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““A REVIEW ON PRECLINICAL A REVIEW ON PRECLINICAL EVALUATION OF ANXIOLYTICS”EVALUATION OF ANXIOLYTICS”

PRESENTING BY,PRESENTING BY,Ms. SONALI B. DIWATEMs. SONALI B. DIWATE

GUIDED BY,GUIDED BY,PROF. R. CHANSHETTI PROF. R. CHANSHETTI

MODERN C.O.P.,MOSHI, PUNE-412105 MODERN C.O.P.,MOSHI, PUNE-412105

To Review and Study the Preclinical Evaluation of Anxiolytics. To Review and Study the Preclinical Evaluation of Anxiolytics.

OBJECTIVEOBJECTIVE

1.TO COLLECT THE INFORMATION ABOUT ANXIETY & ANXIOLYTICS. (LITERATURE 1.TO COLLECT THE INFORMATION ABOUT ANXIETY & ANXIOLYTICS. (LITERATURE SURVEY).SURVEY).

2.TO SURVEY THE STATUS AND STANDARD ANXIOLYTIC DRUGS AVAILABLE IN 2.TO SURVEY THE STATUS AND STANDARD ANXIOLYTIC DRUGS AVAILABLE IN MARKET.MARKET.

3. TO STUDY THE COMMON ADR & USES OF ANXIOLYTICS.3. TO STUDY THE COMMON ADR & USES OF ANXIOLYTICS.4.TO STUDY THE PHRMACOKINETICS,PHARMACODYNAMICS OF ANXIOLYTICS.4.TO STUDY THE PHRMACOKINETICS,PHARMACODYNAMICS OF ANXIOLYTICS.5.TO EXPLAIN INVIVO & INVTRO MODELS FOR PRECLINICAL EVALUATION OF 5.TO EXPLAIN INVIVO & INVTRO MODELS FOR PRECLINICAL EVALUATION OF

ANXIOLYTICS.ANXIOLYTICS.6.TO ANALYZE ALL THE PARAMETERS RELATED TO ANXIOLYTICS6.TO ANALYZE ALL THE PARAMETERS RELATED TO ANXIOLYTICS7.SUMMERIZATION OF THE INFORMATION.7.SUMMERIZATION OF THE INFORMATION.

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CONTENTS…..CONTENTS…..

1. AIM AND OBJECTIVE1. AIM AND OBJECTIVE2.INTRODUCTION TO ANXIETY & ANXIOLYTICS 2.INTRODUCTION TO ANXIETY & ANXIOLYTICS 3.STANDARD ANXIOLYTICS AVAILABLE IN MARKET3.STANDARD ANXIOLYTICS AVAILABLE IN MARKET4. COMMON ADR & USES OF ANXIOLYTICS4. COMMON ADR & USES OF ANXIOLYTICS5. PHARMACOKINETICS & PHARMACODYNAMICS5. PHARMACOKINETICS & PHARMACODYNAMICS6. PRECLINICAL SCREENING METHODS6. PRECLINICAL SCREENING METHODS7. SUMMERY7. SUMMERY8. CONCLUSION8. CONCLUSION9. BIBLIOGRAPHY9. BIBLIOGRAPHY

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INTRODUCTION TO DEPRESSIONINTRODUCTION TO DEPRESSION

1. Anxiety is an emotional state commonly caused by the perception of real or danger that threatens the security of an individual.

2.It allows a person to prepare for or react to environmental changes .3.Anxiety can produce uncomfortable nature.

The Symptoms of Anxiety1. Sweating2. Shaking3. Palpitation4 .Blushing5. GI Discomfort6. Lack Of Sleep7. Lack Of Concentration8. Irritation

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TYPES OF ANXIETY Panic Disorder-unreasonable fear spreading for 10-20minGeneralized Anxiety Disorder-experienced when person thinking about no. of events Social Anxiety Disorder-means fear about interaction with social programs.Obsessive Compulsive Disorder-repeated experience of specific image or thoughts.

HISTORY OF ANTIDEPRESSANTSHISTORY OF ANTIDEPRESSANTSThe drugs or agents which are used in treatment of anxiety is called as

Anxiolytics.Anxiolytics acts by balancing brain neurotransmitters level to ease anxiety.

