Literature search and study design

We evaluated the scientific literature using databases like Google Scholar, Web of Science and PubMed to conduct our literature search, taking into account all works published between November 2003 and October 2023. The following keywords were used: “Ashwagandha, “Withania somnifera”, "herbal medicine," "brain-related disorders," “depression” "inflammation," "oxidative stress," and "plant". The study only uses publications from well-indexed journals (systematic, meta-analysis, research, and review articles) that are linked to brain-related diseases or disorders. The search results were filtered using the titles and abstracts of the reputed journal (Khan & Ibrahim, 2022). Furthermore, from many reports, relevant publications were considered and rigorously examined in order to advance the discussion on the experimental evaluation of ashwagandha in the management of brain-related disorders.

Inclusion criteria

This study included a review or research article with good repute in scientific journals about the prevalence of brain disorders, plants and their ingredients, molecular techniques for oxidative and inflammatory stress, and a thorough discussion of the study (Abdulazeez et al., 2021).

Exclusion criteria

Ethnopharmacological evidence does not include relevant field studies, study localities, ethical information, and data which is not fully accessible to prevent the misappropriation of comprehensives (Moher, Liberati, Tetzlaff, Altman, & Group, 2010).

Rationale for the use of Ashwagandha

For centuries, ashwagandha has frequently been utilised as a most important herbal medicine in various traditional systems to improve numerous diseases and disorders including stress and depression (Ibrahim et al., 2021). Depending on the complexity of the ailment, ashwagandha either alone or in combination with other remedies is typically advised to use. Ashwagandha had a number of phytoconstituents like withanolides, and other alkaloids that are pharmacologically and medicinally important in neurological disorders including stress and depression. TNF-α, IL-6, LPO, SOD, and GSH are examples of inflammatory makers and brain oxidative stressors that ashwagandha modulates (Mikulska et al., 2023). It also reduces apoptosis and modulates multiple inflammatory pathways. This suggests that ashwagandha in conjunction with essential components, may alter the pharmacological efficacy. In this work, we examine a subgroup of ashwagandha that is beneficial for depression, taking into account their features, functional traits, and phytochemical effects. These plants were chosen because it has a long record of being used in Indian medical practices for memory-related disorders, such as stress and depression; (a) phytochemicals from these plant sources have been identified for their potential in treating depression or stress; (b) these herbs' antidepressant and antistress properties have been determined; and (c) pre-clinical or clinical studies have been conducted to verify their reputation.

Antidepressant effect of Ashwagandha

Possibly the most well-known advantage of ashwagandha is its capacity to diminish stress. It belongs to the class of molecules known as adaptogens, which aid the body in adjusting to stress and depression. Additionally, it lessens the activity of the body's hypothalamic-pituitary-adrenal (HPA) axis, which controls your reaction to depression (Salve, Pate, Debnath, & Langade, 2019). A short study on the 58 participants found that those who consume ashwagandha extract (250 or 600 mg) for approx. 56 days experienced significantly less stress than those who took a placebo (Salve et al., 2019). When compared to the placebo group, those who took the ashwagandha supplements also experienced improvements in the quality of their sleep. According to another study on 60 participants, anxiety levels significantly decreased in those who took ashwagandha extract (240 mg) daily for 60 days than in those who got a placebo (Speers et al., 2021).

Ashwagandha can have effects that are similar to imipramine's in terms of its antidepressant qualities. Research might be in favour of the usage of ashwagandha root-derived medications as mood stabilisers and for the treatment of anxiety and depression. Ashwagandha root may also improve learning and memory (Bhat, Akther, Najar, Rasool, & Hamid, 2022; Yeung, Hernandez, Mao, Haviland, & Gubili, 2018). Among the plant's bioactive phytochemicals are withaniamides, withaferin A, withanolide A and D. They all significantly affect its pharmacological properties and actions. A conventional root extract of ashwagandha produced with water and ghee, as well as aqueous, methanolic, and hydroalcoholic extract, showed antidepressant efficacy (Saleem, Muhammad, Hussain, Altaf, & Bukhari, 2020). Additionally, ashwagandha enhanced the effectiveness of popular antidepressants and enhanced the antidepressant effects of fluoxetine, a selective serotonin reuptake inhibitor, and imipramine, a tricyclic antidepressant, in mice and rat models of depression (Speers et al., 2021).

