Skip to main content
NCBI home page
Journal of Ayurveda and Integrative Medicine logoLink to Journal of Ayurveda and Integrative Medicine
. 2023 May 16;14(2):100707. doi: 10.1016/j.jaim.2023.100707

Experimental evaluation of hypnotic and antidepressant effect of pine needles of Cedrusdeodara

Nirmal Kumar E a, Padmaja A Marathe b,∗, Sandhya K Kamat b, Harshitha Havaldar b, Merin Eldhose a, Pritika Mall c
PMCID: PMC10203744  PMID: 37201295

1. Introduction

Depression is the most common psychiatric disorder affecting about 280 million people in the world [1]. The National Mental Health Survey carried out in 2015–16 reported that, one in 20 people in India suffer from depression [2]. Currently used antidepressants pose limitations due to their adverse effects and moderate efficacy [3,4]. Insomnia is regarded as the most common sleep disorder, observed in 30% of the general population [5]. Insomnia is a clinical condition characterized by difficulty in onset and/or maintenance of sleep despite of having adequate circumstances for sleeping [6]. Use of benzodiazepines and Z-drugs are associated with adverse effects like drug dependence, tolerance, amnesia, gastrointestinal upset, residual daytime drowsiness, and withdrawal disorders [7]. The most effective hypnotics developed to date, work by enhancing Gamma-Aminobutyric Acid (GABA) [8].

In clinical practice, insomnia and depression often co-exist [9]. There is a strong association between insomnia and depression. Diagnosing a case of depression in the absence of sleep disturbances should be made with caution. About 3/4th of patients of depression has symptoms of insomnia.There is a need for addressing the problem of sleep disturbances in depressed patients, in order to improve their quality of life and to reduce an important contributing factor for relapse of depression [10].

The involvement of GABA in depression has long been suspected [11]. There are preclinical studies indicating a causal role of GABA deficits in the etiology of depressive disorders. Further, GABAergic transmission plays an essential role in hippocampal neurogenesis and neural maturation, which are known to be the key mechanisms of currently used antidepressants [12,13].

Cedrus deodara (CD) commonly called as “deodar” belongs to the family Pinaceae that grows in abundance in the Himalayan region [14]. Ayurveda, the traditional system of medicine mentions that different parts such as bark, needles, heartwood of CD are used for treatment of various conditions such as inflammation, hyperglycemia, infections, insomnia, mind & skin disorders [15]. A review of literature revealed that all parts of the plant have been tested for various pharmacological activities in preclinical studies such as heartwood extract for anti-convulsant activity, anti-depressant activity and anti-diabetic activity [[15], [16], [17]], root extract for anti-fungal activity [18], needle isolate for anti-microbial activity [19] and cone extract for anti-diabetic activity [20]. CD has been tested in experimental study to explore activities of all parts, as all parts have been shown to be useful. Traditionally, it was used to treat rheumatoid arthritis, arthralgia, sleeplessness, edema, traumatic injury and eczema [21]. CD root oil in the dose of 200 mg/kg had comparable antiulcer effect in gastric ulcer to that of omeprazole in rats [22].

A study conducted by Viswanatha GL. et al. showed that CD demonstrated significant anxiolytic activity through modulation of GABA levels in brain [23]. Dhayabaran. et al. reported that CD showed anticonvulsant activity and it led to increase of GABA levels in the brain of rats [16]. A study conducted by Kumar et al. reported that CD showed significant antidepressant activity in FST whereas it did not significantly alter the immobility time in TST. It also showed raised brain monoamine (serotonin and nor-adrenaline) levels [24]. CD has been found to have anxiolytic, antidepressant & antiepileptic effects in animals through modulation of GABA levels in brain [16,23,24]. There is only one study which has reported hypnotic activity of essential wood oil of CD [25].

It was of interest to study antidepressant and hypnotic effects of CD which appears to act through modulation of GABA. If found effective in preclinical and clinical studies, CD will be a useful and safer treatment option for insomnia and depression based on its potential GABA modulating effect. Hence, the present study was planned using aqueous and alcoholic extracts of pine needles of CD, the part of the plant which has not been explored in the experimental models of depression and hypnosis. It was also decided to evaluate mechanism of hypnotic action of CD using a GABA antagonist.

2. Materials and methods

The study was conducted in two stages. In phase 1, rats were used to explore the antidepressant properties of formulations of C. deodara pine needles. In Phase 2, mice were used to test the hypnotic potential of formulations of C. deodara pine needles. Throughout the experiment, observations were made by observers who were blinded to the treatment allocations. The institutional animal ethics committee's approval was obtained before initiating the study experiments (Approval reference: 08/2019). The study was conducted in accordance with the Committee for the Control and Supervision of Experiments on Animals (CCSEA) recommendations.

2.1. Experimental animals

Animals from the Institute's Central Animal House (registered with CCSEA) were used in this study. Randomly bred animals were chosen for the experiments. Ninety six albino mice of either sex were randomly selected for the hypnotic model; having an age-range of 6–8 weeks and weighing 20–25 g. For the purpose of assessing antidepressant activity, 40 Wistar rats of either sex, weighing between 150 and 250 g, were randomly chosen. The Central Animal House was maintained in compliance with standard norms for animal housing as described in the CPCSEA guidelines, which included a temperature range of 23 °C–4 °C, a relative humidity range of 30–70%, and 12-h light/dark cycles. The animals were contained in stainless steel top grilled polypropylene cages (8 mice and 4 rats per cage) with facilities for supplying food (in the form of Pellets) and drink (free access to UV-filtered water). Paddy husk was utilized as cage bedding.

2.2. Study drugs

The test drug formulations - aqueous (AQ) and alcoholic (AL) extracts of C. deodara pine needles were procured from (Pharmanza Herbals Pvt. Ltd.). The herb extract ratios of the two extracts were AQ - 6.4 and AL - 7.14. Normal saline (0.9%) was administered to the control group. Thiopental sodium obtained from our hospital Pharmacy and was used as hypnosis inducing agent. Diazepam (DZ) obtained from our hospital Pharmacy served as the standard hypnotic control and Fluoxetine (FX) purchased from Sigma (USA) was used as the standard antidepressant control. Picrotoxin, a GABA antagonist was included in the study in Phase 2 to evaluate mechanism of action was bought from Sigma (USA). All study medications were administered intraperitoneally (I.P).

2.3. Stage 1 - evaluation of anti-depressant action of C. deodara using Chronic Unpredictable Mild Stress Model (CUMS)

2.3.1. Study procedure

Chronic Unpredictable Mild Stress Model (CUMS) depression model was already standardized and validated by previous researcher in our institute [26,27]. Hence, due to availability of a standardized validated model and to keep animal usage to minimum, normal control group was not included in this model in the present study [28]. Chronic unpredictable mild stress was given over a total duration of 28 days in order to induce depression in rats. Following are the seven different stressors that were given in random order to establish the model of CUMS induced depression:

  • 1.

