The hypothalamic-pituitary-adrenal (HPA) axis is one of the major biological stress response systems in humans, of which activation is essential to provide an appropriate biological response to stress (Guerry and Hastings, 2011). Activation of the HPA axis in stressful situations is a normal homeostatic mechanism that can contribute to the maintenance of stability and health by enabling adaptive changes in the body. For a healthy population, normal levels return when this stress response is no longer biologically significant. However, studies have found that abnormal activity of the HPA axis, such as excessive activity or persistent disorder, is closely related to the development of depression (Nelson and Davis, 1997; Curtis and Mendels, 1976; Bardeleben and Holsboer, 1989). Previous research has shown that the activity of the HPA axis is governed by vasopressin (AVP) and corticotropin-releasing factor (CRF) which is secreted by the hypothalamus and undergoes a cascade of events. Specifically, when the body is stimulated by stress, the adrenal glands secrete a surge of adrenaline, followed by the activation of the HPA axis. The hypothalamus secretes CRF for release into the portal circulation, and this in turn activates the pituitary to secrete the adrenocorticotropic hormone (ACTH), which ultimately stimulates the adrenal cortex to secrete glucocorticoids and cortisol (Pariante and Lightman, 2008). In addition, cortisol induces inhibitory feedback of the HPA axis by interacting with glucocorticoid (GR) and mineralocorticoid (MR), such as the secretion of ACTH, CRF, and AVP (Pariante and Lightman, 2008; Juruena et al., 2018; De Kloet et al., 1998). The normal physiological activity of the HPA described above maintains the relative homeostasis and health of the organism under certain stressful circumstances. However, excessive or prolonged psychological stress may alter the normal state of homeostasis in the body, which leads to depression, a serious mental illness that affects people’s mental and physical health. Studies have found that the most common biological abnormality in depression is HPA axis overactivity, which is characterized by increased cortisol, adrenal hyperplasia, and negative feedback abnormalities (Dinan and Scott, 2005; Dwyer et al., 2020). Fortunately, previous studies have shown that monoamine-based antidepressants can reverse stress-induced hyperactivity of the HPA axis (Seki et al., 2018). GR receptor antagonists, press-in receptor antagonists, and adrenocorticotropin-releasing hormone receptor antagonists can also exert antidepressant effects by blocking receptor activity to terminate hormone secretion resulting from HPA axis overactivity (Menke, 2019), suggesting that components of the HPA axis may be potential targets for the treatment of mood disorders (Thakore et al., 1997).

Most researchers suggest that PF has promising antidepressant activity by modulating HPA axis dysfunction. Qiu et al. found that intraperitoneal injection of PF (30 or 60 mg/kg) for 4 weeks significantly increased sucrose consumption, and decreased serum corticosterone (CORT) and ACTH levels in the chronic unpredictable stress (CUS) model group, which suggests that PF may exert antidepressant effects in CUS rats by regulating the HPA axis (Qiu et al., 2013). This preclinical study suggests and explores the antidepressant effects of PF and its underlying mechanisms; however, several significant limitations persist. Firstly, the generalizability of the findings is restricted due to the exclusive use of male SD rats as experimental subjects. Additionally, the CUS paradigm employed may not fully replicate the complex stressors encountered by humans. Furthermore, the study utilized only intraperitoneal (i.p.) injection as the method of administration, which may differ from clinical drug delivery methods. Increasing the number of studies on multiple routes of administration, such as oral and transdermal patches, and comparing the antidepressant effects and pharmacokinetic profiles of PF under different modes of administration may provide some reference for the clinical application of the drug. Similarly, the depression model of rats established that the forced swimming trial (FST) was treated with PF (10 mg/kg) or fluoxetine (20 mg/kg) by gavage three times at 24 h, 5 h, and 1 h prior to the behavioral experiments, including the forced swimming and the open field trials. Both fluoxetine and PF could significantly shorten the immobilization time of the 5-min forced swim. In addition, the distance traveled in the open field did not significantly change in both groups compared to the normal control in the Open-Field Test (OFT) (Mu et al., 2020). Further studies revealed that PF was able to increase tolerance to stressors in rats by modulating the hyperactive HPA axis, exerting antidepressant effects similar to those of fluoxetine, as evidenced by the significant reduction of corticotropin-releasing hormone (CRH), ACTH, and CORT in plasma and hippocampus (Yan-xia et al., 2014). Meanwhile, the total glucosides of paeoniflorin (TGP) are active metabolites extracted from Paeonia lactiflora Pall. [Paeoniaceae, Paeonia lactiflora root], including PF, hydroxyl-paeoniflorin, paeonin, albiflorin, benzoylpaeoniflorin. Whereas, PF is the main active metabolite of TGP, which accounts for more than 40% of TGP (Zhang and Wei, 2020). Mao et al. found that TGP significantly inhibited the behavioral and biochemical changes in the chronic unpredictable mild stress (CUMS) mice, and could dose-dependently reduce the serum CORT levels in CUMS-exposed mice. The TGP may also exert an antidepressant-like effect by regulating the function of HPA (Mao et al., 2009a).

