Mechanistic Insights into the Neuroprotective Potential of Sacred Ficus Trees

Ficus religiosa (Bo tree or sacred fig) and Ficus benghalensis (Indian banyan) are of immense spiritual and therapeutic importance. Various parts of these trees have been investigated for their antioxidant, antimicrobial, anticonvulsant, antidiabetic, anti-inflammatory, analgesic, hepatoprotective, dermoprotective, and nephroprotective properties. Previous reviews of Ficus mostly discussed traditional usages, photochemistry, and pharmacological activities, though comprehensive reviews of the neuroprotective potential of these Ficus species extracts and/or their important phytocompounds are lacking. The interesting phytocompounds from these trees include many bengalenosides, carotenoids, flavonoids (leucopelargonidin-3-O-β-d-glucopyranoside, leucopelargonidin-3-O-α-l-rhamnopyranoside, lupeol, cetyl behenate, and α-amyrin acetate), flavonols (kaempferol, quercetin, myricetin), leucocyanidin, phytosterols (bergapten, bergaptol, lanosterol, β-sitosterol, stigmasterol), terpenes (α-thujene, α-pinene, β-pinene, α-terpinene, limonene, β-ocimene, β-bourbonene, β-caryophyllene, α-trans-bergamotene, α-copaene, aromadendrene, α-humulene, alloaromadendrene, germacrene, γ-cadinene, and δ-cadinene), and diverse polyphenols (tannin, wax, saponin, leucoanthocyanin), contributing significantly to their pharmacological effects, ranging from antimicrobial action to neuroprotection. This review presents extensive mechanistic insights into the neuroprotective potential, especially important phytochemicals from F. religiosa and F. benghalensis. Owing to the complex pathophysiology of neurodegenerative disorders (NDDs), the currently existing drugs merely alleviate the symptoms. Hence, bioactive compounds with potent neuroprotective effects through a multitarget approach would be of great interest in developing pharmacophores for the treatment of NDDs.


Introduction
Neurodegeneration or neuronal atrophy results from progressive degeneration of neuronal structures and/or functions. It predominantly affects the elderly population as a spectrum of neurodegenerative disorders (NDDs), including Alzheimer's disease (AD), Parkinson's disease (PD), Huntington's disease (HD), and amyotrophic lateral sclerosis (ALS). NDDs lead to a progressive decline in mental (cognition, memory, orientation, attention) and motor movements (gait, orientation, balance). Neurodegeneration affects diverse neuronal circuitries at different locations in the brain. These alterations lead to the loss of dopaminergic neurons (PD), dorsal striatum neurons (HD), both upper and lower motor neurons (ALS), and axonal neuron degeneration (temporal and hippocampal neurons in AD) [1][2][3][4]. Aggregation of proteins and peptides into their pathological forms are the key events in NDDs [5], for example, amyloid beta protein (Aβ) and Tau in AD [6]; mutant huntingtin (mHTT) protein in HD [7]; α-synuclein (αSN) in PD and TAR DNA-binding protein 43  in ALS [8]. Other factors, such as neuroinflammation, oxidative stress, age, kinase (MAPK), extracellular signal-regulated kinase (ERK), and c-Jun N-terminal kinase (JNK) ones. Additionally, the extract also suppresses the activation of nuclear factor kappa B (NF-κB), which strongly supports the neuroprotective role of F. religiosa in several NDDs [36]. Anxiolytic, memory-enhancing, muscle-relaxant, seizure-modifying effect [84] The neuroprotective effect of the methanolic leaf extract of F. religiosa (MEFR) was studied on aluminum chloride (AlCl 3 )-induced neurotoxicity in rats. Aluminum can cross the blood-brain barrier (BBB) through receptor-mediated transfer where it disturbs the oxidative state of the brain and causes neuronal cell death [64,65]. Histological studies have revealed maximum neurodegeneration in the hippocampal CA3 region of the AlCl 3 -treated group. This region has a key role in memory, susceptibility to seizures, and neurodegeneration [66]. However, the group treated with MEFR (200 and 300 mg/kg body weight) presented a significant improvement in the number and quality of neurons [67].
The neuroprotective effect of the petroleum ether (PE) extract of F. religiosa leaves (PEFR) was studied on a 3-nitropropionic acid (3-NP)-induced HD mouse model [68]. Systemic intraperitoneal (i.p.) administration of 3-NP caused striatum neuronal degeneration as seen in HD [69]. Daily oral administration of the PEFR (400 mg/kg body weight) significantly enhanced cognitive and motor activities compared to the untreated group. Biochemical investigations showed that the extract reduced the levels of oxidative stress and inhibited the acetylcholine esterase (AChE) activity. However, significant results were not observed at lower doses of the PEFR. The PEFR was safe up to the dose of 4000 mg/kg. These findings regarding the neuroprotective action suggest that F. religiosa could be used as an effective therapeutic agent in the management of NDDs. The same group of researchers also evaluated the anti-Parkinson's activity of the PEFR in haloperidol-and 6-hydroxydopamine (6-OHDA)-induced experimental rat models [68]. A significant reduction in catalepsy induced by haloperidol was observed in the group pretreated with the PEFR indicating that the extract can protect dopaminergic neurotransmission in the striatum. A substantial increase in locomotor activity was observed at 200 and 400 mg/kg. However, the extract was able to reduce levels of malondialdehyde (MDA) and increase the levels of catalase (CAT), glutathione (GSH), and superoxide dismutase (SOD) at a high dose (400 mg/kg body weight) only, suggesting the antioxidant effect of the extract in the brain of 6-OHDA-treated animals.
The anti-amnesic and nootropic properties of the ethanolic extract of F. religiosa leaves (100 mg/kg body weight) were observed in amnesia and hypoxia induced by scopolamine and sodium nitrite in rodents. The results were comparable to the positive controls Piracetam (200 mg/kg) and Mentat (100 mg/kg) [70]. However, contradictory results were observed [71] after oral supplementation of the F. religiosa leaf extract in healthy mice where the extract considerably decreased the neuromuscular performance and object recognition ability of male mice only. The conflicting results were, perhaps, due to the extract supplementation to healthy animals as compared to disease models, signifying variable effects of the F. religiosa leaf extract in rodents with different health conditions [71].
In addition, the leaves of F. religiosa were also evaluated for the anticonvulsant effect [72], but the hydroethanolic extract failed to exhibit a protective effect in pentylenetetrazol (PTZ)-and maximal electroshock (MES)-induced mouse models [73].

