Melatonin as a multifunctional modulator: emerging insights into its role in health, reproductive efficiency, and productive performance in livestock

Melatonin has significant potential in alleviating microbial dysbiosis (Gao et al., 2021). Its effects are mainly mediated through toll-like receptor 4 (TLR4), which plays a crucial role in pathogen-associated molecular pattern (PAMP) signaling, especially in relation to lipopolysaccharides (LPS) found in Gram-negative bacteria (Kim et al., 2020). Melatonin’s presence within the GIT is remarkable, with concentrations exceeding those in the pineal gland by up to 400-fold, reflecting the abundance of melatonin receptors and the enzymes required for its production in the gut (Kennaway, 2023). Furthermore, gut microbes exhibit circadian rhythms that mirror those of the host, significantly influencing their metabolic functions (Zheng et al., 2024). Given melatonin’s critical role in regulating the biological clock, it becomes evident that the circadian rhythms of gut microbes and their functions are intricately tied to melatonin.

The identification of rhythmic patterns in the ruminant gut microbiome has prompted additional research, suggesting that melatonin’s effects might stem from its presence in saliva. Salivary melatonin, known for its role in regulating inflammation, promoting antioxidant responses, and accelerating the healing of oral wounds (Elsherbini and Ezzat, 2020), exhibits circadian rhythms similar to those found in ruminal fluid and muscularis (Ouyang et al., 2021). This implies that melatonin present in saliva might affect microbial communities across the GIT through circadian variations. Indeed, the rhythmic changes in rumen microbial populations correspond with fluctuations in melatonin levels, with elevated melatonin associated with a higher relative abundance of the families Preovotellaceae and Muribaculaceae, and a reduction in Succininivibrionaceae and Veillonellaceae (Fu et al., 2023). These findings exhibit melatonin’s capacity to impact Gram-negative bacteria through cytokine production and metabolic regulation (Xue et al., 2023). Fluctuations in melatonin levels within the GIT affect crucial metabolic pathways, thereby impacting the predominant phyla in the rumen, such as Firmicutes, Proteobacteria, and Bacteroidetes (Fu et al., 2023).

Melatonin has recently emerged as a promising regulator of the reproductive tract microbiome in livestock. The reproductive microbiome plays a vital role in fertility and overall reproductive efficiency (Hussain et al., 2021). In cows, melatonin supplementation has been shown to modulate bacterial populations within the uterus and vagina, fostering a more favorable environment for reproduction. This effect is attributed to melatonin’s potent anti-inflammatory and antioxidant properties, which may create optimal conditions for embryo development and implantation (Messman et al., 2021).

Beyond its influence on microbial composition, melatonin also impacts the immune response within the reproductive tract, which is intricately linked to the microbiome (Wang et al., 2021b). By enhancing immune function, melatonin indirectly supports a balanced microbiome, further promoting reproductive success. This dual role underscores the complex interplay between melatonin, the microbiome, and immune health, all of which are crucial for successful reproduction in livestock. Interestingly, melatonin’s effects on the reproductive tract microbiome vary across livestock species (Cosso et al., 2021). Factors such as physiological state, age, and environmental conditions likely contribute to these variations. Understanding these complexities is essential for developing tailored melatonin supplementation strategies that maximize reproductive health benefits across diverse livestock populations.

Melatonin is a potent antioxidant with the unique ability to dissolve in both water and fats, enabling it to function in various environments, such as within cell interiors, body fluids, membranes, and organelles (Zarezadeh et al., 2022). It outperforms conventional antioxidants like vitamins E and C in mitigating oxidative stress. Unlike typical antioxidants, which neutralize only one or a few reactive oxygen species (ROS), melatonin can neutralize up to 10 ROS molecules (Marchena et al., 2020). This remarkable capacity is attributed to its ability to stimulate key antioxidant enzymes, including SOD, CAT, GPx, and GSH, particularly when given in doses between 0.1 and 20 mg/kg per day (Table 3) (Baydas et al., 2002; Cruz et al., 2009; Nayki et al., 2016; Taghizadieh et al., 2016; Olayaki et al., 2018; Aranarochana et al., 2021; Bashandy et al., 2021; Colares et al., 2022; Abdel-Razek et al., 2023; Essawy et al., 2023).