Numerous studies have found that prenatal stress (PS) increases the risk of neurological, endocrine, and metabolic disorders, which induces depression in offspring. To stressed offspring, PF (15, 30, and 60 mg/kg/day) administered for 28 consecutive days significantly increased sucrose intake and reduced immobility time as well as the total number of crossings, center crossings, rearing, and grooming in model rats. However, PF could restore the levels of ACTH, CRH, and COR in PS offspring (Li et al., 2020). For the ovariectomized and CUS-induced menopause depression model rats, PF could downregulate the serum levels of CRH, CORT, and ACTH, correct the hyperfunctioning of the HPA axis, which resulted in a significant improvement in the abnormal behaviors of the model rats (Huang et al., 2015). In summary, PF results suggest that PF could regulate mood disorders and exert antidepressant-like effects by modulating the disordered HPA axis function.

Most trial and clinical evidence have shown that monoaminergic neurotransmitters such as serotonin (5-HT), norepinephrine (NE), and dopamine (DA) have a wide range of biological activities and are important regulators of a series of physiological activities, such as mental activity, behavioral state, and emotion in the CNS (Nemeroff, 2002). Thus, physiological changes in abnormal monoaminergic neurotransmitters, including 5-HT, NE, and DA signal transduction could change receptor regulation and function, or impaired intracellular signal processing (Liu Y. et al., 2018), which trigger various emotional changes (Hamon and Blier, 2013). 5-HT, a biochemical messenger and modulator synthesized by decarboxylation of L-tryptophan, produces a variety of “state-dependent” behavioral responses to different stimuli, which have been shown to be useful in the treatment of anxiety and obsessive-compulsive disorder. The defects in the 5-HT system lead to disorders such as depression, phobias, obsessive-compulsive disorder, generalized anxiety disorder, and post-traumatic stress disorder (De-Miguel and Trueta, 2005). The most common of the pathophysiologic hypotheses in depression is the monoamine hypothesis, which posits that alterations in monoamine neurotransmitters are responsible for the pathogenesis of depression, such as decreased concentrations, abnormal function, and defective transmission across the synaptic gaps in the brain (Hirschfeld, 2000). Evidence for this hypothesis comes mainly from clinical observations and animal experiments. Reserpine, the antihypertensive drug, has been shown to deplete central stores of monoamines, which can trigger depressive-like manifestations such as bradykinesia and sedation (Delgado, 2000). In contrast, isoniazid could increase the concentration of NE and 5-HT in the brain by inhibiting monoamine oxidase, making the subject feel euphoric and active (Buda et al., 2022). The monoamine oxidase inhibitors (MAOIs) iproniazid also have antidepressant effects when used in patients with tuberculosis (Delgado, 2000). Therefore, increasing the monoaminergic neurotransmitter levels and enhancing the function of monoaminergic neurotransmitter systems are effective options for the treatment of depression. According to this hypothesis, various antidepressant drugs have been discovered, such as tricyclic antidepressants (TCAs) that block the reuptake of monoamines in presynaptic neurons, and MAOIs that prevent the breakdown of monoamines after reuptake and enhance neurotransmitters to exert antidepressant effects (Kessing et al., 2024).

The study found that the treatment of PF significantly attenuated the decrease of NA, DA, 5-HT, and metabolite 5-hydroxyindoleacetic acid (5-HIAA) in chronic CUS model mice, as well as the increase in the ratio of 5-HIAA/5-HT in the model (Qiu et al., 2013; Wang Jing-xia et al., 2010). Similarly, Intragastric administration of PF (10 mg/kg) could increase the levels of 5-HT and NE in plasma and hippocampus of the forced swimming test (FST) depression model rats (Mu et al., 2020). Reserpine is a vesicle reuptake inhibitor that irreversibly inhibits the vesicular uptake of monoamine neurotransmitters, including NA, DA, and 5-HT, which are metabolized and depleted by monoamine oxidase (MAO) inducing behavioral and physiological changes in animals (Ye et al., 2024). Based on the above principles, the rifampicin-antagonistic experimental model was the first established animal model of depression (Bourin et al., 1983). Intragastric administration of PF or TGP dose-dependently antagonized hypothermia, akinesia, and blepharoptosis in reserpine-induced mice, and reversed reserpine-induced decreases in monoamine transmitters such as NE, DA, and 5-HT in the brain, which showed an obvious anti-resensitization effect (Mao et al., 2008; Guang-zhi, 2012; Jin Shumei and Cui, 2013). These studies suggested that the antidepressant-like effect of PF or TGP may be realized by protecting monoamine neurotransmitters. In addition, PF or TGP could protect monoamine neurotransmitters by dose-dependently inhibiting the activities of MAO-A and MAO-B in the mouse brain monoamine oxidase (Mao et al., 2009b).