Root
The saponin-rich fraction (SRF) of the hydroethanolic extract of F. religiosa roots (1, 2, and 4 mg/kg) displayed an anticonvulsant effect in mouse models of convulsions [74]. The study was further extended to study the effect of the SRF on cognitive decline and associated depression in a PTZ kindling mouse model of epilepsy [75]. The extract showed marked neuroprotection in the tail suspension test. The SRF considerably raised the monoamine levels and altered the levels of neurotransmitters (noradrenaline, serotonin, γ-aminobutyric acid, dopamine) in the brain. Oral administration of the aqueous root extract of F. religiosa (25,50, and 100 mg/kg) exhibited a dose-dependent and anticonvulsant effect against strychnine-and PTZ-induced seizures. The researchers proposed the involvement of zinc and magnesium present in the extract in the anticonvulsant activity [76].

Fruit
A very high amount of serotonin is known to be present in the fruits of F. religiosa [77]. The role of serotonergic neurotransmission in the protection from seizures by modifying various GABAergic and glutamatergic activities is well-documented [78] and the reductions in brain serotonin levels lead to increased susceptibility to seizures [79]. Anticonvulsant studies with the methanolic fruit extract of F. religiosa (25,50, and 100 mg/kg) showed substantial dose-dependent protection in picrotoxin-and MES-induced convulsion mouse models, with the activity similar (at 100 mg/kg) to that observed in the diazepam-treated group. However, PTZ-induced seizures were not inhibited by the extract [42].
The flavonoid-rich ethyl acetate fraction of the F. religiosa fruit extract (1, 2, and 4 mg/kg i.p.) was used along with a subeffective dose of phenytoin (15 mg/kg) in a PTZ-kindled mouse model. The extract completely suppressed the seizures and reduced the oxidative stress in the brain tissue by decreasing the levels of MDA and increasing the CAT and GSH levels. The extract also decreased the activity of AChE which is responsible for its memory-enhancing effect [75].
The methanolic fruit extract of F. religiosa (10, 50, and 100 mg/kg i.p.) displayed a dose-dependent anti-amnesic effect in a scopolamine-induced amnesia model of mice [80]. Additionally, inhibition of the anti-amnesic effect of the extract by cyproheptadine (a serotonin antagonist) demonstrated the association of serotonergic pathways for memory improvement by the methanolic extract.

Bark
Numerous plants were screened for AChE inhibitory activity in vitro. Among the screened plants, the methanolic extract of the F. religiosa stem bark extract (100-400 µg/mL) displayed the most potent AChE inhibitory activity with an IC 50 value of 73.69 µg/mL [81].

Neuroprotective Effect of Ficus benghalensis
The neuroprotective effect of F. benghalensis extract is summarized in Table 1.

Leaves
Scarce literature is available on the neuroprotective action of F. benghalensis. The methanolic leaf extract of F. benghalensis (200 and 400 mg/kg p.o.) was evaluated for neuroprotective effects against alloxan-induced diabetic neuropathy in rats [82]. The treated animals had better motor coordination in response to stimuli as compared to the disease control group.

Bark
The methanolic F. benghalensis bark extract (100,200, and 300 mg/kg i.p.) displayed a positive anti-amnesic, anti-anxiolytic, and antidepressant effects in a scopolamine-induced behavioral animal model [62]. The phytocompounds isolated from the extract were identified by GC-MS and might interact with glutamatergic, serotonergic, cholinergic, and GABAergic systems in the brain for memory-improving, anxiety-reducing, and depressionresolving activities observed in the study [62]. The aqueous bark extract (150 and 300 mg/kg body weight) had cognitive enhancement activity in scopolamine-induced amnesia in both old and young mice without any toxicity (up to 5 g/kg) [83].

Root
Oral administration of the aqueous root extract (200 mg/kg body weight) displayed anxiolytic, memory-enhancing, muscle-relaxant, and seizure-modifying effects without any toxicity in mice [84]. The phytochemicals present in the extract were suggested to affect muscarinic receptors in the brain.

Neuroprotection by Phytochemicals
A vast library of interesting chemicals has been identified in F. religiosa and F. benghalensis ( Figure 1).