Melatonin not only increases the mRNA levels of these antioxidant enzymes but also inhibits pro-oxidant enzymes such as nitric oxide synthase (Monteiro et al., 2024). By altering membrane fluidity, melatonin safeguards cell membranes from oxidative harm and eliminates free radicals before they damage lipids and proteins, all without promoting pro-oxidant effects (Kopustinskiene and Bernatoniene, 2021). It acts as an effective scavenger of free radicals and an electron donor, neutralizing various reactive species like hydroxyl radicals, hydrogen peroxide, and nitric oxide (Ahmadi and Ashrafizadeh, 2020). When melatonin interacts with hydroxyl radicals, it produces 3-hydroxymelatonin, which is later excreted through urine. Its metabolites, such as AMK and AFMK, display even greater antioxidant potency (Galano and Reiter, 2018; Zarezadeh et al., 2022).

Recent discoveries highlight melatonin’s pivotal role in activating the antioxidant response element (ARE) system, which triggers the transcription of numerous antioxidant proteins and enzymes to neutralize ROS and support essential protein transport (Vriend and Reiter, 2015). Central to this process is the Nuclear Factor Erythroid 2–Related Factor 2 (Nrf2)-ARE signaling pathway, which significantly enhances the activity of key antioxidant enzymes. This pathway provides a protective mechanism that shields livestock from various diseases, helping to preserve their productive potential (Zhang W. et al., 2018; Dong et al., 2020).

Under oxidative stress, melatonin elevates cellular Nrf2 levels, which is crucial for its antioxidant function. Melatonin facilitates the nuclear translocation of the Nrf2 transcription factor and promotes its interaction with ARE, thereby amplifying the expression of antioxidant enzymes (Figure 2) (Deng et al., 2016). Further, melatonin has been demonstrated to reduce stress-related damage by safeguarding the hippocampus through the regulation of the Nrf2/Heme Oxygenase-1 (HO-1) pathway. Although research is still expanding, melatonin is also thought to increase the concentration of inhibitor of nuclear factor kappa B (IκB) alpha, an inhibitor of NF-κB, suggesting an additional mechanism by which it regulates inflammatory responses (Rajput et al., 2017).

Moreover, melatonin reduces the levels of nitrites, inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2), and microsomal PG E synthase-1 (mPGES-1), while also preventing the translocation of NF-κB in peritoneal macrophages. By lowering pro-inflammatory mediators and enhancing HO-1 expression through NF-κB, p38 mitogen-activated protein kinase (MAPK), and Nrf2 signaling, melatonin shows significant potential as a therapeutic agent for conditions involving macrophage over-activation (Deng et al., 2020).

Melatonin acts as a safeguard against the detrimental effects of various chemical compounds that induce oxidative stress. For instance, dizocilpine maleate (MK-801), a compound known to induce oxidative damage in the prefrontal cortex, leads to psychotic symptoms, but melatonin can counteract these effects. Similarly, cadmium negatively impacts male reproductive health; however, melatonin alleviates this toxicity by lowering MDA levels, boosting SOD activity, increasing GSH, and elevating pro-inflammatory cytokines like TNF-alpha and IL-1 beta (Venditti et al., 2021).

Melatonin also protects the cerebellum from damage caused by acrylamide by reducing LPO, boosting antioxidant enzyme activities, and lessening DNA damage (Ozturk et al., 2023). The cerebellum, which is highly susceptible to chemical insults, often undergoes neuron loss and organ shrinkage during development. Lead (Pb) is another toxicant that causes oxidative stress and neurotoxicity; however, melatonin (at 10 mg/kg) has been shown to reduce LPO and protect the cerebellum from Pb-induced toxicity (Bazrgar et al., 2015). Additionally, melatonin exhibits neuroprotective properties against ethanol toxicity in the cerebellum and lowers plasma homocysteine (Hcy) levels, further strengthening its role as a neuroprotective agent (Bagheri et al., 2024).

Animals are exposed to various forms of radiation, against which melatonin provides protection. For instance, tropical animals frequently encounter ultraviolet (UV) radiation. When these animals were treated with melatonin after UV exposure, there was a marked increase in the activity of antioxidant enzymes such as SOD, CAT, and GPx. This enhanced enzyme activity neutralized free radicals, reducing the damage caused by UV radiation. Although UV radiation can indirectly harm spleen tissue, melatonin treatment aided in restoring balance and preventing splenocyte apoptosis, thus maintaining organ function (Goswami and Haldar, 2014). Melatonin also functions as an antioxidant in testicles exposed to microwaves, helping to alleviate oxidative stress and reduce DNA fragmentation (Özgen et al., 2023).