5-HT1A receptors, the largest class of 5-HT receptor subtypes, are mainly distributed in the frontal cortex, hippocampal area, lateral septal nucleus, and dorsal nucleus of the middle suture, which are closely associated with anxiety and depression (Sargent et al., 2010). 5-HT2A receptors are densely distributed in the hippocampal area, amygdala, prefrontal cortex, and olfactory cortex executive area, closely associated with suicide, depression, and schizophrenia (Papp et al., 1994). The mRNA and protein expression levels of the 5-HT1A receptor in the hypothalamus of menopausal depression rats by CUMS model were significantly lower than those of normal rats, while the mRNA and protein expression levels of the 5-HT2A receptor were significantly higher than those of normal rats. After gavage of PF (10 mg/kg) for 2 weeks, the mRNA and protein expression levels of the 5-HT1A receptor in the hypothalamus of the model rats were increased, while the expression level of the 5-HT2A receptor was decreased. These results indicate that PF can treat climacteric depression by adjusting the different 5-HT receptor subtypes in the hypothalamic region of rats (Huang et al., 2015). The model established by ovariectomy combined with long-term CUS in this study, while effectively simulating the physiological changes and stressors associated with menopause, still fails to capture the complex etiology and manifestations of human menopausal depression, and cannot fully replicate the realities of the human condition. In addition, although serum CRH, ACTH, CORT, and prefrontal cortex monoaminergic neurotransmitter levels, mRNA, and protein expression were measured, which confirmed the antidepressant effect of PF at the molecular level. On this basis, we suggest an in-depth exploration of the specific molecular targets where PF acts and the related signalling pathways. In addition, PF could activate the release of monoamines in the rodent brain, inhibit the reuptake of NA and 5-HT, and increase the content of DOPAC and 5-HIAA in the brain, which are the metabolites of 5-HT and DA (Lin et al., 2019; Qingwei, 2018).

Oxidative stress is caused by the overproduction of reactive oxygen species (ROS) or defective antioxidant systems, strongly associated with diseases such as cardiovascular disease, cancer, and diabetes (Poprac et al., 2017). In addition, ROS is widely recognized as a main cause of brain damage. Specifically, the brain effectively regulates oxygen consumption and redox-generating capacity by neutralizing the deleterious effects of ROS production through the antioxidant system, under normal physiological conditions. When this regulatory system is dysregulated, excessive ROS will cause oxidative damage to a series of biomolecules such as DNA, proteins, and lipids, and even lead to functional decline (Patel, 2016). Therefore, oxidative stress may be associated with the development of a range of neurodegenerative diseases or mental disorders, such as Alzheimer’s disease, Parkinson’s disease, cerebrovascular disease, attention deficit hyperactivity disorder, schizophrenia, and autism spectrum disorders (Vavakova et al., 2015). Most studies have shown that oxidative stress is closely related to depression. The depressed patients have elevated levels of ROS, reactive nitrogen species (RNS) (Suzuki and Colasanti, 2001; Dhir and Kulkarni, 2011; Maes et al., 2011), and reduced activity of the antioxidant glutathione (GSH) in postmortem brain samples (Gawryluk et al., 2011). In addition, the expression of the enzymes involved in ROS production (xanthine oxidase and monoamine oxidase) was increased in depressed patients. For example, xanthine oxidase (XO), which catalyzes the oxidation of xanthine to produce superoxide and hydrogen peroxide, has been observed to elevate levels in the serum of depressed patients and in the thalamic region of post-mortem depressed patients (Michel et al., 2010). MAO levels, of which the by-products, such as hydrogen peroxide, leading to excessive production of ROS, are higher in depressed and postpartum depressed patients than in nondepressed subjects (Sacher et al., 2015), resulting in neuronal apoptosis and mitochondrial dysfunction. Therefore, anti-oxidative stress may be an effective strategy for the treatment of depression.