Amyrin
Higher concentration of β-amyrin is present in the B. ceiba leaf extract, which was reported to ameliorate various biochemical parameters (CAT, MDA, AChE) and cognitive functions in rats with scopolamine-induced amnesia [85]. The memory-enhancing effects of αor β-amyrin from the mouse model of scopolamine-induced cognitive impairment involved the activation of extracellular signal-regulated kinase (ERK) and inhibition of glycogen synthase kinase (GSK-3β) in the hippocampus [86]. GSK-3β has a role in tau phosphorylation, which ultimately causes their detachment from microtubules and formation of aggregates [87]; hence, inhibition of GSK-3β is of significance in NDDs. The increased activity of ERK in response to oxidative stress and abnormal phosphorylation has been observed in AD [88]. In addition, both ERK and GSK-3β are known to play an important role in synaptic plasticity and memory processes [89,90]. Moreover, amyrin (25 and 50 mg/kg p.o.) also exhibited anticonvulsant activity by increasing the latency time to 75% and 101% at two doses, probably by inhibiting the protein kinase C (PKC) pathway and by increasing taurine (116% and 76%) and tyrosine (135% and 110%) in the basal ganglia and hippocampus, respectively, and decreasing glutamate (68%), aspartate (65%), and GABA (62%) in the basal ganglia [91]. The PCK pathway negatively regulated the expression of the GABA A receptors by affecting the ion channel function and receptor trafficking [92]. Additionally, amyrin exerted potent anxiolytic and antidepressant effects through the inhibition of monoamine oxidase (MAO) and elevating the GABA levels in the hippocampus [93]. MAO-B is known to have a key role in ROS generation, and its inhibitors (selegiline and rasagiline) are used in PD and AD treatment [94]. Hence, amyrin exerts neuroprotection mainly through enhancing the antioxidant pathway.

Amyrin
Higher concentration of β-amyrin is present in the B. ceiba leaf extract, which was reported to ameliorate various biochemical parameters (CAT, MDA, AChE) and cognitive functions in rats with scopolamine-induced amnesia [85]. The memory-enhancing effects of α-or β-amyrin from the mouse model of scopolamine-induced cognitive impairment involved the activation of extracellular signal-regulated kinase (ERK) and inhibition of glycogen synthase kinase (GSK-3β) in the hippocampus [86]. GSK-3β has a role in tau phosphorylation, which ultimately causes their detachment from microtubules and formation of aggregates [87]; hence, inhibition of GSK-3β is of significance in NDDs. The increased activity of ERK in response to oxidative stress and abnormal phosphorylation has been observed in AD [88]. In addition, both ERK and GSK-3β are known to play an important role in synaptic plasticity and memory processes [89,90]. Moreover, amyrin (25 and 50 mg/kg p.o.) also exhibited anticonvulsant activity by increasing the latency time to 75% and 101% at two doses, probably by inhibiting the protein kinase C (PKC) pathway and by increasing taurine (116% and 76%) and tyrosine (135% and 110%) in the basal ganglia and hippocampus, respectively, and decreasing glutamate (68%), aspartate (65%), and GABA (62%) in the basal ganglia [91]. The PCK pathway negatively regulated the expression of the GABAA receptors by affecting the ion channel function and receptor trafficking [92]. Additionally, amyrin exerted potent anxiolytic and antidepressant effects through the inhibition of monoamine oxidase (MAO) and elevating the GABA levels in the hippocampus [93]. MAO-B is known to have a key role in ROS generation, and its inhibitors

Azelaic Acid
Azelaic acid's neuroprotective potential was evaluated in a rotenone-induced PD model (80 mg/kg p.o.) where a significant reversal in posture, muscular rigidity, and catalepsy was observed after the treatments. Additionally, synergistic effects of azelaic acid with levodopa and carbidopa (100 mg/kg + 25 mg/kg p.o.) [95] were revealed, indicating its promising role in treating PD. Azelaic acid has also been identified as a potential biomarker in urine for personalized healthcare in AD diagnosis [96] as its level correlates negatively with Aβ 42 in cerebrospinal fluid (CSF) and positively with the CSF tau levels. It is also indicative of oxidative damage in the brain of AD patients that may account for changes in brain functions.

Bergapten (5-Methoxypsoralen)
Pieces of evidence support the neuropharmacological effects of bergapten in AD and depression [97,98]. The compound inhibits the AChE and butyrylcholinesterase (BChE) activities in vitro and in silico [99][100][101]. Moreover, bergapten (25 and 50 mg/kg body weight) also improved memory in a scopolamine-induced amnesia model and the cholinergic levels in the hippocampus and the prefrontal cortex without improving motor coordination and locomotor activity [102]. Interestingly, memory improvements were observed after acute and sub-chronic administrations of bergapten, which was independent of the AChE activity and dependent solely on the antioxidant activity of the compound [102]. Additionally, it also exhibited antidepressant effects by inhibiting MAO [103]. Bergapten (25,50, and 100 mg/kg) reversed the paclitaxel-induced neuropathic pain by restoring the levels of oxidative stress markers (GSH, GST, iNOS, LPO) and downregulating the expression of inflammatory mediators (COX-2, TNF-α, NF-κB) [104]. Briefly, neuroprotection by bergapten is exerted mainly through its antioxidant and anti-inflammatory mechanisms. Additionally, higher bioavailability and the potential to cross the BBB make it a favorable candidate for treating neurological diseases [105,106].

