INTRODUCTION:

Depression is a heterogenous mood disorder that has been classified and treated in variety of ways. Although a number of synthetic drugs are being used as standard treatment for clinically depressed patient, they have adverse effects that can compromise the therapeutic treatment. Thus, it is worthwhile to look for antidepressant from plants with proven advantage and favorable benefit to risk ratio. A number of medicinal plants and medicine derived from these plants have shown antidepressant properties by virtue of combined effect of their medicinal constituents.

There are two types of mental depression, namely unipolar depression, in which mood swings are always in the same direction and is common, non-familial, clearly associated with stressful life events, and accompanied by symptoms of anxiety and agitation¹. The second type is bipolar depression, sometimes also called as endogenous depression, shows a familiar pattern, unrelated to external stresses and usually appears in early life, and is much less common, result in oscillating depression and mania over a period of a few weeks¹. Mental depression affects a person’s mood, thoughts, physical health and behavior. Symptoms of depression include biological and emotional components. Biological symptoms include retardation of thought and action, loss of libido, sleep disturbances and loss of appetite. Emotion symptoms include misery, apathy and pessimism, low self-esteem consisting of feeling of guilt, inadequacy and ugliness, indecisiveness and loss of motivation².

Ayurveda the traditional system of Indian medicine mentions a number of plant products which can be used in the treatment of psychiatric disorders. The Ayurvedic concept of Rasayana consists of specialized class of drugs which prevent ageing, increase longevity, impact immunity, improve mental functions and vitality to body³.

Inflammation or phlogosis is a pathophysiological response of living tissue to injuries that leads to the local accumulation of plasmatic fluid and blood cells. Due to severe pain there are incidences that the person losses confidence and feel depressed. Although it is a defence mechanism the complex events and mediators involved in the inflammatory reaction can be induced, maintain aggravate many diseases.The basic way in which the body reacts to infection, irritation or other injury, the key feature being redness, warmth, swelling and pain. Inflammation is now recognized as a type of nonspecific immune response⁴.

Since the depressive disorders as well as inflammation are having a huge impact on our lives, it is worth evaluating the alternative forms of medicines which can be used for its treatment. So in this study, an effort was made to investigate the antidepressant and anti inflammatory effect of ethanolic extract of Curcuma aromatica in experimental animals using different type of models of depression and inflammation.

MATERIALS AND METHOD:

The rhizome of Curcuma aromatica belonging to the family Zingeberaceae were collected from Nambisian’s Kalpavally Stores (Dealers in Ayurvedic pharmaceuticals), Kasaragod , Kerala. It is preserved in the departmental library for future reference.

Animal selection:

Swiss albino mice weighing 18-30 gm, were used for the study. The mice were inbred in the central animal house of the Department of Pharmacology, Karavali College of Pharmacy, Mangalore, under suitable conditions of housing, temperature, ventilation and nutrition were used for antidepressant activity. They were kept in clean dry cages week before the beginning of the experiment to acclimatize with the experimental conditions. The animals were fed with standard pellet diet (Lipton India Ltd., Mumbai) and distilled water ad libitum was maintained at 21°C-23°C under a constant 12hrs light and dark cycle. The animal care and experimental protocols were in accordance with CPCSEA /IAEC.

Preparation of test solution

The freshly collected rhizomes of Curcuma aromatica were washed 2 or 3 times with tap water so that it was made free from all dust materials. They were cut into small pieces and dried. The dried pieces of curcuma aromatica were powdered with the help of mixer grinder and 100g of powder was used for extraction.

A. ANTIDEPRESSANT MODELS:

1. Forced Swim Test

The forced swimming model to test antidepressant activity was developed by Porsolt et al. the model used in the present study was similar to the original method described^5,6.

Procedure:

· Mice were individually placed into a glass cylinder filled with 15 cm of water for 6 min.

· As a measure of depression-like behavior, the total duration of immobility and the number of immobility episodes were recorded. Immobility is defined as the absence of movement, unless they are necessary for the animal to stay afloat (head above water).

· Maintained the temperature of water at 26 ± 1^oC.

· At this height of water, animals were not able to support themselves by touching the bottom or the side walls of the chamber with their hind-paws or tail.

· Water in the chamber was changed after subjecting each animal to FST because “used water” has been shown to alter the behavior.

· Each animal showed vigorous movement during initial 2 min period of the test.

· The duration of immobility was manually recorded during the next 4 min of total 6 min testing period.