Numerous studies have demonstrated that PF could exert neuroprotective effects by inhibiting oxidative stress (Wang X. et al., 2021) (Figure 3). Superoxide dismutase (SOD) is an important antioxidant enzyme that catalyzes the breakdown of superoxide into oxygen and hydrogen peroxide, which is degraded by catalase under physiological conditions (Sies, 2015). In addition, the increase of ROS could lead to lipid peroxidation of the cell membrane to produce a large amount of malondialdehyde (MDA), a product of oxidative stress. Clinical studies have reported higher serum levels of MDA in patients with major depression compared with controls (Sies, 2015). After multiple intragastric administrations of PF, the plasma SOD level of the depression rats in the FST model was increased, and the plasma MDA level was decreased (Mu et al., 2020). PF significantly shortened the 5-min swimming immobility time of the rats in the FST model and increased the 5-min moving distance in the open-field trials, showing an antidepressant effect similar to fluoxetine. These studies, however, still exhibit certain deficiencies in their design. Firstly, the use of fluoxetine exclusively as a positive control in the antidepressant group restricts a comprehensive evaluation of the advantages and disadvantages of PF in comparison to other similar medications. Additionally, the absence of a detailed randomization methodology for the experimental subgroups may compromise the results. Furthermore, the experimental design featured a limited range of behavioral tests, encompassing only the forced swimming test and the open field test, and was conducted at only a few specific time points, which hindered the ability to observe the effects of PF across different time periods. In CORT- or glutamate-treated PC12 cells, PF increased the cell viability, the levels of GSH, the SOD and catalase activities, meanwhile, decreased intracellular reactive ROS and MDA levels in a dose-dependent manner (Mao et al., 2012a; Mao et al., 2011a; Mao et al., 2010). In addition, long-term PF (15 mg/kg and 30 mg/kg, i.p.) treatment of Aβ (1–42) (1 μg/μL) in rats restored the decreased activities of SOD and catalase, increased the level of MDA and the content of reduced GSH, which suggests that the PF can exert neuroprotective effects by alleviating oxidative stress (Lan et al., 2013; Zhong et al., 2009). Furthermore, Mao QQ et al. found that TGP (80 and 160 mg/kg) treatment in CUS mice dose-dependently reduced GSH depletion and MDA production, which further suggests that the antidepressant-like activity of TGP may be mediated by attenuating oxidative stress in the mouse brain (Mao et al., 2009b).

Apoptosis, a form of programmed cell death, plays a critical role in tissue homeostasis, immune cell renewal, and neuronal development, which prevents the damage of surrounding tissues by timely eliminating senescent or damaged cells in the body (McKernan et al., 2009). Apoptosis is a tightly regulated and energy-dependent process, characterized by cytoplasmic shrinkage, chromatin condensation, nuclear pyknosis, ordered DNA fragmentation, cell rounding, and membrane blistering. Cells eventually form the membrane-bound vesicles called “apoptotic vesicles”, which could be phagocytosed by neighboring cells. Most studies have shown that neuronal apoptosis plays a crucial role in most CNS diseases. For instance, excessive apoptosis of a series of neurons can lead to neurodegeneration such as Alzheimer’s disease, Parkinson’s disease, and Huntington’s disease (Su et al., 1994; Jenner and Olanow, 1998; Kim et al., 1999). In addition, apoptosis may also be caused by stress (Lucassen et al., 2006). The study showed numbers of apoptotic and necrotic cells were observed in depressed mice with chronic mild stress (CMS) (Tian et al., 2019). Kosten et al. found that exposure to unpredictable stress decreased Bcl-2 mRNA in limbic structures of the brain and frontal cortex, and Bcl-xL mRNA in the hippocampus (Kosten et al., 2008). Meanwhile, the Administration of alanylcyclopropionate, reboxetine, and fluoxetine upregulated Bcl-2 mRNA levels and also increased Bcl-xL mRNA expression. Moreover, Bcl-2 expression in the hippocampus was increased after 14 days of administration of citalopram, promethazine, and amitriptyline (Kosten et al., 2008), which suggests that potential antidepressants could be developed through an anti-apoptotic mechanism.

