BYMALLAPPA. SHALAVADI,M-PHARM I,HSK COLLEGE OF PHARMACY,BAGALKOT.

INTRODUCTIONIt is a progressive neurodegenerative disorder .Their is a marked deficiency in the dopaminergic

innervation of the basal ganglia owing to degeneration of neurones in the substantia nigra.

Enhancement of dopaminergic transmission restores at least partially motor function.

The decrease in dopaminergic activity in the basal ganglia results in a relative excess of cholinergic influence.

Therefore, dopaminergic agonists, such as levodopa, a precursor of dopamine, and cholinergic (muscarinic) antagonists can be combined in the treatment of Parkinson’s disease.

SCREENING MODELS I. IN VIVO MODELSa)Tremorine and oxotremorine

antagonismb)MPTP model in monkeysc)Reserpine antagonismd)Circling behavior in nigrostriatal

lesioned ratse)Elevated body swing testf)Skilled paw reaching in ratsg)Stepping test in rats

II. IN VITRO AND EX VIVO MODELSa. Experiments using rat striatal slicesb. Dopamine stimulated Adenylyl

Cyclase activityc. Radioligand binding studies for D1

and D2 Dopamine receptorsd. Dopamine release from

synaptosomes

a. Tremorine and oxotremorine antagonism –

PURPOSE AND RATIONALE The muscarinic agonists tremorine and oxotremorine induce parkinsonism-like signs such as tremor, ataxia, spasticity, salivation, lacrimation and hypothermia. These signs are antagonized by anticholinergic

drugs. PROCEDURE Groups of 6-10 male NMRI mice weighing 18–22 g are used. They are dosed orally with the test compound or the

standard (5 mg/kg benzatropine mesilate) 1 h prior the administration of 0.5 mg/kg oxotremorine s.c.

Rectal temperature is measured before administration of the compound (basal value) and 1, 2 and 3 h after oxotremorine injection.

Tremor is scored after oxotremorine dosage in 10 s observation periods every 15 min for 1 h.

Tremor Score absent 0 slight 1 medium 2 severe 3

Salivation and lacrimation are scored 15 and 30 min after oxotremorine injection. absent 0 slight 1 medium 2 severe 3 EVALUATIONHypothermiaThe differences of body temperature after 1, 2 and 3

h versus basal values are summarized for each

animal in the control group and the test groups. The average values are compared statistically.

TremorThe scores for all animals in each group at the

3 observation periods are summarized. The numbers in the treated groups are

expressed as percentage of the number of the control group.

Salivation and lacrimationThe scores for both symptoms for all animals in

each group are summarized at the 2 observation

periods. The numbers in the treated groups are

expressed as percentage of the number of the control group.

MODIFICATIONS OF THE METHOD Johnson et al. (1986) Developed a procedure for quantifying

whole-body tremors in mice. Displacement of a free floating platform

by animal movement created a change in resistance across a strain gauge.

Administration of oxotremorine, 2.5 mg/kg, i.p, produced numerous high-frequency, high-intensity peaks within 5 min.

Clement and Dyck (1989) Constructed and tested a tremor monitor that

quantitates soman- and oxotremorine-induced tremors.

The device consisted of a force transducer, from which a plastic beaker was suspended containing a mouse.

The signal from the force transducer was fed into a tremor monitor and quantitated using the Applecounter from Columbus Instruments.

Coward et al. (1977) Recommended N-carbamoyl-2-(2,6-

dichlorophenyl)acetamidine hydrochloride (LON-1954), a tremorigenic agent, as alternative to oxotremorine for the detection of anti-Parkinson drugs

b. MPTP model in monkeys

PURPOSE AND RATIONALEN-MPTP (N-methyl-4-phenyl-1,2,3,6-

tetrahydropyridine) has been shown to cause symptoms of Parkinson’s disease in exposed individuals.

When administered to primates this compound causes a partial destruction of basal ganglia and a syndrome that resembles Parkinson’s disease.

How MPTP induce parkinsonism?

PROCEDUREBurns et al. (1983) treated 8 adult rhesus monkeys weighing 5–8 kg over a period of 5–8 days with

cumulative intravenous doses of N-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (N-MPTP) up to 10–18 mg/kg.

These animals showed a parkinsonism like disorder

(akinesia, rigidity, postural tremor, flexed posture, eyelid closure, drooling)

Which was reversed by the administration of L-dopa.

