Improved Monospermic Fertilization and Subsequent Blastocyst Formation of Bovine Oocytes Fertilized In Vitro in a Medium Containing NaCl of Decreased Concentration.

Abstract

ABSTRACT. The objective of this study was to evaluate whether changes in NaCl concentration in a fertilization medium could improve normal fertilization and preimplantation development of bovine oocytes. In vitro matured bovine oocytes were inseminated with frozenthawed semen for 18 hr in a Tyrode's medium with albumin, lactate and pyruvate (TALP), to which 114 (TALP-114), 96 (TALP-96) or 78 (TALP-78) mM NaCl was added. Presumptive zygotes were cultured for 192 hr in a modified TALP containing 90 mM NaCl, 1.5 mM glucose, 0.3% (w/v) BSA, minimal essential medium (MEM) essential and nonessential amino acids, and insulin-transferrinselenium complex. Lower polyspermy rate was obtained by the insemination in TALP-96 (7.8 ± 2.3%) than by the insemination in TALP-114 (25.6 ± 1.4%), without decrease in male pronucleus (MPN) formation. Fertilization in TALP-78 also yielded decreased polyspermic fertilization (3.8 ± 1.5%), but significant decrease in MPN formation was found (63.1 ± 3.1%). In preimplantation development, more blastocysts developed from oocytes inseminated in TALP-96 (24.1 ± 1.7%) than from oocytes inseminated in TALP-114 (16.8 ± 1.4%). TALP-78, however, did not improve preimplantation development beyond the 8-cell stage compared with TALP-114. Mean cell number of blastocyst was higher when oocytes were fertilized in TALP-96 (137.0 ± 4.5) than in TALP-114 (123.1 ± 5.1) and in ). These results demonstrate that insemination of bovine oocytes in a TALP with decreased NaCl concentration (96 mM) improves blastocyst formation and embryo viability. Decrease in NaCl concentration below 96 mM, however, may delay or inhibit MPN formation, and inhibits subsequent development in vitro. KEY WORDS: bovine, embryo development, in vitro fertilization, osmolarity, sodium chloride.

Oil-free culture system for in vitro bovine embryo production

“…Liu and Foote (1996) showed a drastic reduction in bovine embryo Oil-free culture for embryo production production when medium osmolality was 300 mOsm kg -1 . In addition, high levels of NaCl impair fertilization, increase polyspermy, decrease blastocyst formation and decrease number of blastomeres (Liu and Foote, 1996;Roh et al, 2002). Therefore, the evaporation probably affected oocyte viability, due to both increase in salt concentration and osmolality.…”

“…In addition, increased medium salt concentration affects oocyte and embryo development, independently of osmolality. Fertilization of oocytes in media with high concentration of NaCl increases polyspermy and impairs embryo development (Roh et al, 2002). Osmolality of culture media is one of the key factors affecting the success of in vitro production (IVP) of embryos (Liu and Foote, 1996).…”

Hardening of the zona pellucida of unfertilized eggs can reduce polyspermic fertilization in the pig and cow

“…Polyspermy is a common problem of in vitro fertilization (IVF) in the cow (Wang et al 1997, Roh et al 2002, Coy et al 2005, sheep (Fukui et al 1988, Slavik et al 2005, goat (De Smedt et al 1992, Mogas et al 1997, human (van der Ven et al 1985, Balakier 1993, Aoki et al 2005, and in particular, pig , Hao et al 2006. Since the reported levels of polyspermy after IVF are higher than 50% of pig oocytes and as high as 25% of cow oocytes, these two species were chosen as models for this study.…”

Macromolecular source as dependent on osmotic pressure and water source: effects on bovine in vitro embryo development and quality

“…Mammalian embryo culture media routinely use NaCl for osmolarity adjustment and ionic balance maintenance [5]. A reduced NaCl concentration in the fertilization medium and in the defined culture medium improves, respectively, monospermic fertilization [6] and embryo development [7] in bovines. In contrast, it has been reported that a high concentration of NaCl during in vitro fertilization is beneficial to the development of rat embryos [8].…”

“…Although little is known regarding the effects of NaCl concentration in media used to produce bovine embryos in vitro, the sodium ion is involved in a large number of cellular functions. High concentrations of exogenous inorganic salts may cause protein structure changes by increasing the intracellular ion levels [6,9]. NaCl concentration is not, however, the only substance responsible for the osmotic pressure regulation of culture medium, since amino acids can act as intracellular organic osmolytes [10,11].…”