3.1 Effect on monoamine neurotransmitters

In the past years, monoamine neurotransmitters have become a focus on the pathophysiology of depression. It is reported that depression is caused by a decline in the function of monoamine transmitters, such as serotonin (5-HT), noradrenaline (NA) and dopamine (DA). Indeed, reduction of plasma 5-HT level has been observed in depressed patients resulting from alcohol withdrawal.22 At present, antidepressants based on monoamine transmitters are still the first-line antidepressants. Through the metabolomics in serum, urine and cerebral tissue of chronic unpredictable mild stress (CUMS) rats, ginsenosides were found to mitigate changes in amino acid neurotransmitters and monoamine neurotransmitters.23 Similarly, pre-treatment with ginseng fruit saponins could significantly increase the 5-HT levels in both serum and platelets but not reverse the decline of 5-HT level in the brain induced by an experimental comorbidity model of myocardial infarction (Akbay's model) and depression (modified forced swimming model). Besides, ginseng fruit saponins could decrease the 5-HT_2AR expression in both brain and platelets.24, 25 Ginsenoside Rb1, active component of the ginseng root, significantly upregulated the 5-HT, 5-HIAA, NA and DA levels in the brain of CUMS rats and also exhibited a synergistic effect when combined with caffeine and fluoxetine.26 Besides, ginsenoside Rb1 and its metabolite compound K acted as antidepressants by regulating 5-HT_2A receptors, which was demonstrated by blockade action of ritanserin pre-treatment.27 Gintonin, an active constituent of the Panax ginseng extract, could improve depression-related symptoms in mice with alcohol withdrawal through elevating the level of plasma 5-HT which was released from intestinal enterochromaffin cells.28 Chronic treatment of total saponins extracted from Panax notoginseng also could improve the rats depression-like behaviours in chronic mild stress (CMS) mice via increasing the levels of 5-HT, DA and NA and decreasing cytoplasmic free intracellular Ca²⁺ concentration.29 Other neurotransmitters, such as acetylcholine and glutamate, also have been found to be strongly associated with depression.

3.2 Effect on hypothalamic-pituitary-adrenal axis

The neuroendocrine hypothesis points out that dysfunction of the endocrine system is the main cause of the occurrence of depression. Hyperactivity of the hypothalamic-pituitary-adrenal (HPA) axis and the consequently potentiated stress response have many adverse impacts on the brain and many other peripheral organs.30, 31 It has been demonstrated that hyperactivity of the HPA axis leads to the depressive behaviours mainly by elevating blood cortisol (CORT) levels and facilitating brain corticotropin-releasing factor transmission.32 Hyperactivity of stress hormones can cause atrophy or apoptosis of neurons and decrease nerve regeneration in the hippocampus, amygdala and other depression-related brain regions. There is growing evidence that neuroendocrine functional disorder might be the physiological basis of pathological changes of depression and other mood disorders.33 Researches over the last couple of years indicated that Rg1 could return the HPA axis to normal function, by improving serum CORT as well as testosterone levels and modulating glucocorticoid receptor (GR) levels in both prefrontal cortex (PFC) and hippocampus in rats with the CUMS-induced depression.34 Ginsenoside Rh1 and Rg3 as oestrogen receptor (ERs) ligands and Rh2 and K as GRs ligands may also affect the function of HPA axis mediated by ERs or GRs.35 Furthermore, Rb1 and Rg3 could attenuate neuronal toxicity, which is related to glucocorticoid activity.36 In a study of CORT-induced depression in mice, researchers found that ginseng total saponins (GTS) treatments improved depressant behaviours without changing the heightened serum CORT levels, as well as increased the downregulated levels of inhibitory GSK-3β phosphorylation in the hippocampus, which was opposite to fluoxetine.37 In addition, studies have shown that some disorders of the gonadal hormones, such as oestrogen and progesterone, might affect feelings, emotions, and cognition which involved in the development of depression. There is strong evidence that menopause is a time of increased vulnerability to dysphoric mood.38 Moreover, in depressed patients, the HPG axis that regulates gonadal function was weakened.39 Rg1 showed antidepressant-like effects not only by regulating the function of the HPA axis, but also the HPG axis. As is known, the level of androgen determines the androgen receptor (AR) expression, and Rg1 possessed an androgen-like effect. It alleviated sleep disruption, decreased the CORT level in serum and increased the androgen receptor (AR) protein content without affecting testosterone level of the PFC of gonadectomized mice.34