The pathological and biochemical changes produced by N-MPTP are similar to the well established changes in patients with Parkinsonism.

EVALUATION The severity of parkinsonian symptoms is rated

by trained observers movement (0: normal; 1: reduced; 2: sleepy) checking movements (0: present; 1: reduced; 2:

absent)attention and blinking (0: normal; 1: abnormal) posture (0: normal; 1: abnormal trunk; 2:

abnormal trunk and tail; 3: abnormal trunk, tail, and limbs; 4: flexed posture)

balance and coordination (0: normal; 1: impaired; 2: unstable; 4: falls)

reactions (0: normal; 1: reduced; 2: slow; 3: absent)

vocalizations (0: normal; 1: reduced; 2: absent).

MODIFICATIONS OF THE METHODKebabian et al. (1992) tested a selective D1

receptor agonist with antiparkinsonian activity in MTPT treated marmosets.

Domino and Sheng (1993) studied the relative potency of some dopamine agonists with varying selectivities for D1 and D2 receptors in MPTP-induced hemiparkinsonian monkeys.

Asin et al. (1997) tested a selective D1 receptor agonist in rats previously given unilateral 6 hydroxydopamine injections and in macaques previously given unilateral, intracarotid infusions of MPTP.

c. Reserpine antagonism

PURPOSE AND RATIONALEReserpine induces depletion of central

catecholamine stores. The sedative effect can be observed in mice

shortly after injection, followed by signs of eyelid ptosis, hypokinesia, rigidity, catatonia, and immobility.

These phenomena can be antagonized by dopamine aonists.

MODIFICATIONS OF THE METHODRats treated with reserpine develop

spontaneous orofacial dyskinesia that has features similar to tardive dyskinesia in humans (Nisewander et al. 1994).

The incidence of tongue protrusions was recorded to quantify the occurrence of oral dyskinesia.

d. Circling behavior in nigrostriatal lesioned rats

PURPOSE AND RATIONALEUnilateral lesion of the dopaminergic

nigrostriatal pathway in the rat by the neurotoxin 6-hydroxydopamine (6OHDA) induces hypersensitivity of the postsynaptic dopaminergic receptors in the striatum of the lesioned side.

The rats rotate in a direction towards the lesioned side (ipsilateral) when an indirect acting compound such

as amphetamine is administered but to the opposite direction (contralateral) when a direct acting dopamine agonist, e.g., apomorphine, or the dopamine precursor L-dopa is given.

Therefore, this test can be used for the study of central dopamine function and the evaluation of dopamine antagonists and agonists, particularly the activity of novel antiparkinsonian drugs.

MODIFICATIONS OF THE METHODEtemadzadeh et al. (1989) described a

computerized rotometer apparatus for recording circling behavior

Hudson et al. (1993) described a 16-channel automated rotometer system for reliable measurement of turning behavior in 6-hydroxydopamine lesioned and transplanted rats

Garrett and Holtzman (1996) compared the effects of apomorphine, cocaine and caffeine on locomotor activity and rotational behavior in rats with unilateral 6-OHDA-induced lesions of the nigrostriatal tract.

e. Elevated body swing testPURPOSE AND RATIONALEBorlongan and Sanberg (1995) proposed the

elevated body swing test as a measure of asymmetrical motor behavior of hemiparkinsonian animals in a drug-free state.

f. Skilled paw reaching in ratsPURPOSE AND RATIONALEBarnéoud et al. (1996) used the skilled paw

reaching test as a model of Parkinson’s disease in the rat.

Unilateral injection of 6-OHDA into the medial forebrain bundle results in an impairment of paw reaching on both sides which can be ameliorated by drug treatment or transplantation of a nigral cell suspension.

g. Stepping test in ratsPURPOSE AND RATIONALESchallert et al. (1992), Olsson et al. (1995)

introduced the stepping test as a clinically relevant unilateral model of Parkinsonian akinesia.

The 6-OHDA lesion induced marked and long-lasting impairments in the initiation of stepping movements with the contralateral paw which can be ameliorated by the systemic application of drugs.

II. IN VITRO AND EX VIVO MODELSa.Dopamine- stimulated Adenylyl cyclase activity.

REFERANCESDrug Discovery and Evalution by H. Gerhard Vogel. II nd Ed.

Drugs Screening Methods By S. K. Gupta.

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