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Neuroprotective and anti-inflammatory effect of Hedyotis corymbosa extract on chronic stress-induced depression model of rat-A in vivo and ex vivo study

Background: Depression is a well-known mood disorder characterized by persistently low mood and loss of interest and a feeling of sadness. Plants and plant-derived agents have recently attracted the interest of researchers for their therapeutic effects against several illnesses, including mental disorders, and several herbal plants and herbal formulations are useful against experimental depression. In this study, the neuroprotective effects of Hedyotis corymbosa extract were investigated in rats induced with chronic mild stress. Methods: Animals were designated into the following groups: control, 0, 150, and 300 mg of extracts. The dose was given for 30 consecutive days via the oral route. Sucrose preference analysis, forced swim, and open field tests were performed, and serum cortisol and monoamine levels in brain tissue were determined. Expression of brain-derived neurotrophic factor (BDNF) was also examined. Results: Supplementation with extract increased the sucrose preference ratio, locomotor activity, and monoamines and decreased serum cortisol levels. The protein and mRNA expression of BDNF in the brain tissue was decreased by 62.58% and 73% in control rats. However, supplementation with extracts significantly increased BDNF mRNA expression (by 107% and 229.6% in groups 150 mg and 300 mg, respectively). Similarly, the protein expression of BDNF increased by 82.3% and 141.2% in groups 150 mg and 300 mg, respectively. Conclusion: In summary, experimental results suggest that supplementation with extracts of Hedyotis corymbosa may be effective against depression.


Background
Depression is a state of aversion to activity and low mood. Depression affects behavior, thoughts, feelings, sense of well-being, and tendencies. It is a chronic, recurring, and (15 minutes), one period of exposure to empty bottle (1 h), one period of continuous light (36 h), one period of paired caging (2 h), one period of wet cage (21 h) and one period of water and food deprivation. The procedure was repeated for 28 days.

Groups and treatments
Rats were designated into four groups: control, 0 mg, 150 mg, and 300 mg of extracts.
The dose was given for 30 consecutive days via the oral route. Each group contained six rats.

Sucrose preference analysis
The sucrose preference analysis was carried out according to Willner et al. (1987). Briefly, the two bottles of sucrose solution (1%) were placed on the rat cage separately. Then, free access was provided to the rats to drink water from these cages for one day. Then, one bottle was continued with the same sucrose solution and another bottle was filled with water for the next 24 h. Bottles position was changed to avoid the influence of bottle position. Then, all the animals were deprived of water for another 23 h, and sucrose preference examination was performed for each rat. Then, the amount of water and sucrose solution consumed was recorded and calculated.

Behavioral test
Forced swim and open field tests were carried out as previously described (Jun et al. 2016). All the rats were placed in the center of open field [square chamber (80 cm), high walls (40 cm) and light (80 lux)] for 180 seconds in a silent room following rat weighed.
Times of rearing and number of crossing squares were recorded.

Determination of serum cortisol
The serum cortisol level was estimated using an enzyme-linked immunosorbent assay (Anil et al. 2018). At the end of the treatment, rats were dissected and blood was collected and processed with the serum for the determination of cortisol level.

Results
The study evaluated the effect of extracts of H. corymbosa on chronic mild stress-induced depression. The quantitative analysis H. corymbosa extract revealed the presence of saponins, flavonoids, carbohydrates, proteins, steroids, tannins, and phenolic compounds ( Figure 1, Table 1). Figure 2 shows the sucrose preference ratio of control and treated rats. The sucrose preference ratio was substantially reduced by 50.5% in 0 mg rats compared to control rats. However, extracts increased the sucrose preference ratio to 74.4% and 93.6% in groups 150 mg and 300 mg, respectively (Figure 2, P < 0.034).
Behavioral parameters, such as rearing, crossing, and immobility time, were determined in control and treated rats. Rearing capacity was substantially reduced by 69.6% in 0 mg rats. However, extracts treatment increased rearing by 83.5% and 187% in groups 150 mg and 300 mg, respectively (Figure 3, P < 0.041). Crossing counts were substantially reduced by 77.3% in 0 mg rats. Extract supplementation increased crossing counts by 166% and 297.2% in groups 150 mg and 300 mg, respectively (Figure 4, P < 0.025).
Immobility time was substantially increased by 125.9% in control rats. However, extracts treatment reduced immobility time by 32.8% and 47.5% in groups 150 mg and 300 mg, respectively (Figure 5, P < 0.05).
The serum cortisol level was substantially increased by 147.3% in 0 mg rats. However, supplementation with extracts reduced the cortisol level by 27.2% and 51.4% in groups 150 mg and 300 mg, respectively ( Figure 6, P < 0.044). The level of monoamines (e.g., 5-HT, noradrenaline, and 5-HIAA) was substantially reduced in brain tissue homogenate.
However, supplementation with extracts significantly increased these monoamine levels to near-normal levels ( Table 3, P < 0.032). In brain tissue, the protein and mRNA expression of BDNF was substantially reduced by 62.58% and 73% in 0 mg rats respectively. However, supplementation with extracts significantly increased protein and mRNA expression BDNF by 107% and 229.6% in groups 150 mg and 300 mg, respectively ( Figure   7, P < 0.041). Similarly, BDNF protein expression was increased by 82.3% and 141.2% in groups 150 mg and 300 mg, respectively (Figure 8, P < 0.043).

Discussion
In this study, we have investigated the biochemical, behavioral and molecular approaches to understand the effect of extracts on chronic mild stress induced depression model of rats. We observed that the extracts treatment exhibited protective effect against depression and prevented the hormone dysregulation. Depression induced rats showed

Conclusion
In this study, BDNF expression was drastically reduced in depression induced rat model, and return to the near normal range following extracts treatment, which confirms the protective effect against depression. Taking all these data together, it is suggested that the extract is a good therapeutic agent against chronic stress induced depression model of rats.          Protective effect of extracts on the protein expression of BDNF in an experimental model of chronic mild stress-induced depression.