Nootropic activity of tuber extract of Pueraria tuberosa (roxb)

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1 Indian Journal of Experimental Biology Vol. 46, August 2008, pp Nootropic activity of tuber extract of Pueraria tuberosa (roxb) N Venkata Rao 1, Basavaraj Pujar 1, S K Nimbal 1, S M Shantakumar 2 & S Satyanarayana 3 1 Depertment of Pharmacology and 2 Pharmaceutical Chemistry, V L College of Pharmacy, Raichur , India. 3 College of Pharmaceutical Sciences, Andhra University, Visakhapatnam , India. Received 8 August 2007; revised 26 May 2008 Nootropic effect of alcoholic (ALE; 50, 75, 100 mg/kg) and aqueous (AQE; 100, 200, 400 mg/kg) extracts of P. tuberosa was evaluated by using Elevated Plus Maze (EPM), scopolamine-induced amnesia (SIA), diazepam-induced amnesia (DIA), clonidine-induced (NA-mediated) hypothermia (CIH), lithium-induced (5-HT mediated) head twitches (LIH) and haloperidol-induced (DA- mediated) catalepsy (HIC) models. Piracetam was used as the standard drug. A significant increase in inflexion ratio (IR) was recorded in EPM, SIA and DIA models. A significant reversal effect was observed on rectal temperature in CIH model, reduction of head twitches in LIH models. However no significant reduction in catalepsy scores in HIC models were observed with test extracts and standard piracetam. The results indicate that nootropic activity observed with ALE and AQE of tuber extracts of P. tuberosa could be through improved learning and memory either by augmenting the noradrenaline (NA) transmission or by interfering with 5-hydroxytryptamine (5-HT) release. Further, the extracts neither facilitated nor blocked release of the dopamine (DA). Thus ALE and AQE elicited significant nootropic effect in mice and rats by interacting with cholinergic, GABAnergic, adrenergic and serotonergic systems. Phytoconstituents like flavonoids have been reported for their nootropic effect and these are present in both ALE and AQE extracts of tubers of P. tuberosa (Roxb) and these active principles may be responsible for nootropic activity. Keywords: Antiamnesic, Anticataleptic, EPM, Pueraria tuberosa, Tuber extracts Dementia is described as a syndrome due to chronic or progressive disease of the brain, leading to disturbance of multiple functions of higher cortical centers including memory, orientation, comprehension, calculation, learning capacity, language and judgment without altering consciousness. Further deterioration of emotional control, social behaviour or motivation may accompany or proceed to cognitive impairment. Alzheimer s disease (AD) is an incurable, progressive brain disorder that causes dementia and abnormal phosphorylation of the intracellular tauproteins, causing abnormalities of microtubule assembly and collapse of the cytoskeleton affected, particularly pyramidal cells of the cortex and sub cortex 1. Hypo function of various neuronal systems particularly the cholinergic neuronal systems with decrease in several cholinergic parameters, e.g., choline acetyltransferase and cholinesterase activity, choline acetyltransferase mrna and acetyl choline receptors were observed 2. Correspondent author Telephone: ; Mobile: Fax: vrngadda@rediffmail.com A protein called casein kinase-1 could slow the formation of β amyloid protein, the main constituents of the amyloid plaques found in the brains of Alzheimer s disease thought to cause the debilitating effects of the disease. β amyloid protein is toxic to mitochondria found in cells and ultimately causes nerve cell death 3. The dementia in the elderly i.e., about 5 million people in the United States are estimated to be afflicted by this disorder and the percentage approximately doubles with every 5 years of age with about 1% of 60 years olds and about 30% of 85 years olds having the disease 4. In the present study a plant Pueraria tuberosa (Roxb) popularly used in folklore medicine was selected as it is reported for its nerve tonic, galactogogue, anti-inflammation 5 brain tonic 6 and as a mind power syrup 7 in Ayurvedic formulations. Earlier the plant has been studied for its antistaphylococcal 5, antitubercular 5, antifungal 5, antiischaemic 5, antihepatotoxic 5, antiimplantation 5, antifertility 8, anti-inflammatory, woundhealing 9, estrogenic 10, contraceptive 11, hypoglycemic 12 and for adaptogenic 13 activities.