3.3 Effect on BDNF

Brain-derived neurotrophic factor (BDNF), one of the neurotrophic factors, does main favour in neurogenesis, neural survival and growth.40, 41 BDNF binds to its tropomyosin receptor kinase B (TrkB) to activate downstream signalling pathways and then completes phosphorylation and activation of the cAMP response element-binding protein (CREB).42 The BDNF reduction in both hippocampus and PFC is closely associated with depression. Besides, the serum BDNF level also decreases in depression patients.43, 44 BDNF has acted as a biomarker for depression in many studies.45-47 The previous study illustrated that the extracts from Panax ginseng produce antidepressant-like effect via accommodation of BDNF expression in various classic animal depression models, including FST, tail suspension test (TST), CMS, CUMS and so on.48-50 It has been reported that ginseng extract G115 could notably improve BDNF expression in the hippocampus and PFC of ethanol-induced depression mice, even superior to amitriptyline, indicating that the antidepressant effects may contribute to some BDNF-related signalling pathway.51 Interestingly, G115 did not influence the BDNF levels in normal brain areas. GTS could reverse alterations in depressive behaviours resulted from CORT through intervening in GSK-3 β-CREB signalling pathway in the hippocampus and reversing the reduction of some proteins related to plasticity.52 In the lipopolysaccharide (LPS) immunoreactive mice model which act as an inflammatory-related animal model of depression, levels of BDNF and TrkB in the hippocampus were upregulated by sesquiterpenoids, suggesting that BDNF/TrkB pathway may be a target of the antidepressant effects of sesquiterpenoids.53 In addition, ginseng pectin also showed the antidepressant effect to some extent,54 which may be related to its neuroprotective role through the activation of ERK/MAPK and Akt survival signalling pathways.55

Ginsenoside monomers could similarly lead to apparent physiological behaviour changes in depressed murine. Ginsenoside Rg1 enhanced hippocampal BDNF signalling pathway while reduced serum CORT level, and it also reversed the decline in both dendritic spine density and hippocampal neurogenesis in the CMS model.56 Moreover, Rg1 could increase CREB phosphorylation and BDNF expression in the amygdala of CUMS rats.57 Downregulation of the Akt/mTOR signalling pathway has been also shown to be associated with decreased senescence of neural stem cells induced by ginsenoside Rg1.58 However, Rg1 seemed to have no distinguishable effect on the monoaminergic system.56 The same as sesquiterpenoids, ginsenoside Rg2 and Rg5 regulated the BDNF/TrkB pathway, which was confirmed by the blocking effect of TrkB shRNA and TrkB inhibitor.59, 60 Beyond this, researches showed that infusion of the BDNF signalling inhibitor blocked the antidepressant effects of Rg3.61

