Cadmium (Cd) is a heavy metal that lacks a physiological function in the human body and is classified as a carcinogen [1]. It poses a health risk for both humans and animals [2, 3]. Prolonged exposure to Cd is associated with an elevated risk of different health issues, including kidney damage, osteoporosis, diabetes, cardiovascular disease, and several types of cancer [1, 2]. Furthermore, experimental studies have linked Cd exposure to cognitive deficits and Alzheimer’s disease (AD) [1, 4].

Alzheimer’s disease is a progressive neurodegenerative disorder that significantly affects cognitive function and behavior, with memory impairment being the most common initial symptom [2, 5]. The exact mechanisms underlying the pathogenesis of AD are still unclear. Numerous hypotheses have been proposed to explain its development, among these hypotheses are amyloid-beta (Aβ) accumulation, tau protein abnormalities, damage to cholinergic neurons, as well as oxidative stress and neuroinflammation [6–7]. Environmental factors, such as heavy metals, are considered important influencing factors in AD [2, 8]. However, their role in the pathogenesis of AD remains complex and not fully understood, despite their established neurotoxic effects. Cadmium is particularly concerning due to its harmful impact even at low levels of exposure. Given the widespread exposure of the general population to this metal, epidemiological studies have highlighted its association with the development of AD, underscoring the necessity of investigating this potent neurotoxin's contribution to the etiology of AD [8].

For its neurotoxicity, Cd enters the central nervous system (CNS) through two primary routes: oral ingestion, which can damage the blood-brain barrier (BBB), and inhalation, which bypasses the olfactory neuroepithelium and olfactory bulb [2]. Once in the CNS, Cd accumulates in neurons and exerts its neurotoxic effects through various mechanisms, one of which is oxidative stress. Cd triggers the generation of reactive oxygen species (ROS), leading to oxidative damage of proteins, lipids, and DNA in brain tissues. This oxidative stress is a key factor in Alzheimer’s pathogenesis, as it promotes the aggregation of Aβ peptides [9]. Cadmium also interferes with tau protein, another critical pathological feature in AD, by accelerating its hyperphosphorylation. Hyperphosphorylated tau leads to the formation of neurofibrillary tangles, which further impair neuronal signaling and contribute to cognitive decline [10]. Beyond oxidative stress, Cd disrupts calcium homeostasis, which is essential for proper neuronal communication and synaptic plasticity. Cadmium mimics calcium (Ca2⁺), entering neurons through Ca2⁺ channels and disturbing intracellular Ca2⁺ levels. This dysregulation contributes to mitochondrial dysfunction, reducing the energy supply for neurons and triggering apoptosis [11]. Additionally, cadmium is associated with chronic neuroinflammation, another significant contributor to AD. Cd activates microglia, eliciting the expression of proinflammatory cytokines, which exacerbate neuronal damage. Chronic neuroinflammation leads to a self-perpetuating cycle of oxidative stress and neuronal injury, accelerating the progression of AD [12].

Medicinal plants have garnered heightened interest due to their abundant bioactive compounds with significant antioxidants, anti-inflammatory, and neuroprotective properties. These natural compounds may counteract the neurotoxic effects of cadmium, thereby offering promising protection against Alzheimer’s disease [13]. Their multifaceted mechanisms position them as valuable candidates for preventing or mitigating heavy metal-induced neurodegeneration. The Coast Coral tree, scientifically known as Erythrina caffra Thunberg, a member of the Fabaceae family, is well recognized for its historical use in traditional medicine. It's rich in bioactive flavonoids and alkaloids, exhibits potent antioxidants, anti-inflammatory, and neuroprotective properties [14]. However, its efficacy against heavy metal-induced neurotoxicity leading to cognitive decline and neurodegeneration remains insufficiently explored. In the present study, we aimed to evaluate whether intracerebroventricular (ICV) administration of cadmium chloride (CdCl₂) recapitulates key pathological features implicated in Alzheimer’s disease, with a specific focus on cholinergic dysfunction, oxidative stress, and neuroinflammation. Additionally, we assessed the therapeutic potential of the ethanolic extract of Erythrina caffra (E. caffra) seeds in mitigating cadmium-induced AD-like alterations in Wistar rats.

