The synthesis of ZnO NPs and Zn(cur)O NPs was conducted according to the methodology described by Moussawi et al. [26]. Before evaluating their antidepressant effects, a thorough examination of the physical and chemical properties of the nanoparticles was performed due to their significant impact on their physiological behavior [33]. The morphology and dimensions of the synthesized particles were examined using various techniques, including transmission electron microscopy (TEM), scanning electron microscopy (SEM), and dynamic light scattering (DLS). Analysis by SEM indicated that ZnO NPs exhibited a spherical shape, whereas Zn(cur)O NPs had a rod-like appearance. TEM images revealed that both ZnO NPs and Zn(cur)O NPs possessed hexagonal shapes, with average diameters of (23.6 ± 1.46) nm and (23.03 ± 5.2) nm, respectively. However, the average sizes determined by DLS were (220 ± 11.74) nm for ZnO NPs and (342 ± 22) nm for Zn(cur)O NPs. This discrepancy in size values between the two methods can be attributed to the fact that DLS measurements encompass the particle size along with its solvation shell, whereas TEM provides size measurements of individual particles excluding the solvation shell [34]. Particle stability was assessed using zeta potential (ZP), which indicates the tendency of particles to aggregate or remain dispersed. The ZP values of ZnO NPs and Zn(cur)O NPs were determined to be (-19.6 ± 4.5) mV and (-25.6 ± 4.61) mV, respectively [35]. These results indicate low stability for ZnO NPs and a high level of stability for Zn(cur)O NPs. The enhanced stability of Zn(cur)O NPs can be attributed to the strong interaction between Curcumin and ZnO, as described by Moussawi et al. [36].
The blood-brain barrier (BBB) serves as a highly selective barrier that prevents the entry of external substances, including pharmaceuticals, into the brain [37]. However, nanoparticles offer a potential solution as drug delivery systems to overcome this obstacle. ZnO nanostructures, upon oral absorption, can either enter the brain through neural transport or breach the BBB. Among the proteins found on the surface of ZnO-NPs, apolipoprotein E plays a crucial role in facilitating the passage of nanoparticles through the BBB [38]. In this study, the antidepressant effects of Curcumin, ZnO NPs, and Zn(cur)O NPs were evaluated using an animal model of depression induced by reserpine. Reserpine was chosen to induce depression as it reduces motor activity and depletes monoamines, which are the primary neurotransmitters involved in mood regulation [37]. Monoamine dysfunction and neuroinflammation are recognized as major contributors to depression, as reported by Yuanzhen et al. (2019) and Belmaker et al. (2008) [39, 40]. Additionally, disrupted motor activity has been associated with deficiencies in neurobiological processes [39, 40].
FST is a common test used to show the development of depression in animal models. The observed decrease in motor activity is indicated by increased immobility time and decreased time of struggling and swimming in the FST. Moreover, the OFT data revealed a decrease in crossed squares, rearing, and grooming following 15 days of reserpine injections daily. The time when the animals are immobilized and unable to get out of the water correlates with depressive-like actions [42].
The present decrease in 5-HT, NE, and DA are due to reserpine's irreversible inhibitory impact on monoamine vesicular absorption. These findings are similar to the study of Ikram et al., who observed that acute treatment of Wistar rats with a low dose of reserpine-induced motor dysfunction and a substantial reduction in DA, NE, and 5-HT in the brain. Recent studies indicate that repeated reserpine administration may be utilized as a practical and cost-effective animal model for the progression of depression for antidepressant compound validation [43].
Curcumin (diferuloylmethane) has a wide range of pharmacological actions, such as monoamine replenishment [44], anti-inflammatory properties [45], and strong antioxidant function. Turmeric, a yellow curry spice was, contains polyphenol curcumin, which has been utilized in traditional Indian dishes and herbal treatments for a long time [46]. In the FST, immobility reflects the desperate behavior of rats, and it is used as a sign of depression's onset. Furthermore, a wide range of clinically effective antidepressant medicines may help to reduce immobility. This behavioral method has shown to be a useful tool for identifying new antidepressants [47]. The current evidence clearly shows that Curcumin reduces the immobility time and increases the serotonin level in the hippocampus and striatum. This might be due to the inhibitory result of Curcumin on indoleamine 2,3-dioxygenase, the enzyme responsible for the degradation of the serotonin precursor, tryptophan, which increases serotonin. Curcumin's antioxidant properties, communicated through MDA levels and enhanced catalase activity, may also play a role in its antidepressant efficacy, particularly as these effects are associated with increased motor activity [48].
ZnO NPs may improve behavioral and cognitive dysfunction in mice with depressive-like behavior via increasing synaptic plasticity in neurons and the area between the perforant pathway and the dentate gyrus [49].In contrast, other studies showed that mice treated with ZnO NPs exhibited a modification of brain monoamines, ions, and histological impairment [50].
In the present study, after injecting rats with reserpine for 15 days, they were treated daily with Curcumin, ZnO NPs, and Zn(cur)O NPs for ten days, which increased the motor activity of animals. Raising and grooming increased this; however, only ZnO NPs raised rearing to the control level. Moreover, only Curcumin and ZnO NPs increased grooming to control. Also, in the FST, Struggling and swimming times increased significantly when rats were treated with Curcumin and or Zn(cur)O NPS to that of the ZnO NPs exhibited values above the control. Using Curcumin, Zn(cur)O NPs, and ZnO NPs were more effective at decreasing immobility time.
