Stimulatory Effects of Melatonin on Porcine In Vitro Maturation Are Mediated by MT2 Receptor

Abstract

Melatonin is a multifunctional molecule with numerous biological activities. The fact that melatonin modulates the functions of porcine granulosa cells via the MT2 receptor suggests the possibility of MT2 receptor-mediation for melatonin to promote cumulus expansion of porcine cumulus-oocyte complexes (COCs). Therefore, we investigated the presence of MT2 in porcine COCs, and the effects of melatonin with or without selective MT2 antagonists (luzindole and 4-P-PDOT) on this process; COCs underwent in vitro maturation culturing with six different conditions (control, melatonin, luzindole, 4-P-PDOT, melatonin + luzindole or melatonin + 4-P-PDOT). Cumulus expansion, oocyte nuclear maturation, and subsequent embryo development after parthenogenetic activation (PA) were evaluated. In experiment 1, MT2 was expressed in both oocytes and cumulus cells. In experiment 2, melatonin significantly increased the proportion of complete cumulus expansion (degree 4), which was inhibited by simultaneous addition of either luzindole or 4-P-PDOT. A similar pattern was observed in the expression of genes related to cumulus expansion, apoptosis, and MT2. In experiment 3, no significant difference was observed in immature, degenerate, and MII oocyte rates among the groups. In experiment 4, melatonin significantly increased blastocyst formation rates and total blastocyst cell numbers after PA, but these effects were abolished when either luzindole or 4-P-PDOT was added concomitantly. In conclusion, our results indicate that the MT2 receptor mediated the stimulatory effects of melatonin on porcine cumulus expansion and subsequent embryo development.

Melatonin-Nrf2 Signaling Activates Peroxisomal Activities in Porcine Cumulus Cell-Oocyte Complexes

“…This result is supported by a study by Ma and Lin et al, who demonstrated a significant decrease in the maturation of mouse oocytes and embryonic development following brusatol treatment [17,46]. In addition, our results are similar to those that were obtained in previous studies [9,10,40] in that melatonin not only increased subsequent embryo development, but also increased the maturation rates of porcine COCs. Moreover, the expression levels of GDF9 and BMP15, which are related to oocyte competence [47], were also significantly influenced in each experimental group.…”

“…Figure 8 shows a schematic of this study. our study shows that melatonin can improve porcine oocyte quality and subsequent embryonic development through its various biological activities [10,40,41]. Melatonin is thought to mediate its effect by binding to receptors on the membrane, i.e., MTs [42], which constitute a superfamily of guanine nucleotide binding protein (G-protein)-coupled receptors [43,44].…”

“…Melatonin is thought to mediate its effect by binding to receptors on the membrane, i.e., MTs [42], which constitute a superfamily of guanine nucleotide binding protein (G-protein)-coupled receptors [43,44]. Specifically, MT2 was found to be fundamentally involved in oocyte maturation, embryo development, and the activation of antioxidant-related signaling in porcine oocytes and embryos [8,40]; accordingly, our results showed that the regulation of MT2 resulted in differences in mRNA and protein levels among the treatment groups. As demonstrated in many studies, melatonin is known to regulate the Nrf2/ARE signaling pathway [8,11,12]; therefore, we employed brusatol, a Nrf2 inhibitor, in order to verify the mechanism.…”

Effect of Melatonin on the In Vitro Maturation of Porcine Oocytes, Development of Parthenogenetically Activated Embryos, and Expression of Genes Related to the Oocyte Developmental Capability

“…Melatonin treatment improves cumulus oophorus expansion of porcine COCs during IVM [24], and also increases the first polar body extrusion rate of the oocytes disrupted by heat stress and mono-(2-ethylhexyl) phthalate exposed in the porcine [7,25]. Melatonin participates in modulating functions of granulosa cell through melatonin receptor 2 (MT2), and also promotes cumulus expansion of COCs via MT2 in pigs [26]. Therefore, melatonin treatment can enhance cumulus oophorus expansion and the first polar body extrusion rate of oocytes through melatonin receptors, which is beneficial for nuclear and cytoplasmic maturation of porcine oocytes during IVM.…”

Importance of Melatonin in Assisted Reproductive Technology and Ovarian Aging

“…Melatonin membrane receptors (MT1, MT2) are located in oocytes and granulosa cells (cumulus cells) [58][59][60][61]; it would be of great interest to determine the intracellular signaling pathway by which melatonin promotes oocyte maturation and embryo development. While some reports have examined these intracellular processes of melatonin in oocytes [62][63][64][65], the details remain unclear. It has been reported that melatonin controls the expression of genes related to oocyte maturation including mitochondrial function [53,60,66,67], antioxidative enzymes [53,59,67,68], apoptosis [52,[65][66][67][68][69], cumulus cell expansion [51,61,70], and oocyte maturation factors [61,67].…”