44 adult male Wistar rats, with an average weight of 309. 54 ± 4.36g and an age of 3 months were used in this study. To ensure their well-being, the rats were maintained in individual pens under standard conditions (Temperature of 24ºC and a 12-hour light/dark cycle). They were supplied with unrestricted availability of standard water and food. All experimental procedures utilized in the present study followed the guidelines outlined in the National Institute of Health’s (NIH) Guide for the Care and Use of Laboratory Animals.

Prior to the surgery, animals were deeply anesthetized (7% chloral hydrate, i.p injection). Following anesthesia, the heads of rats were shaved, and they were securely fixed in a stereotaxic apparatus. The injection site was determined based on the rat brain atlas, ensuring it fell within the established normative range. Specifically, injections were targeted at coordinates averaging 0.8 mm in the anteroposterior direction, 1.5 mm in the medial-lateral direction, and 3.5 mm in the dorsoventral direction.

The rats were randomly distributed into four groups (n = 11). The sham animals received an intracerebroventricular administration of sterile saline (5 µL/ventricle) using a Hamilton syringe. The disease group received a bilateral ICV injection of CdCl₂ (10 µg/kg; 5 µL/ventricle) (Sigma, St. Louis, MO, USA). The treated groups received similar CdCl₂ injections; one group was subsequently treated orally with the ethanolic extract of E. caffra seeds (via gavage) at a dose of 2.5 mg/kg, while the second treated group received oral memantine at a dose of 20 mg/kg, serving as the positive control group.

At the completion of the study, all rats underwent behavioral testing to evaluate memory performance. Thereafter, all rats were fasted overnight, weighed, anesthetized, sacrificed by decapitation, and the brain tissues were collected for histological and neurochemical analyses. These assessments focused on the hippocampus and included quantification of acetylcholine levels, determination of acetylcholinesterase activity, and evaluation of oxidative stress and neuroinflammatory markers (Fig. 1).

To assess spatial learning and memory, a MWM test was employed. The apparatus comprised a water tank (40 cm in depth and 120 cm in diameter). The tank was partitioned into four points: North, South, West, and East. Within each quadrant, a visual signal was positioned on the outer surface of the labyrinth wall, serving as a reference point for the rats. An 11x14 cm platform was concealed beneath the water's surface, precisely 2cm deep, and positioned at the center of one of the four virtual quadrants. The platform remained stationary throughout the trials. During the training session, a total of five consecutive trials were conducted. In each trial, the rats were introduced into the tank, confronting the wall, starting from different positions, and given a maximum of 60 seconds to locate and reach the concealed platform. If a rat successfully reached the platform, it remained there for 10 seconds. If a rat failed to locate the platform within the allotted time, it was gently directed there [18]. The rat’s performance during the test period was recorded. The time it took the rat to locate the platform was measured.

E. caffra is one of the most used species of the genus Erythrina due to its biological activities and therapeutic potential [14, 32]. Previous studies have reported the antioxidant properties, anti-inflammatory effects, and the acetylcholinesterase inhibitory potential of alkaloids derived from E. caffra, which modulate key molecular pathways involved in AD and contribute to memory enhancement [33, 34].

The cholinergic system plays a central role in memory processes, and in AD, cholinergic transmission is impaired, leading to cognitive decline [33]. The results of the current study showed that administering CdCl₂ via the ICV route resulted in significant changes in the cognitive abilities of rats, causing alterations in memory. These changes were accompanied by increased oxidative stress, neuroinflammation, and neuronal loss in the hippocampus, an essential region for memory and one of the brain areas affected by Alzheimer’s [35]. Whereas daily administration of the ethanolic extract of E. caffra seeds significantly impacted cognitive changes, biochemical alterations, and histological abnormalities attributable to cadmium exposure. These findings indicate that cadmium induced AD-like pathological changes and highlight the potential therapeutic effects of the E. caffra seeds extract in attenuating these pathological manifestations of AD.