Eudesmol
β-Eudesmol is a major phytocompound in Atractylodes lancea rhizome extracts, which induces neurite extensions in PC-12 cells at 100 and 150 µM concentrations by promoting transient phosphorylation of MAPKs (ERK1 and 2). P 38 -MAPK is primarily activated by inflammatory cytokines, playing a critical role in neuronal functions [107]. In addition, β-eudesmol also promotes inositol phosphatase accumulation and encourages the activation of phosphoinositide phospholipase C (PLC), which has a key role in nerve growth factor (NGF)-induced differentiation of neurons [108]. Besides, β-eudesmol (150 µM) induced phosphorylation of the cAMP-responsive element-binding protein (CREB), which is a critical regulator of neuronal plasticity and neuroprotection, in rat pheochromocytoma cells (PC-12) [109].
For the treatment of neurological disorders, low-molecular-weight compounds which can easily cross the BBB are preferred. Being a small molecule, β-eudesmol may prove an encouraging lead compound for studying neuronal functions. In short, β-eudesmol displays a distinctive effect on the nervous system by inducing various pathways that are critical for neuronal growth, plasticity, and protection. Studies on the efficacy of β-eudesmol to cross the BBB are in progress [110].

Eugenol
In traumatic brain injury (TBI) rats, pretreatment with eugenol (25, 50, and 100 mg/kg/day p.o., seven consecutive days) ameliorated the neurochemical and behavioral symptoms. Eugenol decreased lipid peroxidation and improved memory and motor activities in the treated group [111]. Eugenol pretreatment (50 and 100 mg/kg) also mitigated the cerebral ischemia/reperfusion (I/R) damage by inducing autophagy activities through the AMPK/mTOR/P70S6K signaling pathway [112]. AMPK and mTOR are the regulators of autophagy, which works through Unc-51-like kinase 1 (ULK1) activation [113]. Activation of AMPK and inhibition of mTOR endorses the autophagy activities. Additionally, P70S6K (ribosomal protein S6 kinase beta-1) is a downstream kinase of mTOR, whose suppression may promote autophagy activities [114].
In another study, eugenol treatment (10 mg/kg bw for 5 weeks) improved the gait in acrylamide-induced neuropathic rats and restored the levels of antioxidant enzymes and dopamine in the brain [115]. Furthermore, cotreatment with eugenol (6 mg/g) in aluminuminduced toxicity in rat brains reduced the AChE, TNF-α, and caspase-3 expression [116]. Additionally, it promoted neurogenesis in the hippocampus by increasing the expression of the metallothionein gene (MT-III) and restored the levels of brain-derived neurotrophic factor (BDNF) and serotonin in the brain [117]. Since BDNF is vital for the preservation of cortical neurons, its initial loss would lead to short-term memory decline in AD [118]. BDNF expression could be induced by NGF through phosphorylating the CREB in the ERK/AKT signaling pathway [119]. Eugenol (0.1, 1, and 10 mg/kg orally) also displayed neuroprotective properties in a hydroxydopamine-induced PD model [120]. In an in vitro ThT (thioflavin T) assay, eugenol (3 mM) also suppressed amyloid formation by delaying the conversion to the β-sheet form [121].
In another interesting study, eugenol and its analogs were reported to interact with vanilloid receptors in the olfactory bulb and displayed a positive effect on memory [122]. Such a possibility can be explored for the treatment of AD and PD.
Eugenol appears to be a wonder molecule because of its ability to cross the BBB [116] and displays the neuroprotective effect through neurogenesis, antioxidant, anti-amyloid, and antiapoptotic ability by affecting multiple pathways. This multitarget approach expands the application of eugenol to multiple neurological diseases.

Kaempferol
Kaempferol is a multipotential neuroprotective agent that affects various pathways in NDDs [123]. In neurological diseases, the antioxidant activity of kaempferol inhibits various metalloproteases (MMP-2, MMP-3, MMP-9) and protects the BDNF modulations responsible for neuronal plasticity. It is known for enhancing cognitive performance in animal models by displaying anti-AChE activity [124,125]. Kaempferol and its degradation products inhibit Aβ oligomerization and plaque formation by interacting with the Aβ protein without involving the Lys 16 and Lys 18 residues [126] and restores the levels of oxidative stress markers (SOD, glutathione, ROS) both in vitro and in vivo [127]. It also displays its neuroprotective potential at 20 mg/kg dose by suppressing microglial activation by inhibiting the NF-κB, MAPK, AKT, and toll-like receptor 4 (TLR4) pathways in an LPSinduced striatum injury mouse model [128].
In a rotenone-induced PD rat model, kaempferol (10 and 20 mg/kg) increased the SOD and glutathione peroxidase (GSH-P X ) levels and decreased lipid peroxidation. It neutralized ROS by blocking apoptosis through the JNK/MAPK pathway [129]. It also reduced the expression of proinflammatory cytokines, COX-2, and the high mobility group box 1 (HMGB1)/TLR4 inflammatory pathway [128]. As a result, motor coordination and dopamine levels increased in the PD model [130]. In summary, kaempferol displays multitarget properties (antioxidant, anti-inflammatory, anti-amyloid, antiapoptotic, and modulating enzymes involved in neurotransmission) for neuroprotection. Moreover, due to its ability to cross the BBB [131], it can prove beneficial in the treatment of NDDs.