2 592 INDIAN J EXP BIOL, AUGUST 2008 Materials and Methods Drugs Piracetam (B & B Pharmaceuticals Ltd.), scopolamine (Cadila Healthcare Ltd.), diazepam (Ranbaxy Laboratories Ltd.), clonidine (Unichem Ltd.), lithium carbonate (Torrent Laboratories Ltd.), amitriptyline (Torrent Laboratories Ltd.), haloperidol (RPG Life sciences Ltd.) and Mentat (Himalaya Drug Company) were used. Animals Albino mice and rats of either sex weighing between g and g respectively procured from Shri Venkateswara Enterprises, Bangalore were used. After procuring, all the animals were acclimatized for 7 days and housed in groups of six under standard husbandry conditions 14,15 (26 ± 2 C, 45-55% RH) and 12:12 hr light/dark cycle. All the animals were fed with synthetic standard diet (Amrut Laboratories Pranava Agro Industries Ltd. Sangli, Bangalore.) and water provided ad libitum under strict hygienic conditions. After obtaining permission from Institutional Animal Ethical Committee (IAEC) of V L College of Pharmacy, Raichur (Karnataka), animal studies were performed as per rules and regulations and in accordance to the guidelines of CPCSEA with registration number 557/02/c/CPCSEA, All experiments were carried out during the light period between hr. Preparation of different extracts with tubers of P. tuberosa Alcoholic extract 16 The tubers powder was extracted with 95% alcohol in a soxhlet apparatus at C for 18 hr, the alcoholic extract was subjected to evaporation in a beaker on a water bath maintained at 50 C till a thick paste of extract remained in the beaker and was kept in refrigerator below 4 C till the experimental study. Aqueous extract 16 The marc remained in the previous extraction was air dried thoroughly to remove the solvent. About 100 g of the marc was taken in a round bottom flask (2000 ml capacity) and macerated with 500 ml distilled water for 24 hr with occasional shaking for every 1 hr and 10 ml of chloroform was added as a preservative. Then the marc was removed filtering the extract, and was concentrated on a water bath at 50 C and kept in refrigerator till the experimental study. Pharmacological activities Determination of acute toxicity (LD 50 ) The acute toxicity of ALE and AQE of tubers of P. tuberosa was determined in albino mice of either sex (16-25 g) maintained under standard husbandry conditions. The animals were fasted 3 hr prior to the experiment. Up and down method (OECD guidelines No. 425) of CPCSEA was adopted for toxicity studies 17. Animals were administered with single dose of extract and observed for mortality during 48 hr study period (short term toxicity). Based on short-term toxicity profile of extract the dose for the next animal was determined as per the OECD guideline No The LD 50 of the two extracts were calculated using AOT 425 software provided by Environmental Protection Agency, USA 18,19. Laboratory models for testing learning and memory Elevated Plus-Maze (Exteroceptive behaviour model) 20 : Eight groups of mice each having 6 animals, weighing between g were used. I was maintained as normal control which was given distilled water (10 ml/kg, po) only once daily for 7 days, group II with piracetam (200 mg/kg, po) which served as standard, and groups III, IV, V and VI, VII, VIII were treated orally with different doses of ALE (50, 75 and 100 mg/kg) and AQE (100, 200 and 400 mg/kg) of tuber extracts of P. tuberosa, respectively once daily for 7 days. On the 7 th day, 90 min after treatment of last dose each mouse was placed at the end of an open arm of EPM facing away from the central platform. Transfer latency (TL) was recorded i.e. the time taken by mouse to move into one of the enclosed arms with all its four legs. If the animal did not enter into one of the enclosed arms within 90 sec, it was gently pushed into any of the two enclosed arms and the TL was assigned as 90 s. The mouse was allowed to explore the maze for next 10 sec and then to be returned to its home cage. Retention of this learned-task was examined 24 hrs after the 7 th day trial. The Inflexion Ratio was calculated by the formula as follows 21 : IR = (L 0 L 1 )/ L 0 where L 0 is the initial TL (s) on first day and L 1 is the TL (s) on the 2 nd day. Scopolamine-induced amnesia (Interoceptive behaviour model) 20 : Nine groups of mice each comprising 6 animals, weighing between g were used. I was maintained as normal control, which was given distilled water (10 ml/kg, po) only once daily for 7 days. In this 90 min after the last dose treatment (distilled water) TL was recorded on EPM