Studies have shown that PF, the natural plant metabolite from the treasure trove of nature, has surprising neuroprotective effects, inhibiting excessive neuronal apoptosis and exhibiting good therapeutic effects on neurodegenerative diseases (AD, Parkinson’s disease, etc.) and mental disorders (anxiety, depression, etc.) (Zhong et al., 2009; Zhai et al., 2019; Li et al., 2014; Zhang et al., 2017; Cong et al., 2019; Zhang et al., 2015; Wang et al., 2013b; Wu et al., 2013; Sun et al., 2012; Zheng et al., 2016). The accumulation of glutamate (Glu) in the synaptic cleft could produce excitatory neurotoxicity that may contribute to depression, anxiety, post-traumatic stress disorder, schizophrenia, cognitive impairment, and other psychiatric disorders (Mehta et al., 2013). Wang X et al. suggest that PF significantly improves the Glu-induced decrease in SH-SY5Y cell viability in human neuroblastoma cells by affecting the expression of Bax/Bcl2, cleaved caspase-3, and cleaved caspase-9, suggesting that PF significantly reduces cell apoptosis and exerts neuroprotective effects through the Bax/Bcl2 pathway (Wang X. et al., 2021) (Figure 4). Moreover, PF exhibited a suppressive effect on apoptosis by attenuating mitochondrial membrane potential, promoting cytochrome c release, and counteracting the upregulation of caspase-3 and caspase-9 in the case of Aβ 25–35 induced PC12 cell damage (Li et al., 2014). Treatment with 200 μM CORT resulted in apoptosis of PC12 cells for 48 h. Interestingly, the treatment of TGP protected PC12 cells against CORT-induced toxicity in a dose-dependent manner, which was associated with the inhibition of caspase-3 activity involved in the mitochondrial pathway and the upregulation of the bcl-2/bax mRNA ratio. Meanwhile, TGP in mice also produced similar antidepressant effects (Mao et al., 2009c). Similarly, XiaoyaoSan, a characteristic Chinese traditional formula, reduced chronic stress-induced anxiety and depression behaviors in mice. PF, one of the main active plant metabolites in XYS, could protect primary neurons from CORT-induced neurotoxicity and reverse neuronal apoptosis caused by miR-200a-3p and miR-200b-3p overexpression (Yuan et al., 2022). It is important to note that XiaoyaoSan, as a Chinese medicine formula, has a complex composition that includes various plant metabolites beyond PF. This study focused solely on miR-200a/b-3p and PF, neglecting the potential roles and interrelationships of other miRNAs and metabolites within the formula. Furthermore, the use of a stereotactic microinjection method to investigate the role of miR-200a/b-3p in stress behavior may pose a risk of damaging rat brain tissue, which could subsequently interfere with the experimental results. The above studies suggest that PF may exert antidepressant effects through an anti-apoptotic molecular mechanism.

Calcium signaling regulates a range of neuronal activities by controlling the release of multiple neurotransmitters (Bezprozvanny and Mattson, 2008; Bergantin et al., 2013). Dysregulation of calcium signaling, such as excessive Ca²⁺ influx, may be associated with aging, Alzheimer’s disease, and major depression (Bergantin and Caricati-Neto, 2016). The abnormality of serum calcium level may be related to cognitive impairment in patients with depression (Grutzner et al., 2018). Normally, the intracellular Ca²⁺ concentration is maintained at a stable low level. Excessive Ca²⁺ influx may over-activate the Ca²⁺/CaM/CaMKII signaling pathway, impair neuronal activity, and induce depression when the homeostasis is changed. In addition, previous studies have found that the development of depression leads to neuronal death in the hippocampus and limbic brain. And the dysregulation of intracellular Ca²⁺ homeostasis seems to be closely related to this abnormal neuronal death (Duman, 2004; Bergantin, 2020). Therefore, maintaining normal Ca²⁺ homeostasis and safeguarding the proper regulation of calcium signaling pathways is one of the mechanisms to protect the nervous system.