    Food deprivation for 12 h

  • 2.

    Water deprivation for 12 h

  • 3.

    Cage tilt for 18 h

  • 4.

    Damp saw dust for 12 h

  • 5.

    Grouped housing for 12 h

  • 6.

    Shaking for 10 min

  • 7.

    Continuous lightning for 24 h

The stressor sequence was changed every week. Fig. 1, presents the type of stressors given over 28 days.

Fig. 1.

Fig. 1

Open in a new tab

Stressors given in randomized manner in CUMS model.

Rats were randomly divided into five groups of eight animals each in stage 1. The study groups were as follows: Vehicle control (Saline -5 ml/kg i.p), Standard control (Fluoxetine - 5 mg/kg i.p) [29,30], two doses of Aqueous extract of CD (250 & 500 mg/kg i.p) [31] and one dose of Alcoholic extract of CD (100 mg/kg i.p) [24].

The study medications were administered to the respective groups for 28 days. On days 27 and 28, the rats were prepared for a forced swimming test (FST). They underwent Open Field Test (OFT) on day 29, which was followed by an experiment involving forced swimming. The purpose of the OFT was to determine whether a change in an animal's immobility during the forced swim test was secondary to a change in their motor activity. This OFT was important to perform to distinguish the general behavioural stimulation (false positive) effect of study medications from the anti-depressant effect [30,32].

The dimensions of the OFT apparatus are as follows: Floor: 60 × 60 cm, Wall height: 25 cm. The floor and wall are black, the ceiling is open kept in a sound-attenuated, dark room, with minimal background lighting. The rat was handled by its tail (base). It was placed in the centre of the OFT apparatus and was allowed to explore the apparatus for 5 min [33,34].

Rat was forced (one at a time) to swim inside the plexiglass cylinder for 15 min then wiped with cotton and dried under the lamp. This preconditioning was done to induce despair. The next day (24 h later) rat was made to swim for 5 min [35,36].

Stage 2: Thiopental-induced hypnosis in mice was the model used in this study. This model was already validated in our institute.

Stage 2 was divided into two parts. Two dosages of the aqueous and alcoholic extracts of CD were used in Phase 2A in order to determine the effective dose. In Phase 2 B, picrotoxin, a GABA antagonist was used to explore the mechanism of hypnotic action of CD. stage 2 study procedures are depicted in Fig. 5, Fig. 6, Fig. 7, Fig. 8.

Fig. 5.

Fig. 5

Open in a new tab

Phase 2A- sleep latency.

VC – Vehicle Control; DZ – Diazepam; AQ-50 - Aqueous Extract of CD (50 mg/kg); AQ-100 - Aqueous Extract of CD (100 mg/kg); AL-250 - Alcoholic Extract of CD (250 mg/kg); AL-500 - Alcoholic Extract of CD (500 mg/kg).

Values expressed as mean ± SD, ∗p < 0.001 compared to VC, #p < 0.05 compared to AL-500, $ p < 0.05 compared to AL-250 using ANOVA with post-hoc Tukey's test.

Fig. 6.

Fig. 6

Open in a new tab

Phase 2A- total sleeping time.

VC – Vehicle Control; DZ – Diazepam; AQ-50 - Aqueous Extract of CD (50 mg/kg); AQ-100 - Aqueous Extract of CD (100 mg/kg); AL-250 - Alcoholic Extract of CD (250 mg/kg); AL-500 - Alcoholic Extract of CD (500 mg/kg).

Values expressed as mean ± SD, ∗∗p < 0.001, ∗p < 0.01 compared to VC, #p < 0.001 compared to AQ-50, AQ-100, AL-250 & AL-500, $ p < 0.05 compared to AQ-100 using ANOVA with post-hoc Tukey's test.

Fig. 7.

Fig. 7

Open in a new tab

Phase 2B- sleep latency on addition of picrotoxin.

VC – Vehicle Control; DZ – Diazepam; AQ-100 - Aqueous Extract of CD (100 mg/kg); P + AQ-100 – Picrotoxin + Aqueous Extract of CD (100 mg/kg); AL-500 - Alcoholic Extract of CD (500 mg/kg); P + AL-500 - Picrotoxin + Alcoholic Extract of CD (500 mg/kg). Values expressed as mean ± SD, ∗∗p < 0.001, ∗p < 0.05 compared to VC, #p < 0.001 compared to AQ-100, AL-500, P + AQ-100 & P + AL-500, $ p < 0.01 compared to AQ-100 using ANOVA with post-hoc Tukey's test.

Fig. 8.

Fig. 8

Open in a new tab

Phase 2B- total sleeping time on addition of picrotoxin.

VC – Vehicle Control; DZ – Diazepam; AQ-100 - Aqueous Extract of CD (100 mg/kg); P + AQ-100 – Picrotoxin + Aqueous Extract of CD (100 mg/kg); AL-500 - Alcoholic Extract of CD (500 mg/kg); P + AL-500 - Picrotoxin + Alcoholic Extract of CD (500 mg/kg). Values expressed as mean ± SD, ∗p < 0.001 compared to VC, #p < 0.01 compared to AQ-100, AL-500, P + AQ-100 & P + AL-500, $ p < 0.001 compared to AQ-100 using ANOVA with post-hoc Tukey's test.

2.4. Stage 2: thiopental induced hypnosis [36,37]

In stage 2A, mice were randomized into six groups of 8 animals each as follows: Vehicle control (Saline -5 ml/kg i.p), Standard control (Diazepam (2 mg/kg i.p) [36], two doses of Aqueous extract of CD (50 & 100 mg/kg i.p) [17] and two doses of Alcoholic extract of CD (250 & 500 mg/kg i.p) [15]. The mice in each group received a single intraperitoneal injection of the vehicle, diazepam (2 mg/kg), and the test medications. This was done 30 min before to giving thiopental (60 mg/kg) intraperitoneally [38].

When turned on its back, an animal's inability to straighten out itself demonstrates the loss of righting reflex. Indicating onset of sleep. After certain period of time, when animal wakes up and adjusts its posture. The first movement made by the mouse to adjust the posture indicates the return of the righting reflex and thus end of sleeping state. Thus, the duration of total sleeping time and the latency of each mouse to fall asleep was noted in all the study groups. After noting the sleep latency and sleep duration for each mouse animal, it was positioned on its back once more and permitted to adjust its posture again. This was carried out to ensure that the righting reflex was completely regained. Once the righting reflex was regained, animal was place back in its cage.