3.4 Effect on the neuroimmune system

Increasing evidence has indicated that inflammatory cytokine disorders may be closely related to the pathogenesis of depression. Some patients with MDD showed the main characteristics of inflammatory cytokine disorders in peripheral blood and cerebrospinal fluid. LPS is an effective and potent inducer of inflammatory cytokines (eg, TNF-α; IL-1β; IL-6) that can acutely activate the innate immune system in the peripheral or central nervous system to induce depression-like behaviours. Therefore, the LPS-induced depression animal model was used to study the relationship between the immune system and depression. Accordingly, anti-inflammatory therapy is becoming an effective treatment for depression and other neurological diseases. The antidepressant role of GTS is basically related to its anti-inflammatory actions. It was found that GTS treatment can alleviate depression-like behaviour impairments and decrease mRNA levels of IL-1b, IL-6, TNF-a and IDO in the hippocampus. Reduced generation of various proinflammatory cytokines was detected in both LPS-challenged mice and RAW264.7 cells after GTS treatment.62 Panax ginseng extract was found to reduce serum ACTH and CORT concentration, enhance Nrf2 and HO-1 activities and inhibit inflammatory activity in the amygdala of chronic-restraint stress (CRS)-induced depressive mice, which resulted in enhanced BDNF activity and ultimately improved depressant-like behaviours.63 Although ginsenoside Rg1, a cognized anti-inflammatory agent, could not act directly in the brain, it was able to effectively mitigate the neuroimmune disturbances and neurochemical disturbances induced by central LPS challenge.64 Rg1 could significantly inhibit the expression of NADPH oxidase in frontal cortex and hippocampus of CRS-induced cognitive impairment mice, thereby reducing the production of reactive oxygen species and reducing the oxidative damage of neurons in frontal cortex and hippocampal CA1 region.65 The inhibition of Nod-like receptor pyrin domain-containing protein 1 inflammasomes was also involved in the mechanisms underlying the effect of Rg1 on the glucocorticoid-induced neuronal injury.66 In addition, ginsenoside Rg1 plays an important role in activating PPAR signalling to prevent apoptosis and inflammation.67, 68 Ginsenoside Rg3 improved depression-like behaviours induced by systemic inflammatory via suppressing neuroinflammation and modulating TRP-KYN metabolism.69 All these certified that targeting the peripheral immune system may work as a new therapy to neuropsychiatric disorders caused by neuroinflammation.

Microglia, acting as cellular effectors of innate immunity, along with astrocytes, can drive the process of neuroinflammation by phagocytosis and cytokine release.70 An increasing body of evidence has validated the relationship between glial cells and neurological diseases. GTS was able to reverse the decrease of astrocyte number, structural changes and hippocampal atrophy in CORT-induced depressed mice.71 Many studies have shown that systemic LPS administration immediately activates microglia in the brain, and this effect could be inhibited by ginsenosides. The effects of GTS and Rh3 were partly achieved by regulating 5'-adenosine monophosphate-activated protein kinase and its downstream signalling molecules, such as PI3K/Akt and NF-κB/ nuclear factor E2-related factor 2, thereby inhibiting the expression of inducible nitric oxide synthase and proinflammatory cytokines and enhancing the expression of anti-inflammatory hemeoxygenase-1.72, 73 The regulatory role of Rh1 is based on the physiological basis of regulating inflammation, neurotoxicity and oxidation through PKA and hemeoxygenase-1.74 Rg3 also decreased the expression of inducible nitric oxide synthase, cyclooxygenase-2, TNF-α, IL-1β and IL-6 in brain tissue after LPS injection.75 Moreover, Rb1 attenuates damage to cerebral cortex neurons by downregulation of nitric oxide, superoxide and TNF-α expression in hypoxia-activated microglia.76 These studies may provide therapeutic potential for the treatment of depression with microglia activation (Figure 1).

3.5 Effect on other mechanisms

Panax quinquefolium has shown the protective effects against olfactory bulbectomy-induced depression-like behaviours and nitric oxide pathway might be involved.77 GTS, ginsenoside Rg3 and Rb1 were demonstrated to resist stress by blockage of the activity of ODC and reduction of the accumulation of PUT, suggesting that they have potential neuroprotective effects in the brain.78 It was found that the water extract of Panax ginseng could protect cells from CORT-induced impairments via the intervention of GR-related functional proteins such as histone deacetylase 6 and heat-shock protein 90 in vitro and succedent functional recovery of endoplasmic reticulum and mitochondria.79 In vitro, Rg3 recovered growth and suppressed apoptosis by regulating the cell cycle in HT22 cells treated by N-methyl-d-aspartic acid.80 Amyloid-β peptides (Aβ), regarded as a main pathogenic factor of Alzheimer's disease, is playing an emerging role in stress response as well as in depression. Aβ administration in rats induced depression-like behaviours, changed the activation of the HPA axis and decreased the levels of 5-HT and neurotrophic factors in the cortex.81 Rg2 might protect PC12 cells from Aβ25-35-induced apoptosis by affecting the PI3K/Akt signalling pathway.82 Moreover, numerous reports have shown that many other kinds of ginsenosides, such as Re, Rg1 and Rf, had the capability to downregulate the Aβ deposition and the tau protein phosphorylation.83-85 These results suggest that these ginsenosides might have antidepressant effects through regulating Aβ. It was known that gap junction intercellular communication (GJIC) dysfunction might be involved in the potential pathogenesis of depression.86 Pre-treatment with Rg1 can ameliorate GJIC in the astrocytes of the PFC and hippocampus in rats treated with CORT. Also, the Cx43 expression was increased and the Cx43 phosphorylation was decreased, which may be of great significance for the treatment of depression.87, 88