Lanosterol
In vitro and in silico studies have indicated that lanosterol can suppress the buildup of misfolded protein aggregations/sequestosomes [132,133] by promoting autophagy activities [134]. Lanosterol (0.5 mM) also protected dopaminergic neurons from 1-methyl-4-phenylpyridinium (MPP+)-induced cell death in a PD cellular model by inducing mild mitochondrial depolarization and promoting autophagy activities. The observed reallocation of lanosterol synthase to mitochondria suggested that lanosterol might exert its survival effect by regulating mitochondrial functions [135]. In addition, reduced levels of lanosterol were observed in the striatum and ventral midbrain regions from 1-methyl-4phenyl-1,2,3,6-tetrahydropyridine (MPTP)-treated mice (PD model), suggesting a potential role in the cholesterol metabolism of NDD pathogenesis. In short, lanosterol promotes autophagy activities by sequestering misfolded proteins.

Linalool
Linalool has shown a protective effect in various neurodegenerative models and is reported to have the ability to cross the BBB [155]. In in vitro experiments, linalool (10 µg/mL) protected PC-12 cells from Aβ exposure by reducing ROS and inhibiting the activity of proapoptotic caspase-3 [156]. Linalool (50 and 100 mg/kg/day for 21 days) also significantly suppressed Aβ-induced ROS, oxidative stresses, and inflammatory responses in an AD fly model [157] without altering the amount of Aβ in the brain. Alterations in the hippocampal phospholipid profiles in ischemic animals were mitigated by linalool, which helped to maintain the phospholipid homeostasis, hence recovering brain functions [158]. The oral administration of linalool (25 mg/kg for 3 months) in a triple transgenic AD mouse model (3xTg-AD) restored memory in the treated animals via reducing β-amyloidosis, astrogliosis, and tauopathy besides reducing proinflammatory markers (MAPK, inducible oxide nitric synthase (iNOS), COX-2, IL-1β) [159]. Linalool (100 µM) is known to modulate glutamatergic neurotransmission by interacting with NMDA receptors [160]. Furthermore, it proved to be neuroprotective in a glutamate-induced oxidative stress in vitro model (HT-22 cells) by reducing ROS, calcium production, and LPO levels [161]. Linalool (162, 324, 648 µM) exerts protective effects in LPS-induced BV2 microglial cells by Nrf2 activation [162] and in mice with Aβ-induced cognitive deficits by restoring the levels of oxidative stress-related enzymes, suppressing caspase-3, and upregulating the Nrf2 and HO-1 expression [163]. Together, these findings suggest that linalool could be used for the development of NDD drugs as it can cross the BBB and provide neuroprotection through anti-inflammatory, antioxidant, and antiapoptotic properties.
Hence, lupeol seems to be a potent candidate for NDD drug discovery as it has an acceptable ADMET profile and the multitarget neuroprotective approach (inhibiting neuroinflammation, reducing oxidative stress, repressing apoptosis and microglial activations).

Myo-Inositol (Vitamin B8)
Even though no direct study has been conducted to investigate the effect of myo-Inositol on the NDD model systematically, it displays a neuroprotective role in ischemic stroke injury in animals exposed to tobacco smoke and in streptozotocin-induced mice by increasing the motor functions after the stroke using in situ brain perfusion and the acute brain slice method at 0.1 µCi/mL [171,172]. In addition, myo-inositol (30 mg/kg for 28 days) ameliorated spatial learning and memory deficits by attenuating cell loss in the hippocampus in a kainic acid-induced epilepsy rat model [173] by a multitarget approach including preserving neuronal circuits and activation of GABA signaling. Myo-inositol also presented the anticonvulsive property in thiosemicarbazide models of seizures by increasing the latent time and decreasing the severity of the seizure [174].
Interestingly, myo-inositol has been considered a noninvasive early marker for assessing various asymptomatic AD stages in comparison with magnetic resonance spectroscopy (MRS) [175]. Increased myo-inositol/creatine levels have been observed in healthy apolipoprotein E E4 genotype (APOE ε4) carriers with normal CSF Aβ42 levels in comparison to ε4 non-carriers, signifying the importance of myo-inositol levels in assessing AD before a noticeable amyloid pathology [176]. Reports suggest that myo-inositol can also be transported across the BBB by simple diffusion as well as through a stereospecific transporter [177].

Myricetin
Myricetin is a lipophilic compound with the ability to cross biological membranes, including the BBB [178]. It is reported to target AD by inhibiting multiple pathways, such as neuroinflammation, autophagy, oxidative stress, chelation, anti-AchE, and Aβ depositions [179]. Myricetin (5 µM) averted Aβ 1-42 oligomer-induced neurotoxicity in human neuroblastoma cells (SH-SY5Y) by exerting antioxidant effects on the cell membranes and mitochondria. Myricetin also restored mitochondrial dysfunctions by decreasing ROS, increasing the expression of manganese superoxide dismutase (Mn-SOD) and ATP generation [180]. The JNK/stress-activated protein kinase (SAPK) pathway is activated by oxidative stress and Aβ. Enhanced BACE-1 levels lead to an increase in Aβ levels [181], which eventually activates the JNK/SAPK pathway, resulting in a vicious cycle of the NDD. Furthermore, oxidative stress could also endorse serine/threonine protein phosphatase (PP2A) inhibition, promoting tau phosphorylation [182] and damaging the mitochondria [183]. Myricetin inhibited the BACE-1 activity (IC 50 , 2.8 µM) to cleave the amyloid precursor protein (APP), increase α-secretase, and decrease Aβ production/oligomerization [184,185]. Additionally, myricetin also interferes with the NF-κB and AMPK/SIRT1 signaling pathway and reduces the levels of inflammatory mediators (IL, TNF-α, iNOS, COX-2) in the brain [186]. Myricetin (1 and 10 µM) also aids in the removal of abnormal Aβ and tau through autophagy activation by inhibiting phosphorylation of mammalian targets of rapamycin (mTOR) in primary neuron cultures [187]. Interestingly, myricetin (25 µM) can regulate the levels of metal ions in the brain to reduce their interactions with Aβ and disassemble the formation of the metal-Aβ complex [188]. Since it was previously reported that Fe 2+ could activate microglia by increasing neuroinflammation, the complexation of myricetin with Fe 2+ could reduce the inflammatory processes and inhibit the expression of transferrin receptor I (TrP1), thus lowering the iron levels [6]. Lastly, myricetin also improves learning and cognition through the anti-AchE activity. Some proinflammatory cytokines (IL-1) activate AchE causing the Ach levels to decrease in the brain, affecting memory. Hence, the anti-inflammatory activity of myricetin also improves memory [189,190].