The neurotoxic effects of Cd are well documented, as it is linked to cognitive decline and the onset of neurodegenerative disorders such as AD [8]. The findings from the NOR and MWM tests indicate that CdCl₂ significantly impairs the memory of experimental rats. Previous studies have demonstrated that Cd is associated with memory deficits in various animal models [36, 37]. The mechanisms underlying memory deficits following Cd exposure include dysregulation of the cholinergic system, oxidative stress, and neuroinflammation.

The cholinergic system dysfunction in the ICV-CdCl₂ animals is consistent with research indicating that exposure to Cd alters AChE activity and disrupts ACh levels. These changes directly impact synaptic transmission and cognitive functions, resulting in cognitive deficits observed in AD-like conditions [8, 38]. Cd is known to disrupt the Ca²⁺ balance in neurons. It interferes with calcium signaling pathways, which are crucial for neurotransmitter release and neuronal excitability. This disruption leads to impairment in the normal influx of Ca²⁺ necessary for the exocytosis of acetylcholine-containing vesicles [39]. More specifically, Cd enters neurons via dihydropyridine-sensitive (L-type) voltage-gated Ca²⁺ channels, where it competes with Ca²⁺ within the channel pore and blocks Ca²⁺ influx at physiological voltages. Consequently, this alteration in the normal influx of Ca²⁺ impairs ACh release [4]. Cd also directly affects cholinergic receptors, inducing alterations in cholinergic muscarinic receptors and AChE variants. Specifically, Cd disrupts M1 and M3 muscarinic receptors, leading to the overexpression of AChE-S and the downregulation of AChE-R. This dysregulation contributes to the degeneration of cholinergic neurons and impairs cholinergic transmission [4, 39]. From the above, CdCl₂ negatively impacts the cholinergic system, leading to elevated AChE activity. AChE is known to interact with amyloid precursor protein and presenilin-1, which leads to the production of Aβ. This interaction contributes to the accumulation of Aβ plaques, an essential hallmark of AD pathology [40].

The enhancement in memory in the treated group with E. caffra extract was associated with increased hippocampal ACh levels, alongside reduced AChE activity in this region. In parallel with these neurochemical and cognitive improvements, histological examination further confirmed the neuroprotective effect of the E. caffra. The hippocampal subfields exhibited preserved neuronal morphology, characterized by intact pyramidal and granule cell layers with minimal signs of degeneration. These findings strongly suggest that the extract not only modulates cholinergic transmission but also protects hippocampal neurons from CdCl₂-induced neurodegeneration. These results are consistent with prior research, particularly the cholinergic hypothesis of AD, which posits that memory impairment in AD is associated with dysfunction of the cholinergic system due to increased AChE activity [41, 42] and decreased ACh level and availability in the synaptic cleft. ACh is a crucial neurotransmitter that plays an essential role in memory function [43]. These observations suggest that therapeutic interventions targeting AChE activity in the brain may be vital for mitigating cognitive decline associated with AD. Previous research on Erythrina alkaloids has indicated their potential as AChE inhibitors [33, 34]. Specifically, alkaloids from the genus Erythrina, including erythraline, erythrinine, and cristanine A, have shown promising inhibitory potential against human acetylcholinesterase, as confirmed through in vitro testing [44]. These alkaloids bind strongly within the active site of the enzyme, forming salt bridges with key residues like Trp86 and Tyr337, and engaging in hydrophobic interactions that stabilize their binding. This interaction prevents the enzyme from hydrolyzing ACh, thereby reducing its activity and alleviating symptoms of neurodegenerative diseases [44]. Furthermore, another study conducted by Gelfuso et al. [45] on the effects of Erythrina-derived alkaloids on memory in a rat model of epilepsy aligns with these findings. This study demonstrated that 11α-hydroxy-erythravine and erythravine alkaloids isolated from Erythrina mulungu exert neuroprotective effects on the hippocampus primarily by modulating cholinergic transmission through their antagonistic action on nicotinic ACh receptors (nAChRs). By inhibiting ACh-activated currents in cells expressing nAChRs, these alkaloids help regulate neuronal excitability and prevent neuronal damage. This neuroprotection preserves the structural integrity of hippocampal neurons, reducing cell death. Additionally, because nAChRs are critical for cognitive functions, including learning and memory, their modulation by the alkaloids results in enhanced cognitive performance. The preservation of hippocampal neurons and the stabilization of cholinergic signaling collectively contribute to improved memory and learning capabilities [45].