Overall, many studies have shown that the regulatory role of Panax ginseng appears to be crucial in depression (Table 4). The action of Rg1 is shown in Figure 2 as an example of intracellular signalling pathways that illustrates the antidepressant action of ginseng active ingredients. Considering that Panax ginseng and its active ingredients were reported to regulate the monoamine neurotransmitters, HPA axis, neurotrophic factors and neurogenesis, the relationship between Panax ginseng and the pathogenesis of depression might be a potential novel target for depression treatment.

4.1 Kai Xin San

Kai Xin San (KXS) is a classical Chinese herbal prescription which is composed of four kinds of medicines: Panax ginseng, Polygala tenuifolia, Poria Cocos and Acorus calamus. As a classic formula still used today, KXS should benefit Qi first; therefore, ginseng is the main medicine in the prescription. Only when ginseng invigorating Qi and cultivating blood, can KXS achieve the effect of calming the mind. Therefore, the compatibility features of the prescription are that Panax ginseng and Poria cocos are used to tonify Qi and nourish the heart, combined with Polygala and calamus to tranquilize the mind. Clinically, the compatibility ratio of KXS varies according to the specific situation, which results in the serial formulations of KXS.

Chemical studies showed that the main components of KXS were ginsenosides, Polygala tenuifolia glycosides and volatile oils of Acorus calamus. It is through these main active substances that KXS exerts neuroprotective and antidepressant effects. Dang et al found that KXS can reduce the expression of AChE in the hippocampus, increase concentrations of various monoamine neurotransmitters in the hippocampus and PFC and reduce the concentration of serum ACTH, significantly improving the depression symptoms in mice subjected to CMS.89 KXS was found to ameliorate 5-HT defects, which is achieved by raising the synthesis, inhibiting the reuptake and ultimately increasing the concentration of intracephalic 5-HT.90, 91 In an earlier study, impacts and potential mechanisms of KXS2012 on depression in both cell and animal models were validated. Moreover, it promoted neurogenesis via inducing the expression of synaptotagmin and dendritic spine density in cultured neurons and increased neurotrophins in astrocytes which might concern the regulation of Erk1/2 and CREB phosphorylation.92 In further studies of Zhu et al,93 KXS was confirmed to promote expressions of neurotrophic factors and TrkB receptors. By combining qPCR, Western blotting and constituent identification analysis, ginsenoside Rg1 and Rb1 may be active compounds in this component that increase NGF and BDNF expression by activating cAMP-dependent signalling pathway and stimulating neurotrophic factor synthase.94 Studies both in vivo and vitro have shown that KXS can facilitate hippocampal synaptogenesis by upregulating synaptic proteins expression.95 There is also evidence that KXS seems to improve memory impairment induced by hindlimb suspension mainly by decreasing serum ROS, 8-OHdG and 3-nitrotyrosine and inhibiting oxidative stress injury, rather than influencing the central cholinergic system.96 In addition, KXS treatment effectively improved depressive symptoms in fluoxetine-resistant depression rats by the reduction of COX-2, IL-2, IL-6, TNF-α levels and increase of IL-10, IFN-γ level.97 With the deepening of research, the neurobiological mechanism of KXS and its specific active ingredients in preventing and treating depression will be further elucidated.