3 RAO et al.: NOOTROPIC ACTIVITY OF TUBER EXTRACT OF PUERARIA TUBEROSA 593 and retention (memory) of learned task was examined 24 hr later. II was injected with scopolamine (0.4 mg/kg, ip) alone on 1 st day and after 45 min TL was recorded on EPM and retention (memory) of learned task was examined 24 hr later. III was administered with piracetam (200 mg/kg, po) and groups IV, V, VI and groups VII, VIII, IX were treated with different doses of ALE (50, 75 and 100 mg/kg, po) and AQE (100, 200 and 400 mg/kg, po) of tuber extracts of P. tuberosa respectively once daily for 7 days. On 7 th day 90 min after administration of piracetam or ALE or AQE, amnesic agent scopolamine was administered. After 45 min TL was recorded on EPM and retention (memory) of learned task was examined 24 hr later. The IR was calculated as described in earlier experiment. Diazepam-induced amnesia (Interoceptive behaviour model) 20 : Nine groups of mice each comprising 6 animals and weighing between g were used. I was maintained as normal control, which was given with distilled water (10 ml/kg, po) once daily for 7 days and on 7 th day 90 min after treatment (distilled water) TL was recorded on EPM and retention (memory) of learned task was examined 24 hr later. II was administered with diazepam (5 mg/kg, ip) alone on 1 st day only and after 45 min TL was recorded on EPM and retention (memory) of learned task was examined 24 hr later. III was treated with piracetam (200 mg/kg, po) groups IV, V, VI and groups VII, VIII, IX were treated with different doses of ALE (50, 75 and 100 mg/kg, po) and AQE (100, 200 and 400 mg/kg, po) of tuber extracts of P. tuberosa respectively once daily for 7 days. On 7 th day 90 min after administration of piracetam, ALE, AQE amnesic agent diazepam was administered. Then 45 min later TL was recorded on EPM and retention (memory) of learned task was examined 24 h later. The IR was calculated as described in earlier study. Behavioural study Clonidine-induced (NA-mediated) hypothermia 22,23 Ten groups of rats each consisting of 6 animals and weighing between g were used. I was treated with distilled water (10 ml/kg, po) only group II with clonidine (1 mg/kg, ip), group III with piracetam (200 mg/kg, po) groups IV, V, VI and groups VII, VIII, IX were treated with different doses of ALE (50, 75 and 100 mg/kg, po) and AQE (100, 200 and 400 mg/kg, po) of tuber extracts P. tuberosa respectively, 30 min later all the groups were administered with clonidine. Rectal temperature was recorded every 30 min. Lithium-induced (5-HT mediated) head twitches 22,23 Eight groups of rats each comprising 6 animals and weighing between g were selected. I was given distilled water (10 ml/kg, po) only, group II treated with lithium carbonate (190 mg/kg, ip) alone, group III with piracetam (200 mg/kg, po), group IV was treated with amitriptyline (20 mg/kg, po), groups V,VI VII and groups VII,VIII, IX were treated with different doses of ALE (50,75 and 100 mg/kg, po) and AQE (100, 200 and 400 mg/kg, po) of tuber extracts of P.tuberosa respectively, 30 min later lithium carbonate was administered to all the groups and the number of head twitches were counted for 60 min for each groups of rats. Haloperidol-induced (DA- mediated) catalepsy Ten groups of rats each comprising of 06 animals and weighing between g were selected. I received with distilled water (10 ml/kg, po) only, group II haloperidol (1 mg/kg, ip) and animals in groups III-X were treated with piracetam (200 mg/kg, po), mentat (100 mg/kg, po), ALE (50, 75 and 100 mg/kg, po) and AQE (100, 200 and 