Calbindin-D28K, one of the major calcium-binding proteins in the brain, could maintain intracellular Ca²⁺ homeostasis by binding excess Ca²⁺ to protect normal neuronal activation and function and inhibit neuronal apoptosis caused by intracellular Ca²⁺ overload (Carter et al., 2008; Meng et al., 2007). Experiments showed that PF could reverse the decrease of Calbindin-D28K mRNA level and the increase of Ca²⁺ concentration in PC12 cells induced by Glu, suggesting that the neuroprotective effect of PF is related to the inhibition of intracellular Ca²⁺ overload (Mao et al., 2010; Mao et al., 2011b). This result may be one of the pathways of antidepressant activity of PF in vivo. Meanwhile, PF treatment widely observed in other in vitro and animal trials could significantly block intracellular calcium influx caused by adverse stimuli such as methyl-4 phenylpyridine ion (MPP⁺) (Wang et al., 2013b), Aβ(1–42) oligomer (Zhong et al., 2009), and Glu (Wang X. et al., 2021; Wang et al., 2013c), which improve intracellular calcium homeostasis to play a neuroprotective role. In addition, previous studies have shown that the Ca²⁺/CaMKI/CREB signaling pathway plays an important role in intracellular signaling pathways involved in cell proliferation, cell survival, inflammation, and metabolism (Meng et al., 2007; Valera et al., 2008; Zhang et al., 2012). Calcium/calmodulin-dependent pathways may be overactivated to irreversible cellular damagewhen calcium is overloaded in neurons. PF reversed the significant reduction of p-CaMKII and p-CREB and regulated the expression of downstream proteins in the Middle Cerebral Artery Occlusion(MCAO) model and N-methyl-D-aspartic acid receptor(NMDA) induced excitatory toxicity model of primary hippocampal neurons, including Bax, Bcl2, Bad, and Caspase3, along the Ca²⁺/CaMKI/CREB signaling pathway (Zhang et al., 2017). Song et al. found that the effect of PF on the Ca²⁺/CaMKI/CREB signaling pathway may be achieved by regulating the current density of voltage-gated Ca²⁺ channel Cav1.2 (Song et al., 2017). Another study found that PF could significantly inhibit Glu-induced CaMKII over-expression and prevent intracellular calcium overload in PC12 cells (Wang et al., 2013c), thus playing an important role in the treatment and remission of affective disorders.

Recently, clinical and preclinical evidence suggestthat neuroinflammation is an important factor in major depressive disorder. Studies using positron emission computed tomography (PET) imaging and 18 kDa translocator protein (TSPO) as a microglia biomarker have demonstrated neuroinflammation in several brain regions in depressed patients (Li et al., 2018; Setiawan et al., 2018). The research in animal models has also revealed the release of proinflammatory factors and activation of microglia in the animal brain, showing signs of anxiety and depression (Munshi et al., 2020; Wang Y. L. et al., 2018). The persistent sympathetic and parasympathetic under-activity in chronic stress situations and major depressive disorder (MDD) increase catecholamine levels and decreases acetylcholine levels, which increases levels of pro-inflammatory cytokines (TNF-α, IL-1β, IL-6, and IL-18, among others), which explains that depression-like symptoms can be induced directly by proinflammatory cytokines (Won et al., 2021). Moreover, proinflammatory cytokines could activate the kynurenine pathway, leading to the increase of neurotoxic metabolites, including 3-hydroxykynurenine, 3-hydroxy-anthranilic acid, and quinolinic acid, to cause brain damage (Kim and Won, 2017). Meanwhile, researchers have found that the activation of microglia by stress stimulation releases a large number of proinflammatory cytokines, which could destroy the neuroprotective mechanisms in the brain, impair neuroplasticity, and inhibit adult hippocampal neurogenesis, leading to the occurrence of depression-related symptoms (McKim et al., 2016).

The researchers have recently found that PF significantly inhibits the neuroinflammatory response by decreasing the over-activation of astrocytes (AST) and microglia, as well as the expression of pro-inflammatory cytokines such as IL-1β, IL-6 and TNF-α (Zhou et al., 2019; Liu H. et al., 2015; Nam et al., 2013) (Figure 5). PF prevented the upregulation of pro-inflammatory mediators (TNF-α, IL-1b, iNOS, COX2, and 5-LOX) in plasma and brain to the sustained activation of hippocampal AST and microglia caused by chronic cerebral insufficiency of cerebral perfusion or cerebral ischemia, suggesting that PF exerts delayed protective effects in ischemia-injured rats by inhibiting the peripheral and cerebral tissue inflammatory responses mediated in MAPKs/NF-kB (Guo et al., 2012; Liu J. et al., 2006). TLR4/NF-κB/NLRP3 signaling has been shown to regulate the inflammatory response of microglia, of which the activation promotes the over-expression and over-release of proinflammatory cytokines, leading to neuronal damage (Zhong et al., 2019). Cheng et al. found that PF can reduce the release of pro-inflammatory cytokines by regulating TLR4/NF-κB/NLRP3 signaling, thus reducing the damage of cytokines to neurons and reversing LPS-induced depression-like behavior in mice (Cheng et al., 2021). The study found obvious changes in inflammation in the amygdala for 4 weeks after subcutaneous injection of Interferon-α 15 × 10⁶ IU/kg, as modeled by depression-inducing mice. Interestingly, the trial after 4 weeks of pretreatment with PF (20 mg/kg or 40 mg/kg) reversed the depression behavior of mice, and the abnormal level of inflammatory cytokines in serum, medial prefrontal cortex(mPFC), ventral hippocampus (vHi) and the amygdala, including IL - 6, IL - 1β, TNF-α, IL - 9, IL - 10, IL - 12 and monocyte chemotactic protein 1 (Li et al., 2017). However, the experiment only established two PF dose groups, specifically 20 mg/kg and 40 mg/kg, which limits the ability to ascertain the optimal effective dose of PF and its dose-effect relationship. Furthermore, while PF was observed to reverse the increase in microglia density and reduce the levels of inflammatory factors, the specific molecular pathways through which PF influences microglia in the context of neuroinflammation and depressive-like behaviors have not been thoroughly investigated. Furthermore, recent studies have found that PF reduces neuroinflammation by inhibiting Casp-11-dependent pyroptosis signaling induced by the overactivation of hippocampal microglia in reserpine-treated mice, representing a novel mechanism by which PF attenuates depression (Tian et al., 2021).