Role of Ashwagandha in oxidative stress

Previous studies reported that ashwagandha leaf extracts have anticancer effects on several human cancer cell types. It has been demonstrated that low dosages of hydroethanolic extracts cause differentiation, whereas high doses cause cancer cells to undergo apoptosis or growth arrest (Mikulska et al., 2023). As a result, they were offered as potential natural medications for differentiation-based therapy. Moderate amounts of withanone and aqueous extract have also been demonstrated to be protective against the excitotoxicity and memory impairment caused by glutamate and scopolamine, respectively. There hasn't been any information on comparative research on how alcohol and water extracts react to oxidative stress, a major contributor to neurodegenerative illnesses (Kumar et al., 2014). Both C6 (glioblastoma) and IMR32 (neuroblastoma) cells demonstrated protection against stress and maintained the differentiation state after being treated to oxidative stress and recovering in media supplemented with either ethanol or aqueous extract. Antioxidant effects of Ashwagandha's and its active phytoconstituents neuroprotection against stress in rats (Kaul et al., 2016; Sharma & Kaur, 2018). A study investigated the beneficial effect of Ashwagandha on recognised indicators of oxidative stress and DNA damage (H2AX) using cell-based assays. It is known that DNA double-strand breaks cause the phosphorylation of H2AX at Ser139 in mammalian cells. Using an anti-H2AX antibody for immunostaining, DNA damage foci in the nucleus can be quickly detected as being composed of the phosphorylated version of H2AX (H2AX) and other DNA damage response proteins (ATM, ATR, CHK-1, and CHK-2). These tests showed that ashwagandha extracts reduced the formation of ROS and H2AX generated by H2O2 and glutamate, indicating that their anti-oxidative characteristics had a role in neuroprotection, at least in part (Shah et al., 2015). This study reported that the protective effects of the water and alcohol extracts were equal (Shah et al., 2015). They reported recovering differentiated glial and neuronal cells in media supplemented with extract or withanone reduced glutamate-induced oxidative stress and DNA damage. Shah et al reported that better recovery was seen when extract and withanone were combined. The ability of the cells to survive was improved, along with the maintenance or induction of differentiation, as well as a decrease in ROS accumulation and the creation of H2AX foci (a sign of a reaction to DNA damage). Reductions in GFAP (a marker of glial cell differentiation), NF-200 (an axonal marker), and MAP2 (a diagnostic of dendritic structure) indicate that either H₂O₂ or glutamate-induced oxidative stress affects the primary cytoskeletal elements (myelinated axons and microtubules), which are essential for differentiated neurons (Kataria, Wadhwa, Kaul, & Kaur, 2012; Rajasankar, Manivasagam, & Surendran, 2009; Rajdev & Sharp, 2000). Furthermore, it has been observed that rats subjected to prolonged restraint stress experience changes in the expression and location of MAP2 in the cortex and hippocampus. It should be mentioned that in the current study, GFAP, NF-200, and MAP2 protein levels increased in cells treated with extract or withanone, indicating the preservation and protection of the functional states of both glial and neuronal cells (Kataria et al., 2012; Yan et al., 2010). These findings suggested that ashwagandha extracts and their constituent parts may have neuroprotective and neuro-differentiating properties, which are probably mediated via NF-200 and MAP2 signalling. Withanone was found to be less harmful to differentiated cells overall and to be more effective than withaferin A in all experiments (Kaul et al., 2016; Wang et al., 2021). It has been established that the alcoholic and water extracts of leaves contain different chemical components. The alcoholic extract contains withaferin A and withanone but not water; the latter was noted to include triethylene glycol. Because of the combined action of the active ingredients, which may exert their effects via several pathways, combination therapy is expected to provide improved protection. More investigation is necessary given the molecular characterization of these pathways (Shah et al., 2015). It's interesting to note that, in the absence of retinoic acid (RA), nuclear mortalin was shown to enhance the malignant properties of cancer cells by inactivating p53 and activating telomerase and hnRNP-K proteins. Mortalin was observed to translocate into the nucleus of neuroblastoma cells treated with retinoic acid, bind to retinoic acid receptors, and then increase their recruitment to the retinoic acid response element, where they were transcriptionally activated to produce genes involved in neuronal differentiation and reduce their proteasome-mediated degradation (Ryu et al., 2014; Shih et al., 2011). Mortalin was demonstrated to have a new role in actively promoting neuronal development by causing a significant drop in RA-triggered gene expression when it was knocked down (Grover et al., 2012). Ashwagandha extract or withanone administration was seen to generate nuclear enrichment of mortalin in IMR32 cells, which is similar to the impact of RA and suggests that these phytochemicals have the capacity to promote neuro-differentiation (Shah et al., 2015; Shih et al., 2011). Withanone and Ashwagandha leaf extracts are suggested as strong natural neurotherapeutic medicines based on these findings, which include (i) protection against oxidative stress, DNA damage, and glutamate excitotoxicity, and (ii) maintenance and activation of differentiation (Shah et al., 2015). To determine the signalling pathways and mechanisms involved in the therapeutic potential of each of these extracts and phytochemicals alone and in combination, more research is necessary.