In stage 2B, mice were randomized into six groups of 8 animals each as follows: Vehicle control (Saline -5 ml/kg i.p), Standard control (Diazepam (2 mg/kg i.p), Aqueous extract of CD (100 mg/kg i.p), Alcoholic extract of CD (500 mg/kg i.p), Aqueous extract of CD with Picrotoxin (1 mg/kg I.P) [39] and Alcoholic extract of CD with Picrotoxin (1 mg/kg I.P).

The dose of each type of extract of CD that produced the greatest effect was selected to further evaluate the mechanism of hypnotic action of CD. The procedure was the same as described in stage 2A, with the exception that picrotoxin was given intraperitoneally 30 min before thiopental administration. The extracts were administered to the two CD groups following administration of picrotoxin.

2.5. Statistical analysis

Data of each study group was compiled and was expressed as mean ± SD. The data was analysed using Graphpad-InStat version 3.06. Level of significance was set at p < 0.05. Normality was checked by Kolmogrov–Smirnoff Test. The study variables were analysed using one-way ANOVA followed by post hoc Tukey's test.

3. Results

3.1. Stage 1: Evaluation of antidepressant activity

3.1.1. Time spent in the centre in the open field apparatus

The time spent in the centre of the open field apparatus by the rats in all groups is presented in Fig. 2. There was statistically significant increase in the time spent in the centre by the rats in the fluoxetine (FX) group as compared to those in the vehicle and all the three CD groups (p < 0.001). There was no significant change in the time spent in the centre in aqueous extract (AQ-250 and AQ-500) groups when each group was compared to the vehicle control group. The alcoholic extract (AL-100) group showed increase in the duration of time spent in the centre which was significantly higher compared to the vehicle control (p < 0.001). The rats treated with AL-100 stayed for significantly longer time in the centre when compared to those treated with AQ-250 and AQ-500.

Fig. 2.

Fig. 2

Open in a new tab

Time spent in the centre in the open field Apparatus.

VC – Vehicle Control; FX – Fluoxetine; AQ-250 - Aqueous Extract of CD (250 mg/kg); AQ-500 - Aqueous Extract of CD (500 mg/kg); AL-100 - Alcoholic Extract of CD (100 mg/kg).

Values expressed as mean ± SD, ∗p < 0.001 compared to VC, #p < 0.001 compared to AQ-250, AQ-500 & AL-100, $ p < 0.001 compared to AQ-250 & AQ-500 using ANOVA with post-hoc Tukey's test.

3.1.2. Number of lines crossed in the open field apparatus

The number of lines crossed in 5 min by the rats in all the groups is given in Fig. 3. The statistical comparison among the groups showed that there was no difference in the number of lines crossed.

Fig. 3.

Fig. 3

Open in a new tab

Number of lines crossed in 5 min.

VC – Vehicle Control; FX – Fluoxetine; AQ-250 - Aqueous Extract of CD (250 mg/kg); AQ-500 - Aqueous Extract of CD (500 mg/kg); AL-100 - Alcoholic Extract of CD (100 mg/kg).

Values expressed as mean ± SD, ∗p < 0.05 compared to VC, using ANOVA with post-hoc Tukey's test.

3.1.3. Immobility time in forced swim test

The results of immobility time noted for 5 min in all the groups are presented in Fig. 4. There was statistically significant decrease in the immobility time in FX (p < 0.001), AQ-500 group (p < 0.01) and AL-100 (p < 0.01) group compared to the vehicle control. FX group significantly reduced the immobility time compared to all the CD groups: AQ250, AQ-500 and AL-100 (p < 0.001). Comparison of AQ 250 and AQ 500, showed that there was a slight dose dependent trend for reduction in the immobility time although only the AQ-500 showed significantly better response compared to the vehicle group. Also AL-100 group showed more reduction in the immobility time compared to the AQ-500 group (p < 0.01).

Fig. 4.

Fig. 4

Open in a new tab

Immobility Time in 5 min.

VC – Vehicle Control; FX – Fluoxetine; AQ-250 - Aqueous Extract of CD (250 mg/kg); AQ-500 - Aqueous Extract of CD (500 mg/kg); AL-100 - Alcoholic Extract of CD (100 mg/kg).

Values expressed as mean ± SD, ∗∗p < 0.001, ∗p < 0.01 compared to VC, #p < 0.001 compared to AQ-250, AQ-500 & AL-100, $ p < 0.01 compared to AQ-500 using ANOVA with post-hoc Tukey's test.

3.2. Stage 2A: dose finding study

3.2.1. Sleep latency

The values for sleep latency of the mice in all the groups are presented in Fig. 5. There was statistically significant decrease (p < 0.001) in sleep latency in diazepam (DZ) and in AL-500 group compared to the vehicle control group. Between the DZ and AL-500 group, the sleep latency reduced significantly more in the DZ group (p < 0.05). The other three extract groups (AQ-50, AQ-100 and AL-250) did not reduce sleep latency significantly as compared to the vehicle control. Although the sleep latency with AQ-50 and AQ-100 was comparable to the vehicle control; when they were compared to each other, a trend for dose dependent hypnotic effect was observed. Similarly, the higher dose of alcoholic extract (AL-500) reduced the sleep latency significantly (p < 0.05) compared to the lower dose (AL-250). Thus, both the aqueous and alcoholic extracts showed a dose dependent effect on sleep latency.

3.2.2. Total sleeping time

The total sleeping time of the mice in all the groups is depicted in Fig. 6. There was statistically significant increase in the total sleeping time in DZ, AQ-100, AL-250 and AL- 500 groups (p < 0.001) and AQ-50 group (p < 0.01) when each group was compared to the vehicle control. The total sleeping time was maximum in the DZ group as compared to the four CD groups: AQ-50, AQ-100, AL-250 and AL-500 groups (p < 0.001). The two higher dose groups of each extract (AQ-100 and AL-500) showed significantly longer total sleeping time as compared to their respective low dose groups (AQ-50 and AL-250) (p < 0.001) indicating the dose dependent effect. Between the two groups- AL-500 and AQ-100, AL-500 group showed significantly better effect (p < 0.001). We selected the higher doses of each extract i.e. AQ-100 and AL-500 for the phase 2B study to test mechanism of action in combination with picrotoxin.