Mitochondria, the energy factory of eukaryotic cells, produce energy through the metabolism of lipids, steroids, and proteins, which play a key role in maintaining cellular stability by regulating Ca²⁺ levels, maintaining ROS levels, and regulating apoptosis (Nunnari and Suomalainen, 2012). Mitochondrial biosynthesis occurs more rapidly during neuronal development because neuronal differentiation requires an increase in the mitochondrial genome and mitochondrial proteins (Calingasan et al., 2008; Cuperfain et al., 2018; Kasahara and Kato, 2018). Therefore, mitochondrial dysfunction prevents cells from meeting their energy requirements and involves impairment of neuronal communication, cellular resilience, and hippocampal neurogenesis, which leads to mood and psychiatric disorders (Perkins et al., 1997; Nagashima et al., 2020). Most studies have shown that mitochondrial dysfunction in brain regions may be closely related to the development of depression. For example, brain mitochondrial dysfunction and ultrastructural damage have been reported in mouse models of depression (Gong et al., 2011). 54% of patients were also found to exhibit depressive symptoms in a study of the prevalence of psychiatric comorbidities in subjects with mitochondrial cytopathies (Feng et al., 2020), suggesting that targeting mechanisms of mitochondrial dysfunction may be a possibility for developing new treatments for depression.

Studies have shown that PF could exert neuroprotective effects by preventing mitochondrial dysfunction. PF could significantly improve the viability of SH-SY5Y cells and human neuroblastoma cells damaged by Glu or Ab25-35, and significantly inhibit the increase of mitochondrial membrane potential and calcium concentration to protect SH-SY5Y cells from Glu-induced excitatory neurotoxicity (Wang X. et al., 2021; Li et al., 2014). Daily administration of PF (10 mg/kg once a day) for 21 days was able to significantly ameliorate the cognitive dysfunction for streptozotocin (STZ)-induced in mice model, This research has demonstrated that PF may be related to the significant attenuation of the STZ-induced mitochondrial dysfunction, including a significant increase in cytochrome C oxidase activity and ATP synthesis to significantly restore the function in hippocampal area and cerebral cortex (Wang D. et al., 2018). Moreover, PF can regulate mitochondrial membrane potential and maintain mitochondrial membrane integrity to attenuate or restore PC12 cell injury induced by unfavorable stimuli such as Aβ25-35, MPP⁺, Glu, and CORT (Li et al., 2014; Wang et al., 2013b; Sun et al., 2012; Zheng et al., 2016; Mao et al., 2009c). Further studies revealed that this preventive-protective effect by PF might be achieved by inhibiting the MKK4-JNK signaling pathway (Cong et al., 2019).

Autophagy, a process of cellular waste removal and regeneration, transports cytosolic components such as proteins and organelles to lysosomes for degradation and recycling to provide a constitutive mechanism for the turnover and recruitment of cellular components (Xiao et al., 2018; Miranda et al., 2018). In the CNS, autophagy is involved in cell surveillance, neuroinflammation, and neuroplasticity. The investigators found that neurons are vulnerable to autophagy defects and depend heavily on the level of autophagy composition for survival (Hara et al., 2006; Nishiyama et al., 2007). Several preclinical and clinical studies have shown that dysfunction of cellular autophagy may be an important factor in the occurrence and progression of psychiatric disorders such as depression (Komatsu et al., 2006; Nouri et al., 2020; Zhao et al., 2017). For example, autophagy biomarkers, such as the LC3II/LC3I ratio and BECN1, are significantly reduced in animal models of depression. Notably, PF can exert neuronal protection by modulating the autophagy pathway (Jiang P. et al., 2017). PF can have a significant protective effect against acidosis or MPP + -induced injury in PC12 cells; further research suggested that PF improved acidosis-induced activation of acid-sensing ion channels (ASIC).