· Mice were considered to be immobile when they ceased struggling and remained floating motionless in water, making only those movements necessary to keep their head above water. Following swimming sessions, the mice were dried with towel and placed in a cylinder heated under 60 W bulb. The animals were dried under heated cylinder for 15 minutes before returning to the home cages.

Groups of Animals :

The animals were divided as follows.

· Group I – Received 0.05ml/10g of Normal saline intraperitoneally.

· Group II – Received 15 mg/kg Imipramine intraperitoneally.

· Group III – Received 250 mg/kg Curcuma aromatica Oil orally.

· Group IV – Received 500 mg/kg Curcuma aromatica Oil orally.

· Group V – Received 750 mg/kg Curcuma aromatica Oil orally.

2. Tail Suspension Test:

Mice tail suspension test (TST) is one of the most frequently employed test for screening of antidepressant activity. In the tail suspension test, mice initially engaged in vigorous escape behaviours but eventually succumb to immobility. Like the FST, longer duration of TST immobility infer a heightened degree of behavioural despair. As such, TST is a commonly used screening method for antidepressant properties of drug and highly sensitivity to pharmacological manipulations. Antidepressant drug generally decrease the duration of TST immobility in mice⁶.

Procedure:

Animals were moved from their housing colony to laboratory in their own cages and allowed to adopt to the laboratory conditions for 1-2 hr.

· Each mice was individually suspended to the edge of a table, 50 cm above the floor by adhesive tape placed approximately 1 cm from the tip of the tail.

· Each animal under test was both acoustically and visually isolated from other animals during the test.

· Total period of immobility was recorded manually for 6 min.

· Animal was considered to be immobile when it didn’t show any body movement, hung passively and completely motionless.

· The test was conducted in a dim lighted room and each mice was used only once in the test.

· The observer, recording the immobility of animals was blind to the drug treatment given to the animals under study ^6,7.

Groups of Animals :

The animals were divided as follows.

· Group I – Received 0.05ml/10g of Normal saline intraperitoneally.

· Group II – Received 15 mg/kg Imipramine intraperitoneally.

· Group III – Received 250 mg/kg Curcuma aromatica Oil orally.

· Group IV – Received 500 mg/kg Curcuma aromatica Oil orally.

· Group V – Received 750 mg/kg Curcuma aromatica Oil orally.

Statistical Analysis:

Results are prepared as Mean ± SEM. One way ANOVA was used for multiple comparison followed by Dunnett’s multiple comparison tests. For all tests a “P” value of 0.05 or less was considered for statistical significance.

ANOVA (Analysis of variance):

In statistics, analysis of variance is a collection of statistical models and their associated procedures, in which the observed variance is partitioned into components due to different explanatory variables. In its simplest form ANOVA gives a statistical test of whether the means of several groups are all equal and therefore generalize Dunnett’s multiple comparison tests to more than two groups.

B. ANTI-INFLAMMATORY ACTIVITY:

1. Carrageenan paw edema model⁸

Pedal inflammation was produced in rats according to the carrageenan induced paw edema method. 30 albino rats were divided into four groups of six each and fasted overnight for 18 hrs with water. Next day the animals were weighted and numbered. A mark was made on the right hind paw just beyond tibio-tarsal junction, so that every time the paw was dipped in the mercury (Hg) column up to the fixed mark to ensure constant paw volume. The initial paw volume was noted of each rat by mercury (Hg) displacement method. 0.1 ml of 1 % carrageenan was injected into the right hind paw of each rat under the sub plantar aponeurosis. The animals were treated orally with extracts (100,200 and 400 mg/kg), Ibuprofen (100mg/kg) or Saline (10 ml/kg) 1 h before carrageenan injection. The paw volume of each rat after carrageenan injection after one hour, two hour, three hour and five hour and 24^th was recorded by mercury displacement in plethysmograph. This volume is called final volume.

The anti-inflammatory activity of the extract was measured by its potential to prevent edema caused by carrageenan as against the control group which was given the vehicle only. Mean paw edema was calculated for each group as average of paw volume of individual rats belonging to that group. Since, the mean was subjected to positive and negative fluctuations hence; standard error for each group was also calculated.

Standard error (S.E) = standard deviation

√ n

Where, n = number of rats in each group

Percent inhibition of paw edema was calculated according to the following formula:

% Inhibition = a-b/a x 100

a- is the mean paw inflammation volume of control.

b- is the mean paw inflammation volume of test

Statistical Analysis:

The results were expressed as mean + SEM, Statistical significance was determined by one way ANOVA (Analysis of Variance) followed by t test by using the Graph pad Instant version and compared with control.