3.3. Stage 2B: to study mechanism of hypnotic effect of C. deodara

3.3.1. Sleep latency on addition of picrotoxin

Fig. 7 depicts the sleep latency of the mice in all the study groups. As observed in phase 2A, there was statistically significant decrease in sleep latency in DZ (p < 0.001) and AL-500 (p < 0.001) compared to the vehicle control. Also in this experiment, the AQ-100 (p < 0.05) group showed statistically significant reduction in sleep latency as compared to vehicle control which was not observed in phase 2A. Like in Phase 2A, the DZ group showed significantly lower sleep latency compared to all the CD groups (AQ-100, P + AQ-100, AL-500 and P + AL-500 groups) (p < 0.001). Between the two CD groups, the reduction in sleep latency was significantly higher in AL-500 group compared to AQ-100 group (p < 0.01). When combined with picrotoxin (P), the two extract groups also showed significant difference in sleep latency compared to the vehicle control [(P + AL-500) (p < 0.001) and (P + AQ-100) (p < 0.05)]. Moreover, there was no statistically significant difference (p < 0.05) in the sleep latency between AQ-100 group and P + AQ-100 group as well as between AL-500 group and P + AL-500 group.

3.3.2. Total sleeping time on addition of picrotoxin

Total sleeping time of the mice in all the groups is given in Fig. 8. As seen in phase 2A, there was statistically significant increase in total sleeping time in DZ, AQ-100 and AL-500 as compared to the vehicle control. Similar to stage 2A, the DZ group showed significantly higher total sleeping time compared to all the CD groups: AQ-100, P + AQ-100, AL-500 and P + AL-500 groups (p < 0.01). Between the two CD groups, the increase in total sleeping time was significantly higher in AL-500 group compared to AQ-100 group (p < 0.001). When combined with picrotoxin, the combination groups (P + AL-500 and P + AQ-100) showed increased total sleeping time compared to the vehicle control (p < 0.001). Similar to the findings of sleep latency, the total sleep time also did not differ between the AQ-100 vs P + AQ-100 group and between the AL-500 vs P + AL-500 group.

4. Discussion

Extensive literature review on CD revealed that various parts like stem bark, heartwood, needles, roots and cones of CD have been tested in preclinical studies [[40], [41], [42]]. More than 100 phytoconstituents have been extracted and identified from various parts of CD including terpenoids, flavonoids, sterols and essential oils. The phytochemical studies on CD have reported that majority of phytoconstituents are present in the wood and needles of CD [43]. Heartwood and stem bark of CD have been used in experimental animals for evaluating anticancer, analgesic, anti-inflammatory, hypolipidemic, anti-diabetic, gastroprotective and anti-epileptic activities [31,[44], [45], [46]]. In most of the previous experimental studies, the alcoholic extract of either heartwood or stem bark of CD has been used. A number of in vitro studies have explored antimicrobial potential of needles of CD and have demonstrated significant antimicrobial activity of CD against Staphylococcus aureus [41,42]. Many of these herbal product components, including alkaloids, flavonoids, terpenoids, and steroids, have been demonstrated in previous studies to have hypnotic characteristics. Rakhshandeh et al. studied hypnotic effect of hydroalcoholic extract of A. absinthium (which contains terpenoids and flavonoids) in pentobarbital treated mice and found out hypnotic effect possibly due to GABAergic system. GABA receptor type–ionophore complex (GABAA) where the benzodiazepine has a binding site results in increase sleep duration and faster onset of sleep [[47], [48], [49],55].

Majority of the studies on CD have been carried out using its heartwood which requires uprooting of the whole tree. If pine needles are found to be effective for clinical use instead of heartwood or bark, it will prevent cutting of the tree for the same purpose. The pine needles are present abundantly and it is an easily available resource [40,41]. Hence it was decided to use pine needles of CD in the present study. The alcoholic extract (as the parts of CD made into alcoholic extract had shown many benefits in the previous studies) and also the aqueous extract (not explored much till date) of pine needles were chosen in this study.

The authenticated alcoholic and aqueous extracts of CD were received as gift samples from Pharmanza Herbal Pvt. Ltd. The certificate of analysis ensuring quality of each formulation was also received from the manufacturer. The dose chosen of the alcoholic extract for evaluating antidepressant effect in rats was 100 mg/kg. This dose was selected from the previous preclinical study which evaluated 3,4-bis(3,4-dimethoxyphenyl) furan-2,5-dione (BDFD) of heart wood in the experimental model of depression [24] The doses of the aqueous extract in the depression model: 250 and 500 mg/kg were chosen from the experimental study which used the aqueous extract of heartwood of CD in the model of antidiabetic study [31]. Two doses of the aqueous extract used in mice for testing hypnotic effect were 50 and 100 mg/kg. These were derived from the conventional range of dose used in Ayurveda practice (40–80ml/day) for a kwath preparation (decoction of a single herb) [17]. In the hypnosis model, the doses of the alcoholic extract (250 and 500 mg/kg) were chosen based on the dose of the alcoholic extract of CD which was used in the previous experimental study [15]. All the study drugs were administered through Intraperitoneal route in this study based on the preferred route used in the earlier studies on CD. Also, intraperitoneal route was favoured over oral treatment in the interest of preventing the digestive tract and potential biopharmaceutical degradation or alteration [50]. Further studies can be planned using oral route which will be important for achieving clinical relevance.

The FST results showed that although fluoxetine showed the maximum effects, the alcoholic extract group also showed significant decline in immobility time. OFT was conducted to rule out general psychostimulant/increased locomotor effect of CD before subjecting the animals to FST. However, the extracts of CD tested in the present study did not increase the number of lines crossed in the open field test and hence there was no effect of CD on the general locomotor activity. The rats under the influence of an anxiolytic agent tend to spend more time in the centre than at the edges of the OFT apparatus. There was increased time spent in the centre of the open field apparatus by the rats receiving the alcoholic extract compared to the vehicle control indicating its anxiolytic potential. From the results of FST and OFT, it can be inferred that the effect of CD in FST was due to its antidepressant potential and not due to effect on locomotor activity.

Only one study, similar to the present study has been reported in the literature. Kumar et al. showed that 100 mg/kg of 3,4-Bis(3,4-Dimethoxyphenyl) Furan-2,5-Dione derived from heartwood of CD significantly reduced the immobility time in FST compared to control but not in Tail Suspension Test postulating atypical antidepressant like activity of CD. They also found raised noradrenaline and serotonin levels in brain following administration of CD extract [24,[51], [52], [53], [54]]. In our study, unlike in the previous study, two extracts of pine needles of CD were used instead of heartwood in the model of chronic mild stress. Thus, the present study corroborates the evidence for antidepressant potential of CD shown in the earlier study and provides direction for further research on CD.