Meanwhile, PF weakened the autophagy inhibition induced by MPP⁺ and significantly upregulated autophagy and the ubiquitin-proteasome pathway, which prevents α-synuclein accumulation(α-SYN) of synaptic nucleoproteins to reduce cell damage (Sun et al., 2011; Cai et al., 2015; Cao et al., 2010). α-SYN is a characteristic marker of all types of PD, which can be degraded by the ubiquitin-proteasome system (UPS) and autophagy-lysosomal pathway (ALP) (Kaushik and Cuervo, 2012). In another study, systemic administration of PF in a 6-hydroxydopamine (6-OHDA) -induced PD rat model showed that PF enhanced the autophagic degradation of α-SYN to protect DA neurons from 6-OHDA-induced neurotoxicity. Further Western blot results showed that PF could significantly reduce the level of ASIC1a, suggesting that PF may play a neuroprotective role by inhibiting the activation of ASIC, especially ASIC1a (Gu et al., 2016).

Brain-derived neurotrophic factor (BDNF), a key mediator of activity-dependent neuronal plasticity in the brain (Park and Poo, 2013), has a major impact on neuronal morphology, survival, and differentiation, increasing synapse sprouting and synaptic stability, and facilitating long-duration strengthening (Zagrebelsky and Korte, 2014). BDNF synthesis occurs predominantly in regions involved in emotional and cognitive functions, such as the hippocampus and frontal regions (Phillips, 2017). BDNF could bind and activate tropomyosin receptor kinase B (TrkB) to regulate the different cellular processes for the development and maintenance of normal brain function. In addition, several lines of evidence suggest that BDNF/TrkB signaling is involved with adult neurogenesis in the hippocampus and has differential effects on the dentate gyrus (DG) and subventricular zone (SVZ) (Colucci-D'Amato et al., 2020). Numerous studies have confirmed that BDNF is one of the important biomarkers of depression. The low levels of BDNF are associated with reduced synaptic plasticity and neuronal atrophy, which is consistent with the neurogenic hypothesis of depression. Several autopsy studies have demonstrated decreased BDNF expression in the hippocampus and prefrontal cortex of depressed patients (Dwivedi et al., 2003). In addition, significantly elevated BDNF levels were detected in the serum of patients treated with antidepressants. This suggests that BDNF is not only a marker of disease but also a potential predictor of antidepressant efficacy. Taking BDNF signaling as a breakthrough point would be a potential direction for the development of antidepressants.

Fortunately, a large number of studies have found that the natural plant metabolite PF could upregulate the expression of plasma BDNF and BDNF mRNA in the hippocampus and frontal cortex (Mu et al., 2020; Mao et al., 2009a; Qingwei, 2018; Mao et al., 2012b), as well as postsynaptic density protein 95 (PSD 95) in model animals with mood disorders (Liu S. C. et al., 2019), thereby preventing CUMS-induced synaptic plasticity defects and providing neuroprotection in animal models of mood disorders. Previous studies suggested that BDNF combined with p-CREB is the main regulator of neurogenesis and emotion regulation, which may be closely related to neural plasticity (Sasaki and Yoshizaki, 2015; Orlovsky et al., 2014). Hu et al. found that PF therapy plays a positive role in neural and emotional regulation by reversing the decreased expression of BDNF and p-CREB in the hippocampus caused by the MCAO and CUMS model (Hu et al., 2019). In addition, PF also significantly increased the levels of other neurotrophic factors, such as NGF protein and mRNA, in the frontal cortex of CUMS rats. Further study found that the treatment of PF not only significantly enhanced the protein expression and gene transcription of BDNF in CUMS rats, but also activated the expression of TrkB, a high-affinity receptor for BDNF, which promoted the proliferation of neural stem cells, differentiation into AST, and neurogenesis in the DG of the hippocampus in stressed rats. This result suggests that PF may play the role of an antidepressant through the BDNF/TrkB signaling pathway (Chen et al., 2019). Meanwhile, Chen et al. used PF to treat the withdrawal hormone simulated pregnancy (HSP) in a Postpartum depression (PPD) rat model and found that PF improved PPD symptoms by promoting the activation of the transporter TspO and BDNF/mTOR pathways in PPD rats, proving that PF may be an effective anti-PPD and anti-depression drug (Chen J. et al., 2022).