As the pig becomes increasingly used for biomedical research, an effective and efficient in vitro culture system is essential. This study aimed to improve the commonly used porcine embryo culture medium, NCSU23, by altering the energy substrates and adding amino acids, using electrically activated diploid parthenotes from oocytes obtained from the ovaries of prepubertal and adult animals. Morphological development to day 6 and blastocyst cell number were examined. Glucose (5.56 mM) was replaced by pyruvate and lactate (0.2 mM and 5.7 mM, respectively) for either the entire culture period or for the first 48 h only. Blastocyst rates were not different between any of the treatments, and were similar for prepubertal and adult oocytes. When the embryos were cultured with pyruvate and lactate for the first 48 h and then glucose, there was a significant increase in blastocyst cell number compared to glucose only. Blastocysts produced using pyruvate and lactate for the entire time tended to have more cells than those exposed to glucose only and less than those who were cultured in pyruvate and lactate for the first 48 h and then glucose. Nonessential amino acids added for the first 48 h and nonessential and essential amino acids added for the remaining time significantly increased blastocyst cell number only when the embryos were grown in pyruvate and lactate followed by glucose. Blastocyst rates were not different between any of the treatments, and this result was the same when using sow or gilt oocytes. The modified medium was then tested using in vitro matured and fertilized embryos from sow oocytes. Blastocyst rates and cell number were significantly increased in the modified medium compared to those grown in unmodified NCSU23. This shows that altering energy substrates and adding amino acids can increase the quantity and cell number of IVP blastocysts compared with NCSU23.
Embryo CultureReproduction, Fertility and Development 215 GOT (91.73 ± 3.59 µ/L) and GPT (16.09 ± 3.23 µ/L) activities were recorded in high yielder and the lowest mean values of GOP (72.58 ± 4.79 µ/L) and GPT (13.05 ± 1.99 µ/L) activities were observed in low yielder cows. Based on the findings of this study, it was concluded that although the occurrence of moderate fatty liver was 41.67%, this could be associated with the infertility condition of crossbred dairy cows. It is quite possible that cows showing mild liver damage were in recovery stage after severe or moderate liver damage, as the postpartum period in cross bred cows under study varied from 1.5 to 8 months. Oxygen consumption is a ubiquitous parameter which can provide valuable information about metabolic mechanisms and embryo quality. Recently, we succeeded in non-invasively and quantitatively determining oxygen consumption of individual bovine embryos by the scanning electrochemical microscopy (SECM). The aim of this study was to assess by SECM the oxygen consumption of individual bovine embryos at different developmental stages cultured in serum-free and serum-supplemented media. Bovine oocytes were matured in IVMD101 medium [Research Institute for the Functional Peptides (IFP), Shimojo, Yamagata, Japan] and inseminated in BO-based medium. For serum-free culture, inseminated ooocytes were cultured to the blastocyst stage in IVD101 medium in an atmosphere of a low oxygen condition (5% CO 2 /5% O 2 /90% N 2 ) at 38.5 • C. For serumsupplemented culture, inseminated oocytes were cultured in HPM199 medium (IFP) supplemented with 5% calf serum (HPM199 + CS) in the presence of bovine cumulus/granulosa cells in a humidified atmosphere of 5% CO 2 in air. Oxygen consumption by individual bovine embryos was non-invasively quantified by the SECM measuring system. Some embryos were prepared for transmission electron microscopy. The oxygen consumption rates are presented in the table. Oxygen consumption rates (F) of the single embryos were low from 2-cell to 8-cell stages (0.45-0.52 × 10 −14 mol s −1 ). In serum-free culture, an increase in oxygen consumption rate was found at the morula (1.03 × 10 −14 mol s −1 ) stage, and blastocysts showed an even higher oxygen consumption rate (1.86 × 10 −14 mol s −1 ). On the other hand, the oxygen consumption of morulae and blastocysts produced in serum-supplemented medium was lower than that of embryos cultured in serum-free medium. Electron microscopic study demonstrated that many of the mitochondria of morulae and blastocycts cultured in HPM199 + CS medium were an immature form, indicating a correlation between respiration activity and development of mitochondria. These results suggest that the culture conditions affect the respiration activity of bovine embryos. The SECM procedures may have a wide application for judging embryo quality and culture conditions for embryos. Embryo Culture RESPIRATION ACTIVITY OF BOVINE EMBRYOS CULTURED IN SERUM-FREE AND SERUM-CONTAINING MEDIA THE EFFECT OF ALTERED ENERGY SUBSTRATE CON...
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