400 mg/kg, po) of tubers of P. tuberosa respectively. By means of Bar test 25 the catalepsy was observed and scored at 0, 30, 60, 90, 120, 150 and 180 min time interval. The phenomenon was measured as the duration when the rat maintained an imposed position with both front limbs extended and resting on wooden bar (8 8 8 cm). For better measurement of catalepsy (avoiding false results) the animals were tested twice at each time interval. In between measurement of catalepsy scores the animals were returned to their home cages, only the greater duration of time was recorded using the following scoring system: Time (sec) Grade > Statistical analysis The data were subjected to statistical analysis by One- way Analysis of Variance

4 594 INDIAN J EXP BIOL, AUGUST 2008 (ANOVA) followed by Dunnet s t test and P <0.05, 0.01 and were considered as significant. Results Pharmacological investigations Acute toxicity studies Acute toxicity of ALE and AQE extracts were determined in mice, as per OECD guidelines No LD 50 of ALE was found to be 1500 mg/kg while AQE even in greater than 2000 mg/kg dose did not produce any mortality. Laboratory models for testing learning and memory Effect of extracts on IR in mice with EPM model Piracetam (200 qmg/kg), and tuber extracts at different dose levels of ALE (50, 75 and 100 mg/kg) and AQE (100, 200 and 400 mg/kg), treated groups had shown increased IR. Statistically significant reduction in TL was observed with piracetam and both ALE and AQE except with AQE (100 mg/kg) dose (Table 1). Effect of extracts on IR in SIA model Scopolamine treated group induced impairment of memory, as shown by decrease in IR when compared to normal control group which indicates the induction of amnesia. Piracetam and all the doses of ALE (50, 75 and 100 mg/kg) and AQE (200 and 400 mg/kg) treated groups had shown a significant increase in the IR, reduction in TL and reversed the SIA. A similar effect was not observed with AQE 100 mg/kg treated group, which showed statistically insignificant effect (Table 2). Effect of extracts on IR in DIA model Diazepam has induced dose dependent amnesia in this amnesic model, a decrease in IR was observed as compared to normal control group. Piracetam and all the doses of Table 1 Nootropic effect of tuber extracts of P. tuberosa in mice with EPM model [Values are mean ± SE from 6 animals in each group] No. Treatment Dose (per kg) Inflexion ratio I Control 10 (ml, po) ±0.084 (vehicle) II Piracetam 200 (mg, po) ±0.286** III ALE 50 (mg, po) ±0.112* IV ALE 75 (mg, po) ±0.104** V ALE 100 (mg, po) 3.189±0.292** VI AQE 100 (mg, po) ±0.054 ns VII AQE 200 (mg, po) 1.591±0.085** VIII AQE 400 (mg, po) 2.507±4** Significance at P <0.05*, <0.01** and ns-not significant vs control (vehicle) ALE and AQE treated groups had shown an increase in the IR and a significant reduction in TL observed on EPM and diazepam induced amnesia was reversed (Table 3). Behavioural study Effect of extracts on CIH model Clonidine a α 2 - agonist, 90 min after administration produced a fall in rectal temperature of rats from F to 91.9 ±0.72 F when compared to normal control group. Piracetam reduced the body temperature from 97.7 ± F to F. The different doses of ALE (50, 75 and 100 mg/kg) and AQE (100, 200 and 400 mg/kg) significantly reversed the clonidineinduced hypothermia (Table 4). Table 2 Nootropic effect of tuber extracts of P. tuberosa on scopolamine- induced amnesia in mice [Values are mean ± SE from 6 animals in each group] No. Treatment Dose (per kg) Inflexion ratio I Normal control 10 (ml, po) ±0.084 (vehicle) II Scopolamine 0.4 (mg, ip) 0.189±0.096 control III Piracetam 200 (mg, po) ** IV ALE 50 (mg, po) * V ALE 75 (mg, po) 2.218±0.142** VI ALE 100(mg, po) 1.864±0.142** VII AQE 100 (mg, po) 0.437±0.073 ns VIII AQE 200 (mg, po) 1.229±0.133** IX AQE 400 (mg, po) ** Significance at P <0.05*, <0.01** and ns-not significant vs Scopolamine control Table 3 Nootropic effect of tuber extracts of P. tuberosa on diazepam- induced amnesia in mice [Values are mean ±SE from 6 animals in each group] No. Treatment Dose (per kg) Inflexion ratio I Normal control 10 (ml, po) ± (vehicle) II Diazepam 5 (mg, ip) ±0.070 Control III Piracetam 200 (mg, po) 1.574±0.107** IV ALE 50 (mg, po) 0.451±0.058** V ALE 75 (mg, po) 1.182±0.115** VI ALE 100 (mg, po) 1.224±0.137** VII AQE 100 (mg, po) 0.355±0.032** VIII AQE 200 (mg, po) 0.896±0.114** IX AQE 400 (mg, po) 1.127±0.103** Significance at P <0.05*,<0.01** and ns-not significant vs diazepam control