Pinene
Pinene, a monoterpene, can cross the BBB and affect multiple neurotransmitter systems, such as adrenergic, cholinergic, dopaminergic, GABAergic, serotoninergic, and noradrenergic functions in the brain [191]. According to a cell-based study, pretreatment of α-pinene (10 and 25 µM) in PC-12 cells inhibited ROS by increasing the expression of antioxidant enzymes and reducing apoptosis by decreasing the caspase-3 activity [192]. α-Pinene administration (50 mg/kg i.p.) in the Aβ-induced rat model reduced neuroinflammation by overturning the TNF-α/NF-κB pathway and improved memory and learning. Moreover, α-pinene also upregulated the expression of both the nicotinic acetylcholine receptor (nAChR) α7 subunit and BDNF [193], which play a role in the survival and maintenance of neurons. Additionally, α-pinene (100 mg/kg i.p.) restored the levels of antioxidant enzymes (SOD, CAT, GPX) and reduced NO, IL-6, and MDA in the brain of focal ischemic stroke model rats [194]. Furthermore, α-pinene (100 mg/kg i.p.) downregulated Bax with a corresponding upregulation of Bcl-2 expression, resulting in suppression of apoptosis in a rat model of cerebral ischemia-reperfusion [195].
In a mouse model of memory impairment, α-pinene upregulated the expression of AchE, muscarinic receptors, and antioxidant transcription factors in the hippocampus, improving spatial recognition and memory [196]. In summary, the antioxidant, antiinflammatory, and antiapoptotic properties of α-pinene would help treat NDDs.

Psoralen
Psoralen is a natural furanocoumarin with in vitro competitive inhibitory activity against AchE (IC 50 , 370 µg/mL) [197]. A molecular docking study revealed a stable AchE-psoralen complex with π-π stacking (Tyr 334 ) and hydrogen bonding (Gly 119 and Gly 118 ) interactions [197]. Psoralen also displayed inhibitory activities towards MAO-A (IC 50 , 15.2 µM; noncompetitive inhibition) and MAO-B (IC 50 , 61.8 µM; competitive inhibition) in the rat brain [198]. Since MAO is an important enzyme in maintaining levels of monoamine neurotransmitters in the brain, its reduced expression results in decreased Aβ depositions and oxidative stress [199]. In another study, psoralen and iso-psoralen-rich extracts (0.1 and 0.3 mg/kg) of Psoraleae fructus improved amnesia in scopolamine-induced rats, apparently by AchE inhibition (IC 50 , 1.12 mM) and activation of cholinergic neuronal functions [200]. In summary, even though moderate enzyme inhibitory activity has been observed with psoralen, it could serve as a lead molecule to synthesize other potential analogs to treat NDDs.
In essence, quercetin presents a multitarget tactic (anti-inflammatory action, antioxidant action, and enzyme inhibition) to ameliorate the symptoms of NDDs.

Rhein
Rhein (12 mg/kg) displayed antioxidative and neuroprotective effects in controlled cortical impact (CCI) rats by reducing the MDA levels and increasing SOD, CAT, and GSH [163], thereby protecting the brain from oxidative damage. It also protected the BBB through the NADPH oxidase/ROS/ERK/MMP-9 signaling pathway [217]. Additionally, in an I/R rat model, rhein (50 and 100 mg/kg/day for 3 days) restored the levels of antioxidant markers, enhanced Bcl-2, and decreased the levels of Bax and caspase-3 [218]. Rhein (100 mg/kg i.p.) also exerted an anti-inflammatory response by downregulating the inflammatory cytokines in the cortex of animals with TBI [219]. Additionally, rhein (10 mg/kg i.v.) improved the cognitive decline in an APP/PS1 mouse model of AD by activating the SIRT1/PGC-1α pathway, improving oxidative stress by regulating mitochondrial biogenesis [220]. It also exerted an anti-inflammatory response by subsiding the levels of TNF-α and IL-1β in the hippocampus and an antioxidant property by decreasing MDA, increasing GSH, the GSH/GSSG ratio, CAT, and GSH-Px in the same model [220]. The pharmacokinetics of rhein disclose that it can be easily transported by body fluids and can pass through damaged BBB which is very useful in case of brain injury [221]. In summary, rhein could be an important compound for neuronal protection through antioxidant, anti-inflammatory, and antiapoptotic properties.