The imbalance in oxidative stress markers in the hippocampus of CdCl₂ animals was reduced in treated animals with E. caffra and memantine, suggesting that increased oxidative stress may play a crucial role in AD pathogenesis and that the extract may have beneficial effects on brain oxidative stress. Gella and Durany [46] demonstrated that the imbalance between ROS production and antioxidant defense, which leads to oxidative stress, plays a major role in neurodegeneration. Previous studies are consistent with our findings regarding the beneficial effects of Erythrina caffra on brain oxidative stress. These studies demonstrated that Erythrina caffra exhibits strong antioxidant activities [11, 47]. Notably, the essential oil extracted from E. caffra has been shown to decrease oxidative stress markers while enhancing antioxidant enzyme levels, including CAT and SOD [47]. Furthermore, various species within the genus Erythrina, including Erythrina indica, Erythrina senegalensis, and Erythrina × neillii, have demonstrated antioxidant activities, where extracts from these plants restored the activities of CAT, SOD, and GSH [32].

Neuroinflammation has been reported to contribute to AD [48]. In the present study, the direct injection of Cd in the brain caused an elevation in TNF-𝛼 and IL-6 levels, which are among the primary proinflammatory cytokines activated in the CNS during neuroinflammatory responses. While the extract of E. caffra modulates the alteration in the level of these proinflammatory cytokines, indicating that the extract exerts immunomodulatory activity and offers neuroprotection against inflammatory processes associated with neurodegeneration caused by cadmium. Previous studies have shown that Cd can induce neuroinflammation by mechanisms involving BBB leakage, microglia activation, and infiltration of immune cells into the brain [48, 49]. Whereas Erythrina species, including E. caffra, modulate neuroinflammation primarily through their bioactive substances, which exhibit potent antioxidant and anti-inflammatory properties [32]. These compounds inhibit key inflammatory pathways by suppressing proinflammatory cytokines, as well as downregulating the NF-κB signaling cascade. Additionally, certain alkaloids in Erythrina act on nAChRs, engaging the cholinergic anti-inflammatory pathway, which further reduces microglial activation and neuroinflammation. Together, these mechanisms contribute to the neuroprotective effects of Erythrina against inflammation-induced neurodegeneration [32, 50].

In conclusion, our research highlights the neurotoxic potential of cadmium in inducing Alzheimer-like pathology. The present findings demonstrate that intracerebroventricular administration of CdCl₂ in rats replicates pathological features of AD, including increased oxidative stress, disruption of cholinergic transmission, neuroinflammation, hippocampal degeneration, and memory impairments. On the other hand, the ethanolic extract of Erythrina caffra seeds demonstrated neuroprotective effects. The daily administration of this extract over two months reduced oxidative stress by enhancing antioxidant defenses, modulating cholinergic neurotransmission and neuroinflammation, and preserving the structural integrity of hippocampal neurons. These combined effects contributed to the observed improvement in memory. These findings suggest that the ethanolic extract of Erythrina caffra seeds may represent a promising therapeutic potential in mitigating key pathological features of Alzheimer’s disease.