Follicular atresia significantly affects bovine reproductive performance, as it heavily depends on the health of GCs for ovarian follicle development. Disturbances in these cells, whether through apoptosis, autophagy, cell cycle arrest, or accumulation of ROS, can lead to follicular atresia (Wang H. et al., 2018; Ma et al., 2019; Wang Y.-X. et al., 2021). Additionally, alterations in steroid hormone synthesis further influence GC function. Melatonin, known for its ROS-scavenging properties and cellular regulatory abilities, plays a critical role in mitigating follicular atresia by reducing ROS levels and inhibiting apoptosis in GCs through various pathways (Xie et al., 2022).

In the initial phases of follicular atresia, apoptosis predominantly targets the inner layer of GCs, while the cumulus-oocyte complex and outer GCs remain largely unaffected, highlighting the selective nature of this process (Rajin et al., 2022). GCs are crucial for supporting and maintaining follicle growth, and their physiological condition greatly influences the fate of the follicle (Yuan et al., 2019). Mitochondria, as a primary source of ROS, contribute to mitochondrial swelling and apoptosis when ROS levels become excessive. Melatonin helps maintain antioxidant enzyme activity and neutralizes reactive oxygen by regulating ER oxidoreductin 1 (ERO1) and enhancing the activities of antioxidant enzymes (Fernández et al., 2015; Tamura et al., 2020).

Melatonin has been found to mitigate oxidative stress and apoptosis in bovine ovarian GCs caused by β-zearalenol (Yang et al., 2019). The protective effects of melatonin are modulated by its receptors, MT1 and MT2, as inhibition of these receptors can diminish melatonin’s benefits and interfere with cell cycle regulation (Gobbi and Comai, 2019). Melatonin’s influence also varies with environmental conditions such as temperature and oxygen concentration. For example, at 37.5°C and 5% O₂, low concentrations of melatonin promote cell proliferation, while at 40°C, higher concentrations have the same effect (Zeebaree et al., 2018). This temperature-dependent response highlights melatonin’s potential in mitigating heat stress. Nonetheless, differences in physiological conditions and natural melatonin production make the reliable and effective use of supplemental melatonin challenging, highlighting the need for comprehensive data to guide evidence-based practices.

Melatonin supports the development of bovine secondary follicles through membrane-bound receptors, whereas its antagonist, luzindole, inhibits these effects and decreases the expression of antioxidant enzymes in cultured follicles (Paulino et al., 2022). Additionally, melatonin stimulates follicular angiogenesis by increasing VEGF expression, which is crucial for follicular development (Tao et al., 2021). In theca cells, which exclusively express the MT2 receptor, melatonin inhibits androgen biosynthesis and slows ovarian atresia and aging by reducing apoptosis and regulating cell proliferation through the PI3K/Akt pathway (Wang S. et al., 2018; Ma et al., 2023). Abnormal melatonin levels in theca interna cells have also been linked to the development of follicular cysts in sows (Qin et al., 2022).

Beyond these roles, melatonin’s antioxidant properties support oocyte quality and enhance subsequent embryonic development, underscoring its potential in assisted reproduction. Studies have identified MT1 and MT2 mRNA in porcine cumulus-oocyte complexes, revealing that melatonin modulates GC function through MT2, which enhances cumulus expansion and embryonic development (He et al., 2016; Lee et al., 2018). Melatonin is critical for preventing age-related defects in germline-soma communication and aids in the transfer of antioxidant molecules from cumulus cells, preserving oocyte quality (Zhang H. et al., 2022).

Melatonin also shows promise as a pharmacological agent against endocrine disruptors such as Bisphenol A (BPA) and Bisphenol S (BPS), which impair follicular growth and steroidogenesis. It mitigates these harmful effects by increasing estradiol production, promoting GC proliferation, and protecting against mitochondrial apoptosis during oocyte maturation (Park et al., 2018; Wu et al., 2018; Berni et al., 2019). Its protective effects extend to other toxic substances, such as Aflatoxin B1 (AFB1), which induces follicular atresia and oxidative stress. Melatonin’s antioxidant properties and ability to inhibit apoptosis provide effective protection against AFB1-induced toxicity (Cheng et al., 2019).

Furthermore, melatonin enhances results in ovarian tissue cryopreservation and is vital for reproductive processes and blastocyst implantation in various mammalian species. It achieves this by regulating apoptotic mechanisms, enhancing adhesion protein expression, and protecting against oxidative stress that compromises embryo quality and pregnancy success (Najafi et al., 2023). The detection of melatonin in follicular fluid, along with reduced levels in cases of polycystic ovary syndrome (PCOS), further emphasizes its essential role in ovarian function and oocyte maturation (Shi et al., 2009).