RESULTS:

A. Antidepressant activity of curcuma aromatica oil

1. Forced Swim Test :

In FST, Table shows that animals treated with three doses of (Curcuma aromatica oil) CAO (250, 500 and 750 mg/kg) showed decrease in their immobility times, which was significant (137.5±1.30; p<0.05 and 134.12±1.21, 122.15±1.17; p<0.001) when compared with control (142.22±1.73). Similarly, animals treated with Imipramine (15 mg/kg), as expected, showed a significant decrease in the immobility time (64.27±1.17; p<0.001). Animals treated with high dose (750 mg/kg) and moderate dose (500 mg/kg) shows more significant decrease in immobility time when compared with low dose (250 mg/kg).

Table 1.Effect of CAO on Immobility time in FST

Group No.	Drug Treatment	Dose (mg/kg)	Immobility period, mean ± S.E.M [n=6]
I	Control	0.05 ml/10 g	142.22±1.73
II	Imipramine	15	64.27±1.17***
III	CAO (Low dose-LD)	250	137.5±1.30*
IV	CAO (Moderate dose-MD)	500	134.12±1.21***
V	CAO (High dose-HD)	750	122.15±1.05***

GRAPH NO: 1 Effect of CAO on Immobility time in FST

Values were mean ± S.E.M. for (n=6) expressed as the time (in sec) of 6 animals each group. Data analysis was performed using Dunnett’s test.^*P < 0.05, ^**P < 0.01, ^***P < 0.001.

Table. 2 Effect of CAO on Immobility time in TST.

Group No.	Drug Treatment	Dose (mg/kg)	Immobility period, mean ± S.E.M [n=6]
I	Control	0.05 ml/10 g	160.15 ± 1.40
II	Standard	15	72.13±1.68***
III	CAO	250	158.13±1.49*
IV	CAO	500	147.18±1.07**
V	CAO	750	128.78±1.30***

2. Tail Suspension Test :

Animals treated with three doses of CDO showed decrease in their immobility times, which was significant (158.13 ± 1.49; p<0.05, 147.18 ± 1.07; p<0.01 and 128.78 ± 1.30; p<0.001) when compared with control (160.15 ± 1.40). Similarly, animals treated with Imipramine (15 mg/kg) as expected showed a significant decrease in the immobility time (72.13 ± 1.68; p<0.001). Animals treated with high dose (750 mg/kg) showed more significant decrease in immobility time.

GRAPH NO: 2 Effect of CAO on Immobility time in TST

Values were mean ± S.E.M. for (n=6) expressed as the time (in sec) of 6 animals in each group. Data analysis was performed using Dunnett’s test.^*P < 0.05, ^**P < 0.01, ^***P < 0.001 vs. Control.

B. Anti-Inflammatory activity of curcuma aromatica ethanolic extract (CAE)

1. Carrageenan induced paw edema

The effect of standard Ibuprofen at dose of 100 mg/kg and test drug CAE at three different concentrations 100,200 and 400 mg/kg on carrageenan induced acute inflammation is tabulated and also shown in Figure. After injection of carrageenan a progressive increase in paw volume was observed in the control group and found to be maximum at hour 4. After 4 h decrease in paw volume was observed.

In 1^st there was a significant reduction in paw volume was observed in group II (P<0.001), group IV (P<0.01) and group V (P<0.001) animals, when compared to the control group. Animals of group III did not reveal significant reduction in paw volume, compared to the control group animals. In 2^ndhour the animals of group II, group IV and group V were showed significant (P<0.001) reduction in paw volume when compared to control group. The animals of group III did not showed reduction in paw volume compared to control group. In 3^rd a significant (P<0.001) reduction in paw volume of group II, group IV and group V were observed when compared to the control group. The animals of group III did not show significant reduction in paw volume.

In 4^th hour a significant reduction in paw volume of group II (P<0.001), group III (P<0.01), group IV (P<0.001) and group V (P<0.001) were observed when compared to the control group. The animals of group II, group IV and group V showed significant (P<0.001) reduction in paw volume when compared to control group in 5^th hour. Animals treated with Ibuprofen at dose of 100 mg/kg showed significant inhibition of biphasic response of acute inflammation produced by carrageenan. Animals treated with CAE 400 mg/kg body weight showed significant inhibition of both phases of inflammation produced by carrageenan. But animals treated with CAE 200 mg/kg suppressed only the second phase of carrageenan induced inflammation. A dose dependent reduction in edema was observed by CAE extracts and the potency of anti-inflammatory activity of AEEI was found to be: CAE 400 > CAE 200 > CAE 100.