The hypnotic effect of CD was evaluated in phase 2 in the model of thiopental induced hypnosis. In phase 2A, dose selection was done initially taking two doses each of aqueous (AQ- 50 and AQ-100) and alcoholic (AL-250 and AL-500). Although all four groups increased the total sleeping time, the sleep latency was reduced only in AL-500 group. There was a trend for dose dependent increase in sleeping time and reduction in the sleep latency for the four CD groups. The AL-500 group showed the best effect among all the four CD groups although it was not comparable to diazepam. Since both the aqueous and alcoholic extracts showed a trend towards dose dependent effect with higher doses (AQ-100 and AL-500), these two doses were chosen for the phase 2B study. Higher doses of the two extracts showed better hypnotic effects in phase 2A like that observed with the aqueous extract in the depression model, highlighting the need to explore wider dose ranges of CD in future studies.

The selected higher doses (AQ-100 and AL-500) showed significant hypnotic effect by decreasing sleep latency and increasing the total sleeping time compared to vehicle control. Thus, similar results in phase 2B as that of phase 2A consistently proved the hypnotic potential of CD. Interestingly, AQ 100 also showed significant decrease in sleep latency in stage 2 B, unlike in stage 2A. In this phase the mechanism of hypnotic action was evaluated using picrotoxin as a GABA antagonist. It was found that addition of picrotoxin did not reverse the hypnotic effects caused by either AQ-100 or AL-500. Since picrotoxin is a specific antagonist of GABAA/C, CD may be having hypnotic effect mediated by GABAB receptors and not GABAA or GABAC. It is reported that GABAB receptor modulation also causes hypnosis [56]. The hypnotic effect of CD was tested by Gupta et al. in the past, where the authors had used C. deodara wood essential oil (100,300,1000 mg/kg) in pentobarbitone hypnotic test and all three doses had shown increased sleeping time compared to the control. This study did not test effect of CD on sleep latency and also mechanism of action was not elucidated [25]. The present study confirms the hypnotic effect of CD as reported by Gupta et al. Unlike in the previous study this study showed that CD significantly reduces sleep latency as well as increases total sleeping time compared to vehicle control. For a drug to be an effective hypnotic, it has to reduce the sleep latency and increase the total sleeping time. In our study, we found that CD exhibited both the effects albeit less than that of diazepam. The present study had the following limitations. A limited dose range of extracts was used to study hypnotic and antidepressant effects in the two models. Also only one dose of alcoholic extract was used to evaluate antidepressant effect. To evaluate the hypnotic mechanism of action, we had used picrotoxin which is a specific GABAA/C antagonist. The mechanism of action may be mediated through GABAB receptor subtype which was not evaluated in the present study. Also other than GABA, there are other neurotransmitters involved in the control of sleep and maintaining wakefulness and these Glutamate, Acetylcholine, Histamine, Serotonin and Norepinephrine [57]. Literature shows that CD extract modulates levels of noradrenaline and serotonin in brain [24]. Effect on these other neurotransmitters was not explored in this study.

A range of doses of CD may be tested in future studies and mechanism of action should be explored further.

Despite of the above limitations, the study results are encouraging and provide impetus to future research. The study results revealed that the alcoholic extract of the pine needles of CD showed consistent and dose-dependent antidepressant and hypnotic effects albeit to a lesser extent than the respective standard controls. Zang et al. had found lignans, flavonoids, phenolic compounds, glycosides as the predominant chemical constituents of pine needles of CD which were isolated from petroleum ether, ethyl acetate and n-butanol extracts [58]. Lignan concentrate from the pine needles of CD specifically inhibited the growth of the A 549 cancer cells [59] It would be worthwhile to specifically test different phytoconstituents of pine needles to find out antidepressant and hypnotic activity.

4.1. Limitations

Acute toxicity study was not done as the dose range was lower and it was selected based on earlier studies. An Iranian traditional medicine Portulaca oleracea was tested for its hypnotic effect on Pentobarbital-Induced Sleep in Mice taking its hydroalcoholic extract and its different fractions. The N hexane fraction (n-HF) decreased sleep latency and increased sleep duration and was found to be safe as per single dose acute toxicity test and non-nerutoxic [49]. Unlike, this study, our study did not include acute toxicity and neurotoxicity testing. This is a limitation of our study. Further studies may be done on the formulation and its fractions to address these aspects.

5. Conclusion

Alcoholic and aqueous extracts of pine needles of C. deodara in the doses that were tested in this study, demonstrated antidepressant and hypnotic effects. The alcoholic extract showed better and more consistent antidepressant and hypnotic effect compared to the aqueous extract.

Source of funding

The authors gratefully acknowledge the financial support of Diamond Jubilee Society Trust (DJST), Seth GSMC and KEM Hospital, Mumbai for conducting this study. This is institutional funding that was granted for this study.

Author contribution

The authors confirm contribution to the paper as follows: Study conception and design: Nirmal Kumar E, Padmaja A. Marathe, Sandhya K. Kamat; Data collection: Nirmal Kumar E, Pritika Mal; Analysis and interpretation of results: Nirmal Kumar E, Padmaja A. Marathe, Harshitha Havaldar, Merin Eldhose; Draft manuscript preparation: Nirmal Kumar E, Padmaja A. Marathe, Sandhya K. Kamat. All authors reviewed the results and approved the final version of the manuscript.

Conflict of interest

The authors declare that there was no commercial or financial association that can be considered as conflict of interest when the research was planned and conducted.

Acknowledgement

The authors thank Pharmanza Herbal Private Limited, Anand, Gujarat for the gift sample of Cedrus deodara Aqueous and Alcoholic extract.

Footnotes

Peer review under responsibility of Transdisciplinary University, Bangalore.

Contributor Information

Nirmal Kumar E, Email: nirmal25ega@gmail.com.

Padmaja A. Marathe, Email: pam2671@gmail.com.

