IS USED FOR
TESTING POTENTIAL ANTIDEPRESSANT
DRUGS.
EVALUATION :
1.THE PERCENTAGE INHIBITION AT EACH DRUG CONCENTRATION IS THE MEAN OF 3 DETERMINATIONS.
2.IC50 VALUES ARE DERIVED FROM LOG-PROBIT ANALYSIS.
3.IC50 VALUES FOR THE STANDARD DRUGS DESIPRAMINE & NORTRYPTYLINE ARE AROUND 20nM.

21. CONTI……CONTI…… IN VITROIN VITRO METHODMETHOD
INHIBITION OF [3H]- NOREPINEPHRINE UPTAKE IN RAT BRAININHIBITION OF [3H]- NOREPINEPHRINE UPTAKE IN RAT BRAIN
SYNAPTOSOMESSYNAPTOSOMES
ASSAY PROCEDURE
 200µl OF TISSUE SUSPENSION ARE INCUBATED WITH 800µ l 62.5 Nm NE IN KREBS-HENSELEIT BICARBONATE BUFFER
& 20 µl OF THE APPROPRIATE DRUG CONCENTRATION AT 37’C UNDER 95%O2 /5% CO2.
 FOR EACH ASSAY TUBES ARE INCUBATED WITH 20µl OF VEHICLE AT 0’C IN ICE BATH.
 AFTER THAT CENTRIFUGATION IS DONE 4000g FOR 10 MIN.
 SUPERNATENT LIQUID IS ASPIRATED & PELLETS ARE DISSOLVED IN SOLUBILIZER.
 SHAKING IS DONE & DECANTED INTO SCIENTILLATION VIALS & COUNTED IN 10 ML OF LIQUID SCINTILLATION
COCKTAIL.
 ACTIVE UPTAKE IS THE DIFFERENCE BETWEEN cmp at 37’C & 0 ‘C.

22. CONCLUSIONCONCLUSION
 Depression is an incapacitating disease which needs appropriate treatment.
 This presentation reviews the pharmacology of antidepressant drugs and the future perspectives of
treating mood disorders such as depression.
 The foremost theory for explaining the biological basis of depression has been the monoamine
hypothesis.
 Depression is due to a deficiency in one or other biogenic monoamines (serotonin, 5-HT;
noradrenalin, NA; dopamine, DA). Antidepressant drugs are therefore classified according to their
ability to improve monoaminergic transmission.
 Since this first theory, other explanations based on abnormal function of monoamine receptors or
associated with impaired signaling pathways have been suggested.
 Notable progress has been accomplished in the treatment of major depressive disorders with new
compounds recently discovered (selective serotonin reuptake inhibitors: SSRI; serotonin noradrenalin
reuptake inhibitors: SNRI).
 Behavioral, electrophysiological and micro dialysis studies have shown that serotonin (5-HT)
receptors, mainly 5-HT1A, 5-HT1B and 5-HT2C sub-types, exert a key role in modulating
antidepressant activity. 22

23. BIBLIOGRAPHYBIBLIOGRAPHY
1. Lead Medical Assessor Anders Wessling ,Lead Health Economist Joakim Ramsberg, “The review of
Antidepressants”
2. “Antidepressants The Old and The New” ,October, 1998 iii
3. Review on Pharmacology of Antidepressants ,ELLIOTTRICHELSON, MD, Mayo Clin Proc. 2001;76:511-
527.
4. Ayflegül Y›ld›z, M.D., “Mechanism of Actions of Antidepressants: Beyond the Receptors”, 2002;12:194-
200.
5. http://ajp.psychiatryonline.org/cgi/reprint/157/11/1901
6. http://www.webmd.com/depression/
7. http://pn.psychiatryonline.org/content/41/24/21.full
8. http://www.mayoclinic.com/health/maois/MH00072
9. http://www.springerlink.com/content/b9b8668ff59f89d7/fulltext.pdf
10. http://www.emsam.com/pi_emsam.pdf
11. H.GERHARD VOGEL, “DRUG DISCOVERY & EVALUATION” PHARMACOLOGICAL ASSAY,SECOND
EDITION2002.
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24. Thank
You…..--------------------------------------------------------------------------------
Thank
You…..--------------------------------------------------------------------------------