Table 3. Effect of CAE on carrageenan induced acute inflammation in rats

Groups	Treatment	Mean change in paw volume (ml) and percentage protection ± SD
Groups	Treatment	1h	2h	3h	4h	5h
I	Control	0.81±0.04	0.95±0.05	1.03±0.05	1.07±0.10	1.02±0.09
II	Ibuprofen Standard	0.40±0.04*** (51.75±5.04)	0.49±0.02*** (48.47±2.13)	0.56±0.03*** (45.38±3.62)	0.46±0.03*** (56.80±3.54)	0.32±0.03*** (68.16±3.09)
III	CAE 100	0.78±0.04 (5.02±5.64)	0.88±0.04 (7.42±4.58)	0.95±0.05 (7.63±5.63)	0.92±0.5 (13.61±5.11)	0.80±0.06 (21.22±6.46)
IV	CAE 200	0.74±0.02** (10.55±3.11)	0.80±0.03*** (15.28±3.94)	0.85±0.02*** (18.07±2.63)	0.79±0.04*** (25.68±3.74)	0.71±0.04*** (30.20±4.02)
V	CAE 400	0.61±0.02*** (26.13±2.52)	0.66±0.01*** (30.56±1.43)	0.69±0.02*** (33.33±2.48)	0.65±0.03*** (39.29±3.30)	0.57±0.03*** (43.26±3.25)

Values are mean±SD of six animals, values in bracket indicate percentage protection.

***P<0.001; **P<0.01 when compared to control group.

GRAPH NO: 3.1 Effect of CAE (Curcuma aromatica extract) on carrageenan induced inflammation.

GRAPH NO: 3.2 Percentage protection shown by CAE (Curcuma aromatica extract) on carrageenan induced edema

DISCUSSION:

The purpose of current research was to investigate the anti-depressant activity of Curcuma aromatica oil in experimental animals. The main finding of present investigation suggests the antidepressant activity of CDO in different animal models of depression. The review on pathophysiology of depression reveals that the depression occurs due to many reasons and many hypothesis has been proposed on it. But widely accepted mechanism involved in pathogenesis of depression is monoamines deficiency. Certain biological monoamines like NA and 5-HT, dopamine especially decrease in NA and 5-HT causes depressive episodes in patients suffering from depression which makes their life miserable.

The incidence of depression in the community is very high and is associated with lot of morbidity. Hence, it is very important to address these problems and find effective remedies. Though several drugs are available, all are associated with some limitations and there is an urgent need for alternative medications for these disorders. Despite the widely popular use of CAO for treating nervous disorders, there is an absence of scientific reports about the evaluation of its pharmacological effects. In this work, it was demonstrated that the administration of different doses of CAO in mice was able to induce antidepressant effects.

In present study, behavioural models namely forced swim test and tail suspension test were employed. Both represents the behavioural despair models, claimed to reproduce a condition similar to human depression. The tests were based on the observation that animals, following initial escape oriented movements, develop an immobility posture when placed in an inescapable chamber. The immobility is thought to reflect either a failure of persistence in escape directed behaviour or development of passive behaviour that disengage the animal from active form of coping with stressful stimuli⁹.

We observed that following oral administration of CAO demonstrated significant (compared to control treated group) dose dependent reduction in duration of immobility. It has been established that the shortening of immobility time in the forced swimming and the tail suspension tests depends mainly on the enhancement of central 5‐HT and catecholamine neurotransmission¹⁰. Early evidence of a role for noradrenaline in depression came from the discovery that drugs, either causing or alleviating depression, acted to alter the noradrenaline metabolism. Furthermore, depletion studies carried out in treated and untreated patients indicated a role for serotonin and noradrenaline in depression¹¹.

The results indicate that essential oil of Curcuma aromatica may have an antidepressant like effect and the immobility time observed in the test reflected a state of lowered mood or hopelessness in animals, thus, this animal model is the most widely used tool for preclinical screening of putative antidepressant agents. The FST shows a strong sensitivity to monoamine alterations and is a very specific cluster of stress-induced behaviours that are not related to depression symptoms in humans, but which are nonetheless exquisitely sensitive to monoaminergic manipulations. It also provides a useful model to study neurobiological and genetic mechanisms underlying stress and antidepressant responses.