References

  • 1.https://www.who.int/news-room/fact-sheets/detail/depression Depression [Internet]. [cited 2021 Nov 13]. Available from:
  • 2.Murthy R.S. National mental Health Survey of India 2015–2016. Indian J Psychiatr. 2017 Jan 1;59(1):21–26. doi: 10.4103/psychiatry.IndianJPsychiatry_102_17. pmc/articles/PMC5419008/ [Internet] [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 3.Antoniuk S., Bijata M., Ponimaskin E., Wlodarczyk J. Chronic unpredictable mild stress for modeling depression in rodents: meta-analysis of model reliability. Neurosci Biobehav Rev. 2019 Apr 1;99:101–116. doi: 10.1016/j.neubiorev.2018.12.002. https://pubmed.ncbi.nlm.nih.gov/30529362/ [Internet] [DOI] [PubMed] [Google Scholar]
  • 4.Lader M. Limitations of current medical treatments for depression: disturbed circadian rhythms as a possible therapeutic target. Eur Neuropsychopharmacol. 2007 Dec;17(12):743–755. doi: 10.1016/j.euroneuro.2007.05.004. https://pubmed.ncbi.nlm.nih.gov/17624740/ Available from: [DOI] [PubMed] [Google Scholar]
  • 5.Ohayon M.M. Epidemiological Overview of sleep Disorders in the General Population. Sleep Med Res. 2011 Apr 30;2(1):1–9. https://www.researchgate.net/publication/285076723_Epidemiological_Overview_of_sleep_Disorders_in_the_General_Population Available from: [Google Scholar]
  • 6.Sateia M.J., Buysse D.J., Krystal A.D., Neubauer D.N., Heald J.L. Clinical practice guideline for the pharmacologic treatment of chronic insomnia in Adults: an American Academy of sleep medicine clinical practice guideline. J Clin Sleep Med. 2017;13(2):307–349. doi: 10.5664/jcsm.6470. https://pubmed.ncbi.nlm.nih.gov/27998379/ Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 7.Wagner J., Wagner M.L. Non-benzodiazepines for the treatment of insomnia. Sleep Med Rev. 2000;4(6):551–581. doi: 10.1053/smrv.2000.0126. https://pubmed.ncbi.nlm.nih.gov/12531036/ Available from: [DOI] [PubMed] [Google Scholar]
  • 8.Hypnotics and Sedatives | Goodman & Gilman’s: The pharmacological basis of therapeutics, 13e | AccessMedicine | McGraw Hill Medical [Internet]. [cited 2022 Jan 12]. Available from: https://accessmedicine.mhmedical.com/content.aspx?bookid=2189&sectionid=169518968.
  • 9.Jindal R.D., Thase M.E. Treatment of insomnia associated with clinical depression. Sleep Med Rev. 2004;8(1):19–30. doi: 10.1016/S1087-0792(03)00025-X. https://pubmed.ncbi.nlm.nih.gov/15062208/ Available from: [DOI] [PubMed] [Google Scholar]
  • 10.Nutt D.J., Wilson S., Paterson L. Sleep disorders as core symptoms of depression. Dialog Clin Neurosci. 2008;10(3):329. doi: 10.31887/DCNS.2008.10.3/dnutt. pmc/articles/PMC3181883/ [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 11.Sanacora G, Mason GF, Rothman DL, Behar KL, Hyder F, Petroff OA, et al. Reduced cortical gamma-aminobutyric acid levels in depressed patients determined by proton magnetic resonance spectroscopy. Arch Gen Psychiatr 1999 Nov;56(11):1043–7. 10.1001/archpsyc.56.11.1043. [DOI] [PubMed]
  • 12.Caldji C., Francis D., Sharma S., Plotsky P.M., Meaney M.J. The effects of early rearing environment on the development of GABAA and central benzodiazepine receptor levels and novelty-induced fearfulness in the rat. Neuropsychopharmacology. 2000;22:3. doi: 10.1016/S0893-133X(99)00110-4. https://www.nature.com/articles/1395434 [Internet]. 2000 Mar [cited 2022 Jan 14];22(3):219–29. Available from: [DOI] [PubMed] [Google Scholar]
  • 13.Luscher B., Shen Q., Sahir N. The GABAergic deficit hypothesis of major depressive disorder. Mol Psychiatr. 2011 Apr;16(4):383–406. doi: 10.1038/mp.2010.120. https://pubmed.ncbi.nlm.nih.gov/21079608/ Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 14.Mukherjee P.K. Exploring botanicals in Indian system of medicine—regulatory perspectives. Ceased. 2003;20(3):249–264. https://www.tandfonline.com/doi/abs/10.1081/CRP-120023840 Available from. [Google Scholar]
  • 15.Chaudhary A.K., Ahmad S., Mazumder A. Cedrus deodara (roxb.) loud.: a review on its ethnobotany, phytochemical and pharmacological profile. Phcog J. 2011 Jul 1;3(23):12–17. [Google Scholar]
  • 16.Dhayabaran D., Florance E.J., Nandakumar K., Puratchikody A. Anxiolytic and anticonvulsant activity of alcoholic extract of heart wood of Cedrus deodara roxb. in rodents. J Med Plants Res. 2010;4(14):1374–1381. http://www.academicjournals.org/JMPR Available from: [Google Scholar]
  • 17.Singh P., Khosa R., Mishra G. Evaluation of antidiabetic activity of ethanolic extract of Cedrus deodara (Pinaceae) stem bark in streptozotocin induced diabetes in mice. Niger J Exp Clin Biosci. 2013;1:33–38. Available from: https://www.researchgate.net/publication/264862394_Evaluation_of_antidiabetic_activity_of_ethanolic_extract_of_Cedrus_deodara_Pinaceae_stem_bark_in_streptozotocin_induced_diabetes_in_mice. [Google Scholar]
  • 18.Parveen R., Azmi M.A., Tariq R.M., Mahmood S.M., Hijazi M., Mahmud S., et al. Determination of antifungal activity of cedrus deodara root oil and its compounds against CANDIDA albicans and aspergillus fumigatus. Pakistan J Bot. 2010;42(5):3645–3649. [Google Scholar]
  • 19.Wu Y., Bai J., Liu X., Liu L., Zhong K., Huang Y., et al. Antibacterial effect of 3-p-trans-coumaroyl-2-hydroxyquinic acid, a phenolic compound from needles of Cedrus deodara, on cellular functions of Staphylococcus aureus. RSC Adv. 2018 Jan 24;8(9):4969–4975. doi: 10.1039/c7ra13457f. https://pubs.rsc.org/en/content/articlehtml/2018/ra/c7ra13457f Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 20.Xu F., Gu D., Wang M., Zhu L., Chu T., Cui Y., et al. Screening of the potential α-amylase inhibitor in essential oil from Cedrus deodara cones. Ind Crop Prod. 2017 Sep 1;103:251–256. [Google Scholar]