Increased levels of reactive oxygen species (ROS) can lead to lipid peroxidation, resulting in the formation of malondialdehyde (MDA) and damage to cell membrane lipids. Clinical studies have shown that individuals with severe depression exhibit higher MDA levels in their blood compared to control groups [51]. Furthermore, research has indicated that MDA levels are elevated during depression and that antidepressant therapy can reduce lipid peroxidation levels [52, 53]. Glutathione (GSH), an endogenous antioxidant, plays a crucial role in DNA repair and regulates various metabolic processes [54]. Individuals with depression have been found to have lower GSH levels [55, 56]. Catalase (CAT), an important antioxidant enzyme, converts hydrogen peroxide into oxygen and water. Clinical trials have demonstrated that catalase activity decreases when excessive oxidative stress leads to catalase deficiency, resulting in increased levels of hydrogen peroxide and ROS in the body [57]. However, some studies have shown an increase in catalase activity during acute periods of depression [58–60].
According to the findings of this study, all three treatment formulations improved the oxidative stress status, as evidenced by changes in MDA, CAT, and GSH levels, in the cortex, hippocampus, and striatum. Treatment with ZnO NPs and Zn(cur)O NPs significantly reduced MDA levels in the brain compared to the control group. Additionally, all treatment formulations significantly increased GSH levels compared to the control. In the striatum, all three treatments significantly decreased MDA levels compared to the animal model of depression. Notably, Zn(cur)O NPs led to a substantial increase in GSH levels and significantly increased CAT activity in the hippocampus compared to the control group.
Monoamine neurotransmitters play a crucial role in brain neurochemistry. Norepinephrine (NE) is involved in controlling behavior, attention, prefrontal cortex activity, memory processing, and behavior [61, 62]. The original catecholamine theory of major depression suggests that depletion of NE at key synapses contributes to the development of depression through changes in the central nervous system [63]. Serotonin (5-HT) and its receptors have a modulatory effect on almost all brain functions, and dysregulation of the serotonergic system has been implicated in the pathogenesis of various psychological and neurological disorders [64, 65]. Dopamine (DA) is a neurotransmitter that regulates reward, motivation, working memory, and attention [66]. In the context of the interaction theory between depression and impaired DA transmission, compensatory upregulation of D2 receptor density in the basal ganglia/cerebellum has been observed in depressed patients compared to healthy individuals [67]. These findings highlight the significance of monoamine neurotransmitters in the etiology of depression and emphasize the role of NE, 5-HT, and DA in regulating various cognitive and emotional processes in the brain.
Significant improvements in monoamine levels were observed in the cortex, hippocampus, and striatum following three different therapies. Among them, Zn(cur)O exhibited a promising effect by significantly increasing norepinephrine (NE) levels above the control group in the cortex. In the hippocampus, all three treatments effectively increased serotonin (5-HT) levels compared to the depression model. However, treatment with Zn(cur)O NPs demonstrated superior efficacy in elevating 5-HT levels to the control group. Similarly, in the striatum, Zn(cur)O NPs significantly increased 5-HT levels above the control values. These findings indicate that Zn(cur)O has a more pronounced influence on the restoration of reserpine-induced depletion of monoamines compared to free Curcumin or ZnO NPs therapy. The selective impact of Zn(cur)O NPs can be attributed to several factors. Curcumin exerts its antidepressant properties by inhibiting monoamine oxidase activity, leading to elevated levels of the monoamine transmitters 5-HT and DA. The rise in 5-HT levels is associated with increased expression of the 5-HT receptor 1A mRNA. Additionally, Curcumin exhibits anti-inflammatory properties by inhibiting enzymes involved in oxidative stress and apoptosis, including nuclear factor kappa-light-chain-enhancer of activated B cells (NF-kB), tumor necrosis factor-alpha (TNFα), and interleukin 6 (IL-6).
Furthermore, Curcumin has demonstrated antidepressant effects by increasing the levels of serotonin (5-HT), dopamine (DA), and norepinephrine (NE) [68]. Moreover, the utilization of ZnO nanoparticles (ZnO NPs) as a potential formulation for antidepressant purposes enhances the consistency, efficacy, and delivery of Curcumin. Zinc (Zn) plays a crucial role in safeguarding the blood-brain barrier (BBB) against oxidative stress caused by free radicals and is essential for the synthesis of coenzymes involved in the metabolism of biogenic amines, thereby maintaining brain homeostasis and preventing neurological disorders [69]. Zinc is known to regulate calcium reflux and prevent oxidative stress. Zinc deficiency can lead to the activation of nitric oxide synthase (NOS), mitochondrial dysfunction, and oxidative stress due to calcium influx [70]. Consequently, the antidepressant effects of ZnO NPs may be potentiated through their conjugation with Curcumin, thereby offering a synergistic therapeutic approach.
The data presented in this study demonstrates that the effectiveness of native Curcumin in restoring oxidative stress and neurochemical changes associated with depression in the rat brain is limited. This ineffectiveness can be attributed to the low bioavailability of Curcumin due to its poor water solubility. However, when Curcumin is combined with ZnO nanoparticles (ZnO NPs), the pharmacokinetics of Curcumin are enhanced, resulting in increased availability. This improved bioavailability may account for the remarkable antioxidant activity observed with Zn(cur)O NPs compared to Curcumin and ZnO alone. Furthermore, the augmented bioavailability of Curcumin through its conjugation with ZnO NPs enhances its inhibitory effects on monoamine oxidase, an enzyme responsible for the degradation of monoamines. This dual action leads to an elevation in monoamine levels and a reduction in the production of free radicals generated during monoamine oxidative catabolism.
Finally, the conjugation of Curcumin with ZnO NPs offers a promising strategy to enhance its therapeutic efficacy by addressing the challenge of low bioavailability due to limited water solubility. This approach capitalizes on the potent antioxidant and monoamine-modulating properties of Curcumin, thus holding potential for the treatment of depression. The study findings provide valuable insights into the role of Zn(cur)O NPs as a novel therapeutic avenue for the management of depressive disorders.