Forced swim test is most widely used test for screening of antidepressants involves placing mice in cylinder of water from where there is no escape and measuring the animal’s behaviour for several minutes. Initially, rodent’s displays escape orientation behaviours, however, their behaviour changes eventually into movement that are just sufficient to keep their head above the water-termed immobility. This was originally interpreted by Porsolt et al as “behaviour despair” such that the animals has lost the motivation to perform escape oriented behaviour. This behavioural immobility reflecting a state of despair is reduced by a broad spectrum of antidepressant drugs¹².

In tail suspension test, it has been argued that the TST is less stressful than FST and has greater pharmacological sensitivity. In this test, mice are suspended by their tail for defined period of time and duration of their immobility is assessed. Typically, animals are immediately engaged in escape oriented behaviour followed by progressive increasing period of immobility. We observed that following oral administration of CAO demonstrated significant (compared to control) a dose dependent reduction in duration of immobility¹³.

In both FST and TST, the results clearly revels the fact that standard treated animals showed better response as compared to CAO. The CAO treated animals showed good response as compared to control. Data in the literature demonstrated that drugs that alter general motor activity may give false‐positive/negative results in the forced swimming test. The exact mechanism underlying antidepressant effect of essential oil of Curcuma aromatica is not clear but it may be apparently related to active compounds present in CAO are reviewed¹⁴. We cannot discard the possibility that more than one compound are the responsive for its antidepressant activity. Chemical studies have reported the presence of several monoterpenoid compound in the essential oil of Cinnamomum camphora, primarily, β-pinene, β-thujone, limonene and also linalool are reported to have antidepressant activity¹⁵.

The results indicate that essential oil of Curcuma aromatica may have an antidepressant like effect. However further experiments evaluating the levels of noradrenaline and serotonin in different regions of brain are necessary to confirm this hypothesis.

The therapeutic applications of flavonoids in inflammation have been previously reported⁴. Therefore, intake of flavonoids is very important in the management of this condition. In addition, flavonoids are known to prevent the synthesis of prostaglandins. Biochemical investigations on the mechanism of action of flavonoids have shown that this compound can inhibit a wide variety of enzymes¹⁶_, which are involved in inflammation processes. The ability of flavonoids to inhibit both cyclooxygenase and lipooxigenase pathways of arachidonate metabolism have been suggested to contribute to its anti-inflammatory action. In vivo and in vitro anti-inflammatory effects have been reported for several flavonoids thus, some 6-methoxylated flavones have been found to be active in different experimental models. B-ring substituted flavones of this type are capable of inhibiting cotton pellet induced granuloma and carrageenan edema in rats¹⁷.

Inflammation plays an important role in many serious diseases, including cancer, Alzheimer’s and AIDS¹⁷. Inflammation induced by carrageenan involves three distinct phases of the release of the mediator, including serotonin and histamine in the first phase (0 to <2h), kinins in the second phase (3 h) and prostaglandin in the third phase (>4h)¹⁸. It is a standard experimental model of acute inflammation. Carrageenan is the phlogistic agent of choice for testing anti-inflammatory drugs as it is not known to be antigenic and is devoid of apparent systemic effects¹⁹.

CONCLUSION:

The present study was aimed to expose the antidepressant activity of Curcuma aromatica Oil in Swiss albino rat using four animal models of depression namely, Forced swim test and Tail suspension test.

The data obtained was satisfactory and conclusive and so as to accomplish our objectives. In conclusion the present data indicate that the administration of Curcuma aromatica oil to rat has shown significant dose dependant antidepressant activity supporting folk information regarding antidepressant activity of the essential oil, relatively sub-chronic study may be necessary to arrive at a better picture. The exact mechanism underlying antidepressant effect is not clear but it may be apparently related to active compounds present in Curcuma aromatica oil. Hence further studies would be necessary to evaluate the contribution of active chemical constituents for the observed antidepressant activity as it still remains to be determined which components were responsible for these effects.

The ethanolic extract of Curcuma aromatica was used to evaluate the anti-inflammatory and it was found that Curcuma aromatica exhibited dose dependent anti-inflammatory activity. The exact mechanism by which Curcuma aromatica exhibit anti-inflammatory activity is still to be studied it is presumed that it might be due to the presence of flavonoids in Curcuma aromatica.