Rutin
Rutin has emerged as an important pharmacological flavonoid in various NDDs [222] as it can easily cross the BBB [223]. Rutin improves memory by reducing Aβ oligomerization, oxidative stress, neurotoxicity, and neuroinflammation in several AD animal models [224][225][226][227]. The anti-amyloidogenic property could be due to the destabilization of Aβ by direct interaction of the aromatic ring of rutin with the hydrophobic β-sheet of amyloid aggregates. Rutin (100 µL rutin suspension/per 10 g bw for 30 days) decreased the tau levels by regulating phosphorylation through increased PP2A activities. It also reduced inflammation by downregulating NF-κB besides securing neuronal morphology and improving cognition through synapse preservation in the brain of the Tau-P301S mouse model [228]. In addition to the AD models, rutin (25 mg/kg bw orally for 3 weeks) protected dopaminergic neurons by reducing oxidative stress and apoptosis in the 6-OHDA-induced rat model of PD [229,230]. It also improved HD symptoms by activating autophagy and the insulin/insulin-like growth factor I (IGF-1) pathway at 15-120 µM concentrations [231]. APP phosphorylation is suppressed by insulin/IGF-I in vitro which favors the non-amyloidogenic pathway [232]. This pathway regulates neurogenesis and is known to improve neuronal survival, learning, and memory through the PI3K/Akt and ERK pathway [233][234][235].
In brief, rutin provides neuroprotection through its antioxidant, anti-inflammatory, anti-amyloid, and autophagic activation properties.

Stigmasterol
Stigmasterol, a phytosterol, demonstrated its neuroprotective effects through multitarget approach in vitro and in vivo. Studies revealed that phytosterols can cross the BBB through the scavenger receptor class B member 1 (SR-BI)-dependent pathway and via apolipoprotein E (ApoE) [236], improving the cognition [237]. It reduced amyloid plaque formations [238], inhibited the AchE activity (IC 50 , 644 µM) in vitro [239], and decreased the elevated ROS [240]. Stigmasterol (1 µM) displayed its neuroprotective effects in hydrogen peroxide-induced oxidative stresses in SH-SY5Y cells by modulating the sirtuin 1-forkhead box O3a (SIRT1-FoxO3a) pathway [241]. SIRT1 activates FoxO3a, which consequently stimulates the production of antioxidant enzymes (SOD, CAT), hence protecting against oxidative stress. Stigmasterol also exhibited anti-inflammatory activity in the IL-1β-treated cells by inhibiting proinflammatory cytokines without affecting the levels of anti-inflammatory cytokines suggesting its role in the NF-κB inflammatory pathway [242]. In a recent report, stigmasterol (10 and 20 µM) inhibited NF-κB and NOD-like receptor thermal protein domain-associated protein 3 (NLRP3) signaling by activating AMPK, thereby reducing the Aβ-induced inflammatory response in BV2 cells [243]. Stigmasterol (3, 10, and 30 mg/kg) also exerted ameliorating effects on the scopolamine-induced memory loss in mice through the cholinergic neurotransmission augmentation by N-methyl-D-aspartate receptor (NMDA) activation [244]. Earlier reports had shown progressive modulatory effects of Ach by binding to muscarinic receptors on NMDA, enabling NMDA receptor-mediated synaptic plasticity and long-term potentiation (LTP), which are implicated in learning and memory [245]. Besides, stigmasterol (10 mg/kg p.o.) has been reported to increase the ERK and CREB phosphorylation in the hippocampus of the scopolamine-induced memory loss model in mice, which could also influence a positive effect on memory and learning [244,246].

Synephrine
Synephrine is a sympathomimetic alkaloid with mild CNS stimulant properties [247]. When the effects of oral supplementation of synephrine (20 mg) were evaluated for cognition and exercise performances during a pre-workout, no significant results were observed on muscular endurance during exercise, but it seemed to improve cognitive functions and mental focus [248]. Synephrine has effective inhibitory properties towards AchE (IC 50 , 226.01 nM) and BchE (IC 50 , 92.22 nM) in vitro [249].

β-Sitosterol
β-Sitosterol displayed robust anti-AchE (55 µg/mL) and anti-BchE (50 µg/mL) activity both in vitro and in silico. The in vivo results confirmed that β-sitosterol acts as a free radical scavenger and can reach the brain and inhibit AchE and BACE [255,256]. Its antioxidant effects (15 µM) were observed in a glucose oxidase (GOX)-induced oxidative stress and lipid peroxidation model of HT22 hippocampal cells through the estrogen receptor (ER)-mediated PI3K/GSK-3β signaling pathway [257]. β-Sitosterol seemed to help the PI3K recruitment to the lipid raft, an important region of the membrane in signal transduction [258]. GSK-3β is an important downstream target of PI3K upregulation which eventually increases the intracellular glutathione, a natural antioxidant [259]. Additionally, β-sitosterol augmented the mitochondrial membrane potential (∆Ψm) and adenosine triphosphate (ATP) by integrating into the mitochondrial membrane [257].
In NDDs, neurons are damaged due to persistent neuroinflammation. In a study, β-sitosterol (8 and 16 µM) displayed anti-inflammatory properties in lipopolysaccharide (LPS)-induced BV2 microglial cells by reducing the expression of proinflammatory factors (IL-6, iNOS, TNF-α, cyclooxygenase-2 (COX-2), an inhibitor of nuclear factor kappa B: IκB, NF-κB, ERK/p38) [260]. In animal models, β-sitosterol exhibited positive effects on learning and memory [255,261,262], and prevented plaque deposition in an amyloid protein precursor/presenilin 1 (APP/PS1) model [255,261,262]. Recently, substantial anxiolytic and antidepressant effects of γ-sitosterol were observed from the strong binding affinity with the human serotonin receptor in a molecular docking study [263]. In short, the neuroprotective effects of β-sitosterol would be due to its antioxidant, anti-inflammatory, anti-amyloid, and enzyme inhibition properties. It seems to be a potential candidate for managing memory-related disorders in NDDs. Pan-assay interference structures (PAINS) are chemical compounds which could give a significant false positive signal in the drug screening processes including redox reactivity, fluorescence of small compounds, and covalent changes of target proteins [264,265]. For instance, curcumin, known as the representative PAINS, has failed more than 120 clinical studies for various diseases due to false activity in vitro and in vivo [266]. Moreover, at least 15 studies have been retracted and dozens more have been corrected since 2009 [267]. As a result, it is imperative for accurate assessment of these compounds with structural alerts and elimination from further steps of the drug discovery process [268]. Medicinal chemistry analysis of PAINS was processed by the SwissADME server [269]. Except for myricetin, quercetin, rhein, and rutin, all the phytocompounds derived from Ficus trees were predicted to have no structural alerts or false positive signals ( Table 2).