Role of Ashwagandha in inflammation

In preclinical research, ashwagandha water extract reduced lipopolysaccharide-induced neuroinflammation by inhibiting inflammatory cytokines such as TNF-α, IL-1, and IL-6 as well as the production of nitrooxidative stress enzymes (Mikulska et al., 2023). Previous observation pointed to the possible utility of Ashwagandha in reducing inflammation of the nervous system linked to numerous neurological illnesses. In another study, it was found that inhibition of inflammatory markers including cytokines (TNF-a and IL-6), NO, and ROS, this plant has been shown in preclinical trials to be able to control mitochondrial activity, regulate apoptosis, and lessen inflammation. Meanwhile, the inhibitory effect of Ashwagandha powder was shown in circumstances such as proteinuria and nephritis in a mouse model of lupus (Dar, Hamid, & Ahmad, 2015). Rats in the positive control group (control group) received phenylbutazone treatment. Along with a notable decrease in inflammation, changes in the amounts of a variety of serum proteins, including 2 glycoproteins, acute phase protein 1, and albumin, were seen. In an investigation using the HaCaT human keratinocyte cell line, it was found that a water extract of ashwagandha root inhibited the NF-kB and MAPK (mitogen-activated protein kinase) pathways by decreasing the expression of pro-inflammatory cytokines such as interleukin (IL)-1, IL-6, IL-8, IL-12, and TNF-a and increasing the expression of anti-inflammatory cytokines such as (TGF)‑β1 (Rasool & Varalakshmi, 2006; Sikandan, Shinomiya, & Nagahara, 2018). These findings suggest that Ashwagandha's anti-inflammatory properties may one day be applied to the prevention of skin irritation.

Clinical trials of Ashwagandha related to depression

The effects of ashwagandha extract on depression have been the subject of inconsistent clinical trial research; nonetheless, numerous clinical trials have been conducted to assess the herb's effectiveness in connection to human intake, and the results have been favourable.