REFERENCES:

1. Gabbard, Glen O. Treatment of Psychiatric Disorders. 3^rd ed. Washington, DC: American Psychiatric Publishing; 2000.

2. Gold PW, Goodwin FK and Chrousus GP. Clinical and biochemical manifestations of depression in relation to the neurobiology of stress. Part1. New England Journal of Medicine. 1988; 319: 348 – 353.

3. Sharma PV, Cakradutta A. Treatise on principle and practice of Ayurvedic Medicine.Delhi.1994:173.

4. Sosa S, Balick MJ, Arvigo R, Esposito RG, Pizza C, Altinier GA. screening of the topical anti-inflammatory activity of some central American plants. J. Ethnopharmacol.2002;(8):211-215.

5. Bowden CL, Calabrese JR, McElroy SL, Gyulai L, Wassef A, Pretty F, et al. A randomized, placebo-controlled 12-month trial of divalproex and lithium in treatment of outpatients with bipolar I disorder. Divalproex Maintenance Study Group. Arch Gen Psych 2005; 57(5):481-9.

6. Tylor FB, Prather MR. The efficacy of nefazodone augmentation for treatment-resistant depression with anxiety symptoms or anxiety disorder. Depress Anxiety. 2003;18(2):83-8.

7. Feighner J, Targum SD, Bennett ME, Roberts DL, Kensler TT, D’Amico MF, et al. A double-blind, placebo-controlled trial of nefazodone in the treatment of patients hospitalized for major depression. J Clin Psychiatry 1998; 59(5):246-53.

8. Joshni Punam ,Priya Banu,Gahlot Manoj. Antiinflammatory and analgesic activities of methanol extract of “Vallaris solanaceae leaves”. International Journal of Pharmacy and Pharmaceutical Sciences: 6(2);2014.

9. A.R.Srividya, S.Dhanapal, A comparative study on Genomic activity of Hydro alcoholic extract of Curcuma aromatica and Curcuma zedoaria rhizomes. abstract/ wwww.shodhanga. inflinet.ac.in.

10. www.herbalveda.co.uk/index dated 17/03/2015

11. Gorman JM. Mirtazapine : Clinical Overview. J Clin Psychiatry 1999; 60(17):9-13. Horst WD, Preskon SH. Mechanism of action and clinical characteristic of three atypical antidepressants : Venlafaxine, Nefazodone, Bupropion. J Affect Disorders 1998; 51(3):237-54.

12. Bauer M, Dopfmer S. Lithium augmentation in treatment-resistant depression : Meta-analysis of placebo-controlled studies. J Clin Psychopharmacol 1999; 19(5):427-34.

13. Barbosa L, Berk M, Vorster M. A double-blind, randomized, placebo-controlled trial of augmentation with lamotrigine or placebo in patients concomitantly treated with fluoxetine for resistant major depressive episodes. J Clin Psychiatry 2003; 64(4):403-7.

14. Smith D, Dempster C, Glanville J, Freemantle N, Anderson I. Efficacy and tolerability of Venlafaxine compared with selective serotonin reuptake inhibitors and other antidepressants : A meta-analysis. Br J Psychiatry Suppl 2002; 180:396-404.

15. Keller MB, McCullough JP, Klein DN, Arnow B, Dunner DL, Gelenberg AJ, et al. A comparison of nefazodone, the cognitive behavioural-analysis system of psychotherapy, and their combination for the treatment of chronic depression. N Engl J Med. 2000; 342(20):1462-70.

16. Ruhe HG, Mason NS, Schene AH. Mood is indirectly related to serotonin, norepinephrine and dopamine levels in humans: a meta-analysis of monoamine depletion studies. Mol Psychiatry. 2007; 12(4):331-59.

17. Turner EH, Matthews AM, Linardatos E, Tell RA, Rosenthal R. Selective publication of antidepressant trials and its influence on apparent efficacy. N Engl J Med. 2008; 358(3):252-60.

18. Jorenby DE, Leischow SJ, Nides MA, Rennard SI, Johnston JA, Hughes AR, et al. A controlled trail of sustained release bupropion, a nicotine patch or both for smoking cessation. N Engl J Med 1999; 340(9):685-91.

19. Belmaker RH, Agam G. Major depressive disorder. N Engl J Med. 2008; 358(1):55-68.

Received on 16.03.2016 Accepted on 08.04.2016

Asian J. Pharm. Res. 2016; 6(2): 79-86

DOI: 10.5958/2231-5691.2016.00014.9