The aim of the present study was to improve cytoplasmic maturation of porcine oocytes by the addition of lycopene into in vitro maturation (IVM) media. We designed six experimental groups; IVM medium was supplemented with 10 IU/ml FSH, FSH and 10 IU/ml human chorionic gonadotrophin (hCG), or FSH and 7 μm lycopene in the first half of the IVM culture (0-22 h) followed by further culture (22-44 h) with or without hCG. The addition of lycopene into IVM media delayed the interruption of communication between an oocyte and the cumulus cells. Although meiotic competence was similar among the six groups, the glutathione level of matured oocytes was significantly higher in the lycopene-supplemented group (9.89 pmol per oocyte) than that in other groups (7.25 and 7.81 pmol per oocyte). Fertilization rate was significantly improved in lycopene-supplemented groups (58.3%) more than that in the group supplemented with FSH only (43.1%), whereas there were no differences in developmental competence among the groups (blastocyst rate: 20.1-29.5%). These results indicate that insufficient cytoplasmic maturation during conventional IVM resulted by disconnection of the gap junction between an oocyte and the cumulus cells in the early phase during IVM culture. We concluded that lycopene induced a prolonged sustainment of gap junctional communication between an oocyte and the cumulus cells during porcine IVM culture, which was an effective cytoplasmic maturation of porcine IVM oocytes.

Abstract. Glycerol plays multi-functional roles in cellular physiology. Other than forming the backbone molecule for glycerophospholipid and triglyceride (TG), glycerol acts as an energy substrate for glycolysis. Spermatozoa are known to utilize glycerol for energy production, but there are no reports of this in oocytes. In this study, the value of glycerol as an energy substrate for bovine oocyte maturation (Exp. 1) and the gene expression of glycerol kinase (GK), an enzyme crucial for cellular glycerol utilization, in bovine oocytes and early embryos (Exp. 2) were examined. In Exp. 1, in vitro maturation (IVM) was conducted using synthetic oviduct fluid supplemented with/without glucose (1.5 mM) and/or glycerol (1.0 mM), and maturation rate, degree of cumulus expansion, glucose consumption and lactate production by cumulus-oocyte complexes (COC) were examined. In Exp. 2, to examine the developmental expression of GK mRNA, cumulus cells, oocytes and embryos at the 2-, 8-and 16-cell, morula, expanded blastocyst and hatched blastocyst stages were obtained in separate experiments, and the expression of GK mRNA was quantified using a real-time PCR. Glycerol did not support oocyte maturation or cumulus expansion. Addition of glycerol to glucose-supplemented media significantly decreased the maturation rate. Expression of GK mRNA was very low in cumulus cells, whereas an appreciable level of the transcript was observed in the oocytes. GK mRNA was detected in embryos at all the stages examined, and its expression significantly increased at the morula stage. These results indicate that glycerol, at least at the present concentration, is not beneficial as a constituent of the medium for bovine oocyte maturation. However, the appreciable levels of GK mRNA found in the oocyte and embryo imply a physiological role for glycerol in bovine oocyte maturation and embryo development. Key words: Bovine, Embryo, Glycerol, Glycerol kinase (GK), Oocyte maturation (J. Reprod. Dev. 55: [177][178][179][180][181][182] 2009) o understand the nutritional requirements of oocytes and embryos, studies on metabolism of energy substrates such as glucose, pyruvate, lactate and amino acid have been performed on the bovine oocytes and early embryos [1,2]. Thanks to these studies, it has been clarified that the bovine oocyte and embryo exhibit dynamic changes in energy metabolism during the maturation and preimplantation period [1,2]. During maturation, the oocyte cannot utilize glucose [3], and energy is mainly produced by oxidation of pyruvate supplied by the surrounding cumulus cells through glycolysis [4,5]. Likewise, the early cleavage-stage embryo (up to 8-16 cells) has low glucose metabolic activity and mainly relies on oxidative phosphorylation for generation of over 90% of ATP [6]. At this stage, glucose metabolism occurs effectively via the pentose phosphate pathway to generate NADPH and ribose sugars [7]. At the 8-to 16-cell stages, there is a switch from pyruvate-based metabolism to glucose-based metabolism, which coincides with the...