  • 21.Shi X., Liu D., Zhang J., Hu P., Shen W., Fan B., et al. Extraction and purification of total flavonoids from pine needles of Cedrus deodara contribute to anti-tumor in vitro. BMC Compl Alternative Med. 2016 Dec;16(1):1–9. doi: 10.1186/s12906-016-1249-z. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 22.Mashaal K., Perveen R., Zia Z., Mahmood S.M., Minhas K., Ibrahim S., et al. Histopathological effects on stomach, liver and kidney associated with Cedrus deodara root oil in ulcer induced rats (Wistar strain) Pak J Pharm Sci. 2020 Nov 1;(6):33. [PubMed] [Google Scholar]
  • 23.Viswanatha G.L., Nandakumar K., Shylaja H., Ramesh C., Rajesh S., Srinath R. Anxiolytic and anticonvulsant activity of alcoholic extract of heart wood of Cedrus deodara roxb in rodents. Asian J Pharmaceut Res Health Care. 2009;1(2):217–239. https://www.informaticsjournals.com/index.php/ajprhc/article/view/630 Available from: [Google Scholar]
  • 24.Kumar N., Dhayabaran D., Nampoothiri M., Nandakumar K., Puratchikody A., Lalani N., et al. Atypical antidepressant activity of 3,4-bis(3,4-dimethoxyphenyl) furan-2,5-dione isolated from heart wood of Cedrus deodara, in rodents. Korean J Physiol Pharmacol. 2014;18(5):365. doi: 10.4196/kjpp.2014.18.5.365. pmc/articles/PMC4211118/ Available from. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 25.Tandan SK, Chandra S, Gupta S, Lal J. Pharmacodynamic effects of Cedrus deodara wood essential oil. Indian J Pharmaceutic Sci [Internet]. [cited 2022 Jan 21];60(1):20. Available from: https://www.ijpsonline.com/articles/.
  • 26.Liu Y.N., Peng Y.L., Lei-Liu, Wu T.Y., Zhang Y., Lian Y.J., et al. TNFα mediates stress-induced depression by upregulating indoleamine 2,3-dioxygenase in a mouse model of unpredictable chronic mild stress. European cytokine network. 2015;26(1):15–25. doi: 10.1684/ecn.2015.0362. https://pubmed.ncbi.nlm.nih.gov/26083579/ Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 27.Willner P., Towell A., Sampson D., Sophokleous S., Muscat R. Reduction of sucrose preference by chronic unpredictable mild stress, and its restoration by a tricyclic antidepressant. Psychopharmacology. 1987;93:3. doi: 10.1007/BF00187257. https://link.springer.com/article/10.1007/BF00187257 [Internet]. 1987 Nov [cited 2022 Jan 24];93(3):358–64. Available from: [DOI] [PubMed] [Google Scholar]
  • 28.Sharma AS, Joshi SS. Experimental study to evaluate the effect of Niacin in chronic unpredictable stress induced depression in Wistar rats.[dissertation for MD in Pharmacology discipline in 2021; submitted to Maharashtra University of Health Sciences, Nashik, India [ unpublished) ]].
  • 29.Pawluski J.L., van Donkelaar E., Abrams Z., Houbart V., Fillet M., Steinbusch H.W.M., et al. Fluoxetine dose and administration method differentially affect hippocampal plasticity in adult female rats. Neural Plast. 2014;2014 doi: 10.1155/2014/123026. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 30.Farhan M., Haleem D.J. Anxiolytic profile of fluoxetine as monitored following repeated administration in animal rat model of chronic mild stress. Saudi Pharmaceutic J. 2016 Sep 1;24(5):571. doi: 10.1016/j.jsps.2015.03.006. pmc/articles/PMC5059824/ Available from. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 31.Jain S., Jain A., Malviya N., Kumar D., Jain V., Jain S. Antidiabetic activity of Cedrus deodara aqueous extract and its relationship with its Antioxidant properties. J Pharm Sci Pharmacol. 2015 Jan 9;1(3):187–194. [Google Scholar]
  • 32.Tamburella A., Leggio G.M., Micale V., Navarria A., Bucolo C., Cicirata V., et al. Behavioural and neurochemical changes induced by stress-related conditions are counteracted by the neurokinin-2 receptor antagonist saredutant. Int J Neuropsychopharmacol. 2013 May;16(4):813–823. doi: 10.1017/S1461145712000612. https://pubmed.ncbi.nlm.nih.gov/22695046/ Available from: [DOI] [PubMed] [Google Scholar]
  • 33.Gould T.D., Dao D.T., Kovacsics C.E. Mood and anxiety related phenotypes in mice. Neuromethods. 2009;42:1–20. http://www.springerprotocols.com/Abstract/doi/10.1007/978-1-60761-303-9_1 Available from: [Google Scholar]
  • 34.Walsh R.N., Cummins R.A. The open-field test: a critical review. Psychological Bulletin. 1976 May;83(3):482–504. doi: 10.1037/2F0033-2909.83.3.482. [DOI] [PubMed] [Google Scholar]
  • 35.Vogel H.G., Maas J., Hock F.J., Mayer D. Drug Discovery and evaluation: safety and phrmacokinetic assays. 2nd ed. 2013 Jan 1. pp. 1–1402. [Google Scholar]
  • 36.Mirshafa S.A., Azadbakht M., Ahangar N. Study of antidepressant and sedative-hypnotic activity of hydroalcoholic extract of asperugo procumbens L. Aerial parts in mice. Iran J Pharmaceutical Research: IJPR. 2013;12(3):529. pmc/articles/PMC3813279/ Available from. [PMC free article] [PubMed] [Google Scholar]
  • 37.Shanmugasundaram J., Subramanian V., Nadipelly J.S., Kathirvelu P., Sayeli V., Cheriyan B.V. Sedative-hypnotic like effect of 5-methoxyflavone in mice and investigation on possible mechanisms by in vivo and in silico methods. Biomedicine & pharmacotherapy = Biomedecine & pharmacotherapie. 2018 Dec 1;108:85–94. doi: 10.1016/j.biopha.2018.08.117. https://pubmed.ncbi.nlm.nih.gov/30218862/ [DOI] [PubMed] [Google Scholar]
  • 38.Alnamer R., Alaoui K., Bouidida E.H., Benjouad A., Cherrah Y. Sedative and hypnotic activities of the methanolic and aqueous extracts of lavandula officinalis from Morocco. Adv Pharmacol Sci. 2012;2012 doi: 10.1155/2012/270824. https://pubmed.ncbi.nlm.nih.gov/22162677/ Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 39.Razack S., Kandikattu H.K., Venuprasad M.P., Amruta N., Khanum F., Chuttani K., et al. Anxiolytic actions of Nardostachys jatamansi via GABA benzodiazepine channel complex mechanism and its biodistribution studies. Metabolic Brain Dis. 2018 Oct 1;33(5):1533–1549. doi: 10.1007/s11011-018-0261-z. https://pubmed.ncbi.nlm.nih.gov/29934858/ Available from: [DOI] [PubMed] [Google Scholar]