Intake of 1000 mg/day of ashwagandha extract for 12 weeks ameliorates symptoms of depression and anxiety associated with schizophrenia, according to a randomized, double-blind controlled research (Chandrasekhar, Kapoor, & Anishetty, 2012). Oral intake of ashwagandha standardized extract on 66 patients for a period of 12 weeks improved anxiety and depression. Another study found that taking 300 mg twice a day of ashwagandha root extract for 60 days significantly decreased perceived scores of stress and depression, which dropped to 44% from 5.5% in the placebo group. Serum cortisol levels were similarly lower in the ashwagandha root extract group than in the placebo group (Kelgane, Salve, Sampara, & Debnath, 2020; Speers et al., 2021).

Safety of Use

Ashwagandha has been used medicinally for a very long time, which is mainly proof of its efficacy and the body's high tolerance. In recent years, liver damage caused by ashwagandha has been experienced globally, hence, to dispel concerns regarding its use, a number of experts are currently collaborating. The pharmaceutical industry for herbal supplements is substantial and growing, both domestically and globally. It is therefore even more important to verify its safety (Langade, Kanchi, Salve, Debnath, & Ambegaokar, 2019).

The first evidence linking ashwagandha to liver damage was found in Japan in 2004. It concerned a 20-year-old male patient with congestive liver damage who recovered fully after quitting ashwagandha and taking ursodeoxycholic acid and phenobarbitone for two months to address his symptoms (Siddiqui, Ahmed, Goswami, Chakrabarty, & Chowdhury, 2021). Björnsson et al. claim that ashwagandha caused liver damage in the five volunteers. These incidents demonstrate Ashwagandha's potential hepatotoxic consequences. Liver damage typically manifests as cholestatic jaundice and pruritus, although it is a self-limiting illness that resolves in one to five months with normal liver test findings (Björnsson et al., 2020). Furthermore, a 39-year-old lady in the UK was reported to have had nausea and jaundice after using an over-the-counter herbal medication that contained ashwagandha extract (Lubarska et al., 2023). Hepatotoxic consequences have only sometimes and erratically been documented (Bokan et al., 2023). On the other hand, no toxicity of ashwagandha was confirmed by an Indian investigation including eighty perfectly healthy participants. Each participant took 300 mg of ashwagandha root extract orally twice a day for eight weeks. To ascertain this, variables such as plasma haemoglobin, neutrophil and platelet counts, alkaline phosphatase, aspartate transaminase, alanine transaminase, and body weight were monitored. At the conclusion of the study, there was no discernible change in the values of the aforementioned indicators between the 40 participants in the extract-using group and the 40 participants in the placebo-taking group. Triiodothyronine, thyroxine, and TSH levels were measured in the blood to assess thyroid function; however, no appreciable changes in these hormone levels were observed (Langade, Thakare, Kanchi, & Kelgane, 2021; Verma, Gupta, Tiwari, & Mishra, 2021).

A second study using a smaller sample size (18 volunteers) confirms the lack of significant effects on bilirubin, plasma protein levels, WBC percentage, RBC count, and ESR value. Conversely, blood urea nitrogen levels decreased and serum creatinine increased (Liang et al., 2017). The specialists of the study concluded that this event resulted from the concurrently documented increase in muscle mass. Over the course of ten days, participants ingested ashwagandha sluggard root aqueous extracts at increasing doses, beginning at the equivalent of 6 g and finishing at the equivalent of 10 g (Mikulska et al., 2023). Even though the plant has a lot going for it, pregnant or breastfeeding women shouldn't eat it. At this point, there is not enough information to say with certainty whether or not it is safe to use ashwagandha-containing products during such sensitive developmental periods. Some understanding of this aspect of safety can be gained from studies looking at how ashwagandha extract affects pregnant rats (Mikulska et al., 2023). The majority of emphasis was focused on the first five to nineteen days of pregnancy. Crucially, this is an exceptionally fragile period due to the fetus's accelerated organogenesis and histogenesis. The maximum dosage, administered orally, was 2000 mg/kg/day. The number of corpus luteum, body weight, or embryo implantation of the pregnant women did not alter as a result of the study, and there were no negative side effects (Mikulska et al., 2023).