Conclusions and Future Directions
Even though plant extracts have been used for centuries for treating a spectrum of diseases in traditional medicine, no scientific validation for their therapeutic effects has been presented. In the modern era, the use of plants for therapeutic purposes was underestimated initially. Recently, after the discovery and use of numerous important drugs from herbal sources [270], such as quinine (an antimalarial drug from the Cinchona bark), atropine (anticholinergic medicine from Belladona), digoxin (obtained from Digitalis to treat cardiac arrhythmia), colchicine (extracted from Colchicum for treating gout), and galantamine (AChE inhibitor from Galanthus spp.), the interest in plant-based research Major phytochemicals act as inhibitors against oxidative stress and inflammation pathways. Oxidative stress is modulated by triggering antioxidant enzymes or transcription factors and suppressing ROS-mediated proteins. The cascade of inflammation is hampered by the downregulation of inflammatory transcription factors and caspase-related components. Abbreviations: CAT, catalase; GPx, glutathione peroxidase; GR, glutathione reductase; GSH, glutathione; GSSG, oxidized glutathione; GST, glutathione S-transferase; SOD, superoxide dismutase; ROS, reactive oxygen species; MDA, malondialdehyde; PKC, protein kinase C; HMGB1, high mobility group box 1; TLR4, toll-like receptor 4; Bcl-2, anti-apoptotic B cell lymphoma-2; Bax, Bcl-2-associated X; NF-κB; nuclear factor kappa B; MAPKs, mitogen-activated protein kinases; PI3K, phosphoinositol-3 phosphate; AKT, protein kinase B; MMP, matrix metalloproteinase; COX-2, cyclooxygenase 2; TNF-α, tumor necrosis factor α; sMAF, small musculoaponeurotic fibrosarcoma protein; Nrf2, nuclear E2-related factor 2; ARE, antioxidant response element; HO-1, heme oxygenase-1.

Conclusions and Future Directions
Even though plant extracts have been used for centuries for treating a spectrum of diseases in traditional medicine, no scientific validation for their therapeutic effects has been presented. In the modern era, the use of plants for therapeutic purposes was underestimated initially. Recently, after the discovery and use of numerous important drugs from herbal sources [270], such as quinine (an antimalarial drug from the Cinchona bark), atropine (anticholinergic medicine from Belladona), digoxin (obtained from Digitalis to treat cardiac arrhythmia), colchicine (extracted from Colchicum for treating gout), and galantamine (AChE inhibitor from Galanthus spp.), the interest in plant-based research has been expanding to determine their mechanistic actions behind the therapeutic potentials and has reached a new height. In an extract, a cocktail of various bioactive compounds may exhibit synergistic effects for better therapeutic activity, questioning whether a purified compound would present similar effects. However, an understanding of a purified bioactive compound should be performed first, even if a cocktail of compounds may be needed down the line. In addition, since the pathophysiology of NDDs would be complex, linking multiple cellular events, a multitarget tactic might be a better approach going forward.
Several bioactive compounds in the sacred Ficus species display neuroprotective properties in vitro and in vivo through multiple pathways, such as antioxidant (PI3K, AKT, AMPK, PKC, ERK, HO-1, Nrf2), anti-inflammatory (SIRT1, NF-κB), anti-amyloid (APP, BACE-1), antiapoptotic (Bcl-2, Bax, caspase), and modulating enzymes in neurotransmission (AChE, BChE). Most of the studies were carried out using rodent models of NDDs that unfortunately cannot recapitulate the complete aspects of AD as it is a uniquely human disease [271]. This is one of the reasons why many drugs were unsuccessful in clinical trials. Therefore, performing research using human tissues would give a more human-centric strategy.
Additionally, the negative results of the anti-Aβ strategy (amyloid cascade hypothesis) in clinical trials on AD patients demonstrate that it is not the only pathogenic factor involved. Pulling down cerebral Aβ can only delay cognitive decline but it cannot stop it, indicating the role of other etiological factors (oligomer cascade hypothesis) in the pathogenesis of AD [272]. Hence, scientists must follow a multitarget approach, leading to the treatment of complex diseases, such as NDDs.
In conclusion, Ficus spp. extracts and bioactive compounds present effective neuroprotective properties (in vivo and in vitro) by modulating several important pathways. The results from in vivo experiments also indicate the nontoxic nature of the extracts/phytocompounds at the doses tested. Yet, they have not been translated into clinical trials. Henceforth, the need to take the research to next level is of great significance in the treatment of NDDs.