The quality of oocytes is essential for reproductive success in female animals and is a key factor in ruminant embryo transfer. Fresh embryos generally lead to significantly higher live birth rates compared to those that have been cryopreserved (Insogna et al., 2021). Cryopreservation of oocytes has long been challenging due to issues with survival, fertilization, and developmental rates (Chen et al., 2003). To overcome these challenges, enhancing the quality of oocytes cultured in vitro before cryopreservation is crucial, and melatonin has proven to be a promising approach.

Studies show that melatonin can notably improve the developmental potential of oocytes, both in vitro and in vivo (Sananmuang et al., 2020). In cattle, melatonin improves oocyte developmental competence and embryonic growth by reducing ROS levels (Figure 4) (Gutiérrez-Añez et al., 2021). It also mitigates ROS in heat-stressed oocytes, increases maturation rates, boosts the proportion of embryos developing into blastocysts, and upregulates genes related to mitochondrial function (Yaacobi-Artzi et al., 2020). Melatonin also safeguards bovine oocytes from damage inflicted by harmful agents like paraquat, thus preserving their developmental potential (Pang et al., 2019).

Substantial evidence underscores melatonin’s role in advancing oocyte development in bovines. It boosts the production of antioxidant enzymes via specific membrane and nuclear receptors, aiding in the removal of ROS. In cumulus-oocyte complexes, acetylserotonin O-methyltransferase (ASMT) may play a role in melatonin synthesis (El-Raey et al., 2011). By reducing oxidative stress via the MT1 membrane receptor, melatonin preserves spindle body function, a critical factor for oocyte development.

In cattle, administering melatonin from days 190–262 of gestation enhanced uterine blood flow, probably due to its impact on steroid metabolism (Brockus et al., 2016b). Melatonin also positively influenced estradiol metabolism, improving utero-placental development (Lemley and Vonnahme, 2017). During estrus synchronization and AI in cattle, external melatonin significantly elevated P₄ levels, boosted antioxidant enzyme activity, and reduced MDA concentrations in the blood, resulting in a marked improvement in pregnancy rates (Guo et al., 2021).

A functional melatonergic system is essential for luteal function in mammals (Wang et al., 2021a). ROS, primarily generated from normal metabolic processes, are involved in both luteogenesis and luteolysis (Mierzejewski et al., 2023). During cholesterol transport for P₄ synthesis and in the regression phase, LPO can damage the luteal plasma membrane, leading to impaired luteal function (Taketani et al., 2011; Cruz et al., 2014; Xu et al., 2021; Al-Shahat et al., 2022). Melatonin safeguards luteinizing GCs in the ovulatory follicle from ROS, boosts P₄ production after ovulation, and helps prevent premature luteolysis in the newly developed CL. Elevated indolamine levels during the luteal phase underscore melatonin’s direct involvement in these processes. High concentrations of melatonin synthesis enzymes and receptor expression suggest that this site may serve as a key area for hormone synthesis and regulation (Xiao et al., 2018).

Melatonin deficiency in small ruminants disrupts follicular and luteal dynamics, resulting in decreased P₄ synthesis (Kárpáti et al., 2023). In these animals, melatonin enhances P₄ secretion by regulating autophagy through the AMPK/mTOR pathway (Duan et al., 2024). In equine corpus lutea, both MT1 receptor mRNA and protein are present. Melatonin reduces P₄ production and P450scc expression in a dose-dependent manner. This inhibition can be reversed by luzindole, a non-selective melatonin receptor antagonist, indicating that functional melatonin receptors are present in luteal cells (Pedreros et al., 2011).

Melatonin’s role in luteal function has been evaluated during early pregnancy, given the endocrine organ’s importance in the initial stages of gestation (Verteramo et al., 2022). In heat-stressed cows, melatonin improves luteal hemodynamics (Abdelnaby and Abo El-Maaty, 2021). Local melatonin synthesis in the luteal cells of pregnant animals suggests a paracrine or autocrine role (Zhang et al., 2022b). In these animals, luteal cells during pregnancy show hormone receptor expression and increased P₄ levels that correlate with melatonin concentration, mediated by upregulated P450scc and steroidogenic acute regulatory (StAR) protein (Zhang Y. et al., 2018). Moreover, melatonin stimulates GnRH and LH production in the luteal cells of pregnant animals, indicating a regulatory role through these hormones (Zhang et al., 2022c). Administering melatonin boosts the expression of genes related to pregnenolone synthesis, supports the development of the CL, aids in embryonic implantation, and enhances uterine receptivity during early pregnancy.