5 RAO et al.: NOOTROPIC ACTIVITY OF TUBER EXTRACT OF PUERARIA TUBEROSA 595 Effect of extracts on LIH model Lithium treated group had shown 19.0±2.0 head twitches in 60 min. Prior treatment with piracetam decreased the number of head twitches to 3.16±0.83, ALE with 50, 75 and 100 mg/kg and AQE with 200 and 400 mg/kg doses had showed a statistically significant reduction in number of head twitches i.e., 14.16±0.98, 12.0±0.73, 3.5±0.45 and 13.6±1.28, 2.33±0.55. AQE 100 mg/kg treated group has exhibited an insignificant effect. Amitriptyline had reduced number of head twitches to 1.83±0.47 (Table 5). Effect of extracts on HIC model In the present study haloperidol produced time dependent increase in cataleptic state. In haloperidol treated animals, the peak catalepsy was observed at 150 min. Piracetam potentiated the effect of HIC at all time intervals (0, 30, 60, 90, 120, 150 and 180 min). Mentat treated group showed a significant reduction in severity of catalepsy at all time intervals. All the doses of ALE (50, 75 and 100 mg/kg) and AQE (100, 200 and 400 mg/kg) neither reduced nor potentiated HIC at all time intervals in the experimental study (Table 6). Discussion Nootropics popularly referred as smart drugs which boost human cognitive abilities. Typically, these are alleged to work by increasing the brain s supply of neurochemicals, improving brain s oxygen supply or by stimulating nerve growth. Despite the extensive experimental and clinical studies, the neurochemical basis for learning and memory remains controversial but a predominant role of cholinergic mechanism has long been emphasized in learning and memory processes. The role of the central cholinergic system is fairly well established and its deficiency being implicated in memory deficits 26. Though a large number of other receptor systems too are now reported to be involved in the behavioral expression of dementia in animals and human beings as well the role of these neurotransmitter systems cannot be ignored 27. Table 5 Effect of tuber extracts of P. tuberosa on lithiuminduced (5-HT mediated) head twitches in rats [Values are mean ± SE from 6 animals in each group] No. Treatment Dose (per kg) No. of head twitches for 60 min session I Normal control 10 (ml, po) - II Lithium control 190 (mg, ip) 19.0±2.0 III Piracetam 200 (mg, po) 3.16±0.83** IV Amitriptyline 20 (mg, po) 1.83±0.47** V ALE 50 (mg, po) 14.16±0.98* VI ALE 75 (mg, po) 12.0±0.73** VII ALE 100 (mg, po) 3.5±0.42** VIII AQE 100 (mg, po) 15.0±1.63 ns IX AQE 200 (mg, po) 13.6±1.28* X AQE 400 (mg, po) 2.33±0.55** Significance at P <0.05*, <0.01** and ns-not significant vs lithium control Table 4 Effect of tuber extracts of P. tuberosa on clonidine- induced (NA mediated) hypothermia in rats [Values are mean ±SE from 6 animals in each group] No Treatment Dose (per kg) Rectal temperature ( F) mean ± SE at (min) I Normal control 10 (ml, po) II Clonidine control 0.1 (mg, ip) III Piracetam 200 (mg, po) IV ALE 50 (mg, po) V ALE 75 (mg, po) VI ALE 100 (mg, po) VII AQE 100 (mg, po) VIII AQE 200 (mg, po) IX AQE 400 (mg, po) ± ± ± ± ± ** 96.41* 97.46** ± 96.48* 97.76** 97.4 ± ± ** ± ** 96.96** 97.33** 97.75** ± ± ** 94.85** ± ** ± ** 96.61** ± ** 97.75** ± ** 95.73** ± ** ± 97.36** 97.68** ± ** ± 96.3* 97.5** 97.41** 97.11** ± ** ±0.28 Significance at P <0.05*, <0.01** and ns-not significant vs clonidine control

