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The effect of sodium chloride concentration in chemically defined medium on development of bovine embryos in vitro
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Theriogenology
THE EFFECT OF SODIUM CHLORIDE CONCENTRATIONIN CHEMICALLY DEFINED MEDIUM ON DEVELOPMENTOF BOVINE EMBRYOS IN VITRO S. Roh~, J.I. Park, T.Y. Shin, B.C. Lee and W.S. Hwang Department of Theriogenology, College of Veterinary Medicine Seoul National University, Seoul 151-742, Republic of Korea 1AnimalLife Resources, Ansung National University, Kyonggi-do 456-749, Korea High NaC1 concentrations in culture medium are known to inhibit development of murine embryos (Lawitts and Biggers, 1991, Biol Reprod, 45:245-251). Although a completely chemically defined condition is necessary for studying specific requirements for early embryonic development, many researchers add serum or BSA to media for bovine embryo culture. The present study was carried out to develop a completely defined culture system and determine if high NaC1 concentrations in defined (PVA added) or semi-defined (BSA added) medium is toxic to bovine embryos. Oocytes from slaughterhouse ovaries were matured and fertilized using previously published protocol (Fukui et al., 1991, J Reprod Fert, 92:125-131). After 30 h of insemination, only 2-ceU embryos were selected and cultured for this experiment. The culture media used were as follows. "ILP (ll4mM of NaC1) + BSA (3mg/ml), TLP + PVA (lmg/ml), m'ILP (90mM of NaC1) + BSA, mqLP + PVA. Six to ten embryos were placed into a 30-microliter drop of each medium and the embryos were examined 10 d post-insemination without medium renewal. The experiment was replicated 4 times. All data were analyzed by chi-square and results shown in the Table 1. Table 1. Effects of NaC1 concentration in TLP medium and macromolecules (BSA or PVA) on bovine embryo devel )merit in vitro. NaC1 Macromolecule No. of Total Hatched Cone. (Cone.) 2-cell embryos Blastocysts (%) blastoeysts (%) ll4mM BSA (3mg/ml) 26 (21.6)" 14 (11.7)a 120 .__ _vv_A_0_m~J __. 3 {1._~. 1 .(0.6)~ 172 45 (17.2)a 26 (9.9)a 90raM BSA (3mg/ml) 262 PVA (lmg/ml) 188 38 (20.2)" 23 (12.2)a abDifferent superscripts within the same column differ significantly (P<0.005). It is interesting that in BSA containing medium, development of embryos was not affected, but in the completely defmed medium, embryonic development was highly affected by high NaC1 concentration. This study shows that reduced NaC1 concentration in completely defined medium is beneficial for development of bovine pre-implantation embryos in vitro. Further studies are needed to determine the interaction between BSA and sodium or chloride ions.
Theriogenology
THE EFFECT OF SODIUM CHLORIDE CONCENTRATIONIN CHEMICALLY DEFINED MEDIUM ON DEVELOPMENTOF BOVINE EMBRYOS IN VITRO S. Roh~, J.I. Park, T.Y. Shin, B.C. Lee and W.S. Hwang Department of Theriogenology, College of Veterinary Medicine Seoul National University, Seoul 151-742, Republic of Korea 1AnimalLife Resources, Ansung National University, Kyonggi-do 456-749, Korea High NaC1 concentrations in culture medium are known to inhibit development of murine embryos (Lawitts and Biggers, 1991, Biol Reprod, 45:245-251). Although a completely chemically defined condition is necessary for studying specific requirements for early embryonic development, many researchers add serum or BSA to media for bovine embryo culture. The present study was carried out to develop a completely defined culture system and determine if high NaC1 concentrations in defined (PVA added) or semi-defined (BSA added) medium is toxic to bovine embryos. Oocytes from slaughterhouse ovaries were matured and fertilized using previously published protocol (Fukui et al., 1991, J Reprod Fert, 92:125-131). After 30 h of insemination, only 2-ceU embryos were selected and cultured for this experiment. The culture media used were as follows. "ILP (ll4mM of NaC1) + BSA (3mg/ml), TLP + PVA (lmg/ml), m'ILP (90mM of NaC1) + BSA, mqLP + PVA. Six to ten embryos were placed into a 30-microliter drop of each medium and the embryos were examined 10 d post-insemination without medium renewal. The experiment was replicated 4 times. All data were analyzed by chi-square and results shown in the Table 1. Table 1. Effects of NaC1 concentration in TLP medium and macromolecules (BSA or PVA) on bovine embryo devel )merit in vitro. NaC1 Macromolecule No. of Total Hatched Cone. (Cone.) 2-cell embryos Blastocysts (%) blastoeysts (%) ll4mM BSA (3mg/ml) 26 (21.6)" 14 (11.7)a 120 .__ _vv_A_0_m~J __. 3 {1._~. 1 .(0.6)~ 172 45 (17.2)a 26 (9.9)a 90raM BSA (3mg/ml) 262 PVA (lmg/ml) 188 38 (20.2)" 23 (12.2)a abDifferent superscripts within the same column differ significantly (P<0.005). It is interesting that in BSA containing medium, development of embryos was not affected, but in the completely defmed medium, embryonic development was highly affected by high NaC1 concentration. This study shows that reduced NaC1 concentration in completely defined medium is beneficial for development of bovine pre-implantation embryos in vitro. Further studies are needed to determine the interaction between BSA and sodium or chloride ions.