4.2 Shen Yuan Gan

Ginseng and Polygala tenuifolia are often used together in the clinical treatment of depression and have shown good antidepressant effects in animal experiments and clinical studies. Shen Yuan Gan (SYG), a Chinese herbal compound, consists of GTS and Polygala tenuifolia total glycosides (PTG) in a proportion of 2 to 1. Literature reports showed that the effective components of Panax ginseng and Polygala tenuifolia against depression were GTS and PTG, respectively. The antidepressant effects of SYG have been confirmed in various depression models. In previous studies, Sun et al98 found that SYG administration could alleviate depression in TST and FST, as well as regulate CORT levels of learned helplessness rats and BDNF levels in the hippocampus of CMS rats. In subsequent trials, Sun et al99 found SYG not only ameliorated CMS-induced changes, but also recovered concentrations of 5-HT, DA, NA and acetylcholine (Ach) in the PFC, which indicated that antidepressant actions of SYG are likely to be connected with the level of monoamine neurotransmitters. Besides, SYG may also enhance cognition by increasing ACh in the PFC. Therefore, SYG has a clear antidepressant effect and can be used as a potential selective antidepressant.

4.3 Xiaochaihu Decoction

Xiaochaihu Decoction (XD), originated from Shanghan Zabing Lun of Zhang Zhongjing-a distinguished physician in Eastern Han Dynasty, has been used in the treatment of Shaoyang syndrome which involves depressive-like symptoms in China for thousands of years. After many years of evolution, the prescription of XD has the merits of strict compatibility and definite curative effect. It is composed of seven herbs: Bupleurum as the monarch drug, Scutellaria as the minister drug, Panax ginseng, Pinellia and Glycyrrhiza as the assistant drug and Ginger and Jujube as the guide drug.100 Among them, Scutellaria, Panax ginseng and Glycyrrhiza might be the core components of XD, which has the function of strengthening the spirit and tonifying Qi. Clinical and modern pharmacological studies of XD have proved that it has a satisfactory antidepressant effect, and the active ingredients are mainly flavonoids and saponins.

Lots of animal researches have proven that the therapeutic effects of XD on depressant-like behaviours are likely mediated by upregulating the monoamine neurotransmitter, neurotrophin expression and neurogenesis in related brain regions of rodents. Su et al101 found that XD improved depressant-like behaviours, including TST, FST, hypothermia induced by reserpine and head-twitch induced by 5-HTP, which may be related to the increased levels of 5-HT, 5-hydroxyindoleacetic acid and 5-HT turnover in the hippocampus and PFC. In their further study, the 5-HT and DA concentrations, and the BDNF, NGF, TrkB and TrkA expressions in the hippocampus also significantly increased after XD administration, which was reduced in CUMS rats.102 Moreover, XD treatment in chronically isolated rats might increase neurogenesis by upregulating 5-HT levels, activating the 5-HT_1A receptor and then promoting expressions of downstream CREB and BDNF in the hippocampus.103 XD could remarkably alleviate chronic CORT-induced anxiety/depression-like behaviours, which were probably attributed to promoting hippocampal neurogenesis and remodelling the integrity of the negative feedback loop on the HPA axis.104 Additionally, XD was found for the first time to enhance the expression of monoamine neurotransmitter synthase (TPH2 and TH), inhibit the expression of 5-HT transporter and decrease the expression of monoamine neurotransmitter degrading enzyme (MAOA) in the hippocampus of social isolation-reared mice.105 These studies lay the foundation for the study of pharmacodynamic substances of XD's antidepressant effect.

As mentioned above, Panax ginseng herbal formulae have multiple functions on anti-depression and the confirmation of the mechanisms is progressing (Table 5). For the classical prescriptions used so far, there are few systematic studies on their specific effective material basis for the treatment of depression. In recent years, in the study of the modernization of traditional Chinese medicine, it is a hot spot to take the traditional Chinese formulae as the natural combinatorial chemical library and to study it by the method of combinatorial Chinese medicine. That is to say, by separating and extracting every single drug in the compound, the specific effective molecules of traditional Chinese medicine are combined together to synthesize a relatively safe natural combinatorial chemical library with curative effect. It is also the direction for further systematic research.