6 596 INDIAN J EXP BIOL, AUGUST 2008 Table 6 Anti-cataleptic effect of tuber extracts of P. tuberosa on Haloperidol- induced (DA mediated) catalepsy in rats [Values are mean ±SE from 6 animals in each group] No Treatment Dose (per kg) Catalepsy score after (min) I Normal control 10 (ml, po) II Haloperidol control 1 (mg, ip) 0 1.5± 1.83± ± 3.33± 3.83± 3.0± 0.36 III Piracetam 200 (mg, po) 0 1.5± ns 2.33± ns 3.16± ns 4.0± ns ± ns 0.00 IV Mentat 100 (mg, po) ± ** 0.5±** 1.0±** 1.33±** 1.83±** ±** 0.33 V ALE 50 (mg, po) 0 1.0± ns 1.5± ns 2.33± ns 3.33± ns 3.5± ns VI ALE 75 (mg, po) 0 1.0± ns 2.0± ns 2.0± ns 3.5± ns 0.34 VII ALE 100 (mg, po) ± ns 2.0± ns 3.0± ns ± ns ± ns 0.33 VIII AQE 100 (mg, po) ± ns 1.66± ns 2.1± ns 3.33± ns ± ns 3.16± ns 0.30 IX AQE 200 (mg, po) ± ns 1.66± ns 2.5± ns 3.16± ns 3.83± ns X AQE 400 (mg, po) ± ns 2.0± ns ± ns Significance at P <0.05*, <0.01** and ns-not significant vs haloperidol control A simplistic generalization cannot be made in view of the controversial reports available about the role of the central catecholaminergic neurotransmitter system in general and NA system in particular, in learning and memory. It is well known that amphetamines, which markedly augment central noradrenergic activity, leads to mental confusion and retards memory consolidation. The amnesic effect of electroconvulsive shock, which is attenuated by piracetam, is known to produce marked increase in the turnover of NA in rat brain 28. The peripheral as well as central administration of NA was found to suppress avoidance behavior but was facilitatory in some experiments 29. The present study revealed that different extracts (ALE and AQE) of tubers of P. tuberosa contained flavonoids which might be exhibited nootropic activity in view of its facilitatory effect on retention (memory) of acquired learning in mice as these are reported with nootropic activity 22. This observation has been supported by the findings with ALE (50, 75, 100 mg/kg) and AQE (100, 200, 400 mg/kg) of tuber extracts of P. tuberosa that they have shortened the TL in the EPM model indicating an improvement in the memory which is in accordance with the hypothesis proposed by Itoh et al 40. The impairment of learning and memory induced by scopolamine, (0.4 mg/kg), an anticholinergic agent, was reflected by prolonged TL from the open arm to the closed arm i.e., decrease IR 23 was observed with EPM. The ALE (50, 75, 100 mg/kg) and AQE (100, 200, 400 mg/kg) of tuber extracts of P. tuberosa have reversed the amnesia induced by scopolamine, indicates that they are acting through Ach receptors because they had reversed the amnesic effect of scopolamine which is a muscarinic receptor antagonist. Diazepam, a GABA mimetic agent induces memory impairment and the subsequent inhibition of GABA-B receptors has been found to facilitate learning and memory 30,31. Diazepam (5 mg/kg) prolonged TL from the open arm to the closed arm i.e., decreased IR 23. The ALE (50, 75, 100 mg/kg) and AQE (100, 200, 400 mg/kg) of tuber extracts of P. tuberosa have decreased TL from the open arm to the closed arm i.e., increased IR 23 thus confirms their nootropic activity. This protective effect offered against diazepam- induced amnesic model may be due to indirect facilitation of Ach in the brain 37. A reversal action of CIH was observed with ALE (50, 75, 100 mg/kg) and AQE (100, 200, 400 mg/kg) of tuber extracts of P.tuberosa. It can be concluded that there may be an improvement in the transmission

7 RAO et al.: NOOTROPIC ACTIVITY OF TUBER EXTRACT OF PUERARIA TUBEROSA 597 of NA in the hypothalamus as NA activates the hypothalamic cold sensitive neurons, leading to hyperthermia. It has been indicated that an increase in serotonergic transmission in the median raphe of mid brain will interfere with learning acquisition and memory consolidation 32. In the present study with LIH model it was observed that piracetam (200 mg/kg), amitriptyline (20 mg/kg) and different doses of ALE (50, 75,100 mg/kg) and AQE (200, 400 mg/kg) of tuber extracts of P. tuberosa diminished serotonergic function indicated by reduction in number of head twitches as the decrease in brain 5-HT activity can results with anxiolytic activity in this model 33. Controversial reports are available on the involvement of DA activity in learning