CLASSIFICATION OF ANXIOLYTICS

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INTRODUCTION TO INTRODUCTION TO ANXIOLYTICS

1. BENZODIAZEPINES-ALPRAZOLAM,CHLORDIAZEPOXIDE,DIAZEPAM,LORAZEPAM

2. AZAPIRONES-BUSPIRONE,TANDOSPIRONE,GEPIRONE

3. BETA BLOCKER-PROPRANOLOL

4. CARBAMATES-MEPROBAMATES

STANDARD DRUGS AVAILABLE IN MARKETSTANDARD DRUGS AVAILABLE IN MARKET

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SR.NO DRUG MARKETED PREPARATION DOSE

1 ALPRAZOLAM Xanax 0.25mg

2 DIAZEPAM Valium 5mg

3 BUSPIRONE Buspar 5mg

4 PROPRANOLOL  Deralin 2.5mg

5 MEPROBAMATE Miltown 5mg

COMMON ADVERSE DRUG REACTIONCOMMON ADVERSE DRUG REACTION

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Very common (>10% incidence) adverse effects include:

DizzinessHeadache

Common (1-10% incidence) adverse effects include:

NervousnessInsomniaDisturbance in attention

Uncommon (0.1-1%) adverse effects include:

HypertensionRedness and itching of the eyesAltered tasteConjunctivitisFlatulenceAnorexiaIncreased appetiteSalivation

DRUG INTERACTIONS

• Alprazolam is primarily metabolised via CYP3A4.Combining CYP3A4 inhibitors such as cimetidine, erythromycin, fluoxetine, fluvoxamine,itraconazole, ketoconazole, nefazodone, propoxyphene, and ritonavir delay the hepatic clearance of alprazolam, which may result in excessive accumulation of alprazolam. This may result in exacerbation of its adverse effect profile.

• Imipramine and desipramine have been reported to be increased an average of 31% and 20%, respectively, by the concomitant administration of alprazolam tablets in doses up to 4 mg/day .

• Combined oral contraceptive pills reduce the clearance of alprazolam, which may lead to increased plasma levels of alprazolam and accumulation.

• Alcohol is one of the most important and common interactions. Alcohol and benzodiazepines such as alprazolam taken in combination have a synergistic effect on one another, which can cause severe sedation, behavioural changes, and intoxication. The more alcohol and alprazolam taken the worse the interaction. Combination of alprazolam with the herb kava can result in the development of a semi-comatose state.Hypericum conversely can lower the plasma levels of alprazolam and reduce its therapeutic effect.

CONTRAINDICATIONS

• Myasthenia gravis.• Severe respiratory impairment e.g sleep

apnoea, COAD.• Pregnancy • Lactation

The mechanism of action of BENZODIAZEPINES: Benzodiazepines acts selectively on GABAA receptor which mediate fast inhibitory

synaptic transmission throughout the CNS. They enhance the response to GABA by facilitating opening of GABA- activated chlorine channels & increases affinity of GABA for the receptor.

The mechanism of action of BUSPIRONE: It is a partial agonist at 5HT1A receptor. These are the auto receptor that reduces

release of 5HT & other mediators. They also inhibits the activity of noradrenergic locus cerulus neurons & thus interferes with arousal reaction.

The mechanism of action of BETA BLOCKER: Many symptoms of anxiety such as palpitation, rise in BP, tremor due to over

activation of sympathetic system. Beta blocker help anxious patient by cutting vicious cycle & provides symptomatic relief. They do not affect psychological symptoms.

The mechanism of action of CARBAMATES: Acts on several levels of CNS,including limbic system, thalamus & spinal cord. It has

mild to moderate tranquilizing, anticonvulsant & skeletal muscle relaxing activity. 10

PHARMACOKINETICS & PHARMACODYNAMICS OF ANTIDEPRESSANTSPHARMACOKINETICS & PHARMACODYNAMICS OF ANTIDEPRESSANTS

1) GIVEN BY ORAL ROUTE2) METABOLISM BY LIVER3) EXCREATION THROUGH URINE

SCREENING METHODSSCREENING METHODS

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IN VIVO METHODS IN VITRO METHODS

CLASSIFICATION OF SCREENING METHODSCLASSIFICATION OF SCREENING METHODS

IN VIVO IN VIVO METHODSMETHODS

1.Foot shock induced aggression2.Isolation induced aggression3.Maternal induced aggression4.Light & Dark model5.Open field test6.Elevated plus maze model7.Anticipatory anxiety in mice8.Marble burying behavior model9.Staircase test10.Marmoset test11.Cork gnawing test in rats

IN VITRO IN VITRO METHODSMETHODS

1.GABA receptor binding2.GABA receptor binding3.Benzodiazepine receptor binding4.Serotonine receptor binding

[5HT1A, 5HT2A]

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FOOT SHOCK INDUCED AGGRESSION

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PURPOSE: Induction of aggression in pair of mice by giving shock to feet of mice. Anxiolyticsreduces fighting behavior in mice. EVALUATION:Record fighting behavior before & after treatment.

Procedure:1.Place pair of mice in a box consisting of grid floor, which is made up of steel rods kept at distance 5-6 mm & constant current(0.8mA) is allowed to pass for 5sec & 5sec intermission. 2.Record fighting behavior in pair of mice for 5min before & after treatment.