  • 40.Bisht A., Jain S., Misra A., Dwivedi J., Paliwal S., Sharma S. Cedrus deodara (Roxb. ex D.Don) G.Don: a review of traditional use, phytochemical composition and pharmacology. J Ethnopharmacol. 2021 Oct 28;279 doi: 10.1016/j.jep.2021.114361. [DOI] [PubMed] [Google Scholar]
  • 41.Wu Y., Bai J., Zhong K., Huang Y., Qi H., Jiang Y., et al. Antibacterial activity and membrane-disruptive mechanism of 3-p-trans-Coumaroyl-2-hydroxyquinic acid, a novel phenolic compound from pine needles of Cedrus deodara, against Staphylococcus aureus. Molecules (Basel, Switzerland) 2016 Aug 1;21(8) doi: 10.3390/molecules21081084. https://pubmed.ncbi.nlm.nih.gov/27548123/ [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 42.Bai J., Wu Y., Liu X., Zhong K., Huang Y., Gao H. Antibacterial activity of shikimic acid from pine needles of Cedrus deodara against Staphylococcus aureus through damage to cell membrane. Int J Mol Sci. 2015;16:27145–27155. doi: 10.3390/ijms161126015. https://www.mdpi.com/1422-0067/16/11/26015/htm [Internet]. 2015 Nov 13 [cited 2022 Jan 21];16(11):27145–55. Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 43.Saab A.M., Gambari R., Sacchetti G., Guerrini A., Lampronti I., Tacchini M., et al. Phytochemical and pharmacological properties of essential oils from Cedrus species. Natural Prod Res. 2018 Jul 2;32(12):1415–1427. doi: 10.1080/14786419.2017.1346648. https://pubmed.ncbi.nlm.nih.gov/28670915/ Available from: [DOI] [PubMed] [Google Scholar]
  • 44.Manne SH, Basavaraju BB, Bindhu V, Keerthi S, Vernekar N, Laturkar V. Study of anti inflammatory and analgesic activity of methanolic extracts of Cedrus deodara. BioTechnol Indian J [Internet]. [cited 2022 Jan 20];2(1). Available from: https://www.tsijournals.com/abstract/study-of-anti-inflammatory-and-analgesic-activity-of-methanolic-extracts-of-cedrus-deodara-1177.html.
  • 45.Sharma A., Arora P. Anti-cancer activity of Cedrus deodara in 1,2- dimethly hydrazine (DMH) induced anti cancer model in rats. Asian J Pharmaceut Res Dev. 2018 Jun 20;6(2):82–86. https://ajprd.com/index.php/journal/article/view/348 Available from: [Google Scholar]
  • 46.Patil S, Prakash T, Kotresha D, Rao NR, Pandy N. Antihyperlipidemic potential of Cedrus deodara extracts in monosodium glutamate induced obesity in neonatal rats. Indian J Pharmacol. 2011;43(6):644–647. [PMC free article] [PubMed]. [DOI] [PMC free article] [PubMed]
  • 47.Hosseini A., Forouzanfar F., Rakhshandeh H. Hypnotic effect of nepeta glomerulosa on phenobarbital-induced sleep in mice. Jundishapur J Nat Pharm Prod. 2016 [Google Scholar]
  • 48.Rakhshandeh H., Heidari A., Pourbagher-Shahri A.M., Rashidi R., Forouzanfar F. Hypnotic effect of A. Absinthium hydroalcoholic extract in pentobarbital-treated mice. Neurol Res Int. 2021 doi: 10.1155/2021/5521019. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 49.Hamedi S., Forouzanfar F., Rakhshandeh H., Arian A. Hypnotic effect of Portulaca oleracea on pentobarbital-induced sleep in mice. Curr Drug Discov Technol. 2019;16(2):198–203. doi: 10.2174/1570163815666180308142543. [DOI] [PubMed] [Google Scholar]
  • 50.Al Shoyaib A., Archie S.R., Karamyan V.T. Intraperitoneal route of drug administration: should it be used in experimental animal studies? Pharmaceut Res. 2020 Ja;37:1–7. doi: 10.1007/s11095-019-2745-x. [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 51.Katz R.J., Roth K.A., Carroll B.J. Acute and chronic stress effects on open field activity in the rat: implications for a model of depression. Neurosci Biobehav Rev. 1981 Jun 1;5(2):247–251. doi: 10.1016/0149-7634(81)90005-1. [DOI] [PubMed] [Google Scholar]
  • 52.Cryan J.F., Markou A., Lucki I. Assessing antidepressant activity in rodents: recent developments and future needs. Trends Pharmacol Sci. 2002 May 1;23(5):238–245. doi: 10.1016/s0165-6147(02)02017-5. https://pubmed.ncbi.nlm.nih.gov/12008002/ [DOI] [PubMed] [Google Scholar]
  • 53.Askew B.M. A simple screening procedure for imipramine-like antidepressant agents. Life Sci. 1962;10(10):725–730. doi: 10.1016/0024-3205(63)90076-6. https://pubmed.ncbi.nlm.nih.gov/14077412 Available from: [DOI] [PubMed] [Google Scholar]
  • 54.Porsolt R.D., Anton G., Blavet N., Jalfre M. Behavioural despair in rats: a new model sensitive to antidepressant treatments. Eur J Pharmacol. 1978 Feb 15;47(4):379–391. doi: 10.1016/0014-2999(78)90118-8. [DOI] [PubMed] [Google Scholar]
  • 55.Li N., Liu J., Wang M., Yu Z., Zhu K., Gao J., et al. Sedative and hypnotic effects of Schisandrin B through increasing GABA/Glu ratio and upregulating the expression of GABAA in mice and rats. Biomed Pharmacother. 2018 Jul 1;103:509–516. doi: 10.1016/j.biopha.2018.04.017. [DOI] [PubMed] [Google Scholar]
  • 56.Kantrowitz J., Citrome L., Javitt D. GABA(B) receptors, schizophrenia and sleep dysfunction: a review of the relationship and its potential clinical and therapeutic implications. CNS Drugs. 2009;23(8):681–691. doi: 10.2165/00023210-200923080-00005. https://pubmed.ncbi.nlm.nih.gov/19594197/ Available from: [DOI] [PMC free article] [PubMed] [Google Scholar]
  • 57.Kayabekir M. Neurophysiology of basic molecules affecting sleep and wakefulness mechanisms, fundamentals of sleep pharmacology. Sleep medicine and the evolution of contemporary sleep pharmacotherapy. 2022 Jan 7 doi: 10.5772/intechopen.100166. [DOI] [Google Scholar]
  • 58.Zhang J.M., Shi X.F., Ma Q.H., He F.J., Wang D.D., Liu D.Y., et al. [studies on the chemical constituents from pine needles of Cedrus deodara (II)] Zhong Yao Cai. 2010;33(7):1084–1086. [PubMed] [Google Scholar]
  • 59.Shi X., Du R., Zhang J., Lei Y., Guo H. Evaluation of the anti-cancer potential of Cedrus deodara total lignans by inducing apoptosis of A549 cells. BMC Compl Alternative Med. 2019;19(1):281. doi: 10.1186/s12906-019-2682-6. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Ayurveda and Integrative Medicine are provided here courtesy of Elsevier

RESOURCES