and memory as learning and memory storage can proceed normally despite depletion of brain DA 34. Piracetam is known to augment dopaminrgic activity 35 and in the present study piracetam (200 mg/kg) potentiated and prolonged the duration of catalepsy. Standard Mentat (100 mg/kg) had showed significant anticataleptic effect and reversed the catalepsy induced by haloperidol, it suggested that Mentat might have increased dopaminergic transmission in the substantia nigra of brain 28,36. The extracts (ALE and AQE) of tubers of P. tuberosa neither reduced nor potentiated, prolonged the catalepsy thus the extracts not interfering with dopaminergic system in the brain. Involvement of chronic inflammation in certain regions of brain and/or free radicals has been implicated in the pathogenesis of AD. Antiinflammatory drugs have been found to be effective in enhancing cognitive function in this condition. The anti-inflammatory action 9 of P. tuberosa (Roxb) reported earlier might be contributing to the memoryenhancing activity observed in the present study. The phytochemical studies with the tuber extracts of P. tuberosa (Roxb) revealed the presence of most of the chemical constituents and one of the important chemical constituent like diadzein, a natural isoflavone found in natural tea plant also reported with this plant which will activate the choline acetyltransferases (ChAT), an enzyme responsible for biosynthesis of acetyl choline in the cholinergic neuronal cells 37. Several studies had shown that the use of estrogen can cause improvement in performance on memory and cognition tests 38, 39 and the earlier studies with this plant had reported that it possess good estrogenic activity 10. Thus the combined effects i.e., activation of ChAT, estrogenic, anti-inflammatory, adaptogenic and neuroprotective roles of tuber extracts of P. tuberosa could be leading to the net memory-enhancing effect Acknowledgement Authors are thankful to management members of AME s V. L. College of Pharmacy, Raichur for encouragement and necessary facilities. References 1 Bennet P N & Brown M J, Clinical pharmacology, 10 th ed., (Churchill Livingstone, New York), 2003, Akio Itoh, Atsumi Nitta, Yuki Katono & Miho Usui, Effects of metrifonate on memory impairment and cholinergic dysfunction in rats, Eur J Pharmacol, 322 (1997) Premnath Shenoy, What is new in the world of pharmaceuticals?, Pharma Times, 39 (2007) Guyton & Hall, Text book of medical physiology, 11 th ed., (Elsevier a division of Reed Elsevier India Private Ltd. India), 2006, Handa S S & Kaul M K, Supplement to cultivation and utilization of medicinal plants, (RRL Jammu-Tawi), 1996, Mentalin: Ayurvedic brain support formula by metagenics(computer program) place/supplements / mind power/desmentalin.html;2004 (cited 2006 Jan 10). 7 Mind power syrup by Maharishi ayurveda, Ayurveda,UK. (computer program). Co.uk/body.soul/mind-power-syrup.asp; 2003(cited 2006 Jan 10). 8 Gupta R S, Rakhi Sharma & Arun Sharma, Antifertility effects of Pueraria tuberosa (Roxb) extracts in male rats, Int J pharmacognosy, 42 (2004) Kambhoja S & Keshavamurthy K R, Wound healing and anti-inflammatory activity of Pueraria tuberosa (Roxb), (APTI.11 th Annual National Convention, Bangalore), 2006, Shukla Sangeeth, Mathur R, & Prakash Ananda O, Effect of butanolic extracts of Pueraria tuberosa DC.on the oestreous cycle of adult rats, Indian J Pharmacol, 19 (1987) Prakash A O, Sexena V, Shukla S & Mathur R, Contraceptive potency of Pueraria tuberosa DC and its hormonal status, Acta Eur Fertil, 16 (1985) Nikam, Hypoglycemic activity of Pueraria tuberosa, J Che Soc, 32 (1980) Babu P V, Rao M A, Kumar S M S & Rao N V, A study on adaptogenic activity of tuber extracts of Pueraria tuberosa, Indian Drugs, 43 (2006) Buger G T & Miller C L, Animal care and facilities, in Principles and methods of toxicology, 2 nd ed., (Wallace Hayes A, Raven Press Ltd. NewYork), 1989, Goyal R K, Practicals in pharamacology, 3 rd ed., (Shah Prakashan, Ahmedabad) 2002, 7.

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