ISOLATION INDUCED AGGRESSION

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PURPOSE:Keeping one of the micealone for prolonged period oftime, so that aggressiondevelops against same sexanimal. Anxiolytic agentsuppress isolation inducedaggression . EVALUATION:Record no. of attacksbefore & after treatment.

Procedure:1.Keep mouse or rat alone for long period of time result into generation of aggression. Place intruder into cage of isolated mouse.2.Isolated mouse attacks on intruder within 10 sec & record no of attacks for 3 min.

MATERNAL INDUCED AGGRESSION

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PURPOSE: 1.Induction of an intruder incage of parturient female rat,induced high level aggressionagainst such intruder.2. Maternal aggression ischaracterized by high intensityattacks on head or neck ofintruder. Anxiolytics reducesthe aggression. EVALUATION:Record fighting behavior before & after treatment.

Procedure:1.One male intruder is placed in female’s home cage for 5 min.2.The ongoing behavior is videotaped & analyzed later.3.Each intruder is used only once & sacrificed immediately afterwards with an i.p. overdose of pentobarbital, followed by shaving & describing the wounds on wound charts.

IN VITRO IN VITRO METHODMETHODGABAGABAAA RECEPTOR BINDING RECEPTOR BINDING

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

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PRINCIPLE :1. THE BASIC BEHIND TO FOLLOW

INVITRO SCREENING METHOD IS THAT TO FIND OUT BINDING CHARACTERISTICS OF TEST DRUG WITH SPECIFIC RECEPTOR.

2. FOR THIS WE ARE USING RADIOLEGANG THAT MAY BE AGONIST OR ANTAGONIST.

3. THEN THE REACTIVITY IS MEASURED USING LIQUID SCINTILLATION METHOD

4. CALCULATE TOTAL BINDING,NON-SPECIFIC BINDING,SPECIFIC BINDING.

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.

ASSAY PROCEDURE

IN THIS WE DETERMINE WE DETERMINE TOTAL BINDING & NON-SPECIFIC BINDING.

1.TOTAL BINDING:

INTHIS PELLET SUSPENSION IS UTILIZED

INCUBATE THE TISSUE WITH RADIOLABELLED SUBSTANCE HAVING AFFINITY FOR SPECIFIC & NONSPECIFIC BINDING SITE.

MEMBRANE SOLUTION IS SUBJECTED TO CENTRIFUGATION FOR FAST REACTION OF BINDING

CARRY OUT FILTERATION & SEPARATED FRAGMENTS DISSOLVED IN SCINTILLATION FLUID

MEASURE RADIOACTIVITY USING LIQUID SCINTILLATION COUNTER

..

CONT…… IN VITRO CONT…… IN VITRO METHODMETHOD

GABAA RECEPTOR BINDING GABAA RECEPTOR BINDING

2. NON SPECIFIC BINDING: IN THIS SUCH DRUG IS USED WHICH WILL BIND TO SPECIFIC SITE AND WHICH IS NON

RADIOLABELED.SO THAT NON RADIOLABELED SUBSTANCE WILL REPLACE ONLY RADIOLABELED BINDED DRUG FROM SPECIFIC BINDING SITE AND ONLY RADIOLABELED NON SPECIFIC BINDING REMAINS AS IT IS.

INCUBATE THE TISSUE WITH RADIOLABELLED SUBSTANCE+DRUG(STANDARD/TEST).

SELECT DRUG WITH BINDS ONLY TO SPECIFIC SITE.

NON RADIOLABELED DRUG WILL REPLACE RADIO SUBJECT FROM SPECIFIC BINDING SITE.

MEASURE RADIOACTIVITY USING LIQUID SCINTILLATION COUNTER.

THIS WILL GIVES RADIOACTIVITY OF NON SPECIFIC SITE. 2. SPECIFIC BINDING = TOTAL BINDING- NON-SPECIFIC BINDING

A

CONT…… IN VITRO CONT…… IN VITRO METHODMETHOD

GABAA RECEPTOR BINDING GABAA RECEPTOR BINDING

CONCLUSIONCONCLUSION1. Anxiety is an emotional state commonly caused by the perception of real or danger that

threatens the security of an individual.2.It allows a person to prepare for or react to environmental changes .3.Anxiety can produce uncomfortable nature.

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BIBLIOGRAPHYBIBLIOGRAPHY

1. H.GERHARD VOGEL, “DRUG DISCOVERY & EVALUATION” PHARMACOLOGICAL

ASSAY,SECOND EDITION2002.

2. http://ajp.psychiatryonline.org/cgi/reprint/157/11/1901

3. http://www.webmd.com/depression/

4. http://pn.psychiatryonline.org/content/41/24/21.full

5. http://www.mayoclinic.com/health/maois/MH00072

6. http://www.springerlink.com/content/b9b8668ff59f89d7/fulltext.pdf

7. http://www.emsam.com/pi_emsam.pdf

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Thank You…..

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Thank You…..

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