To investigate the significance of impaired insulin secretion on preimplantation embryo development, outbred ICR female mice received an injection of a single dose of streptozotocin 200 mg.kg-1 14-17 days before fertilization. Oocytes were collected 24-26 h after hCG injection. Morphological evaluation revealed a lower percentage of oocytes with second polar bodies from streptozotocin-treated females in comparison with controls. Furthermore, in this group the incidence of degenerated embryos significantly increased after 120 h in vitro cultivation. Insulin (5 U per 100 g b.w.) administered twice daily to streptozotocin-treated mice significantly improved the Embryonic development. Morphological analysis of oocyte maturation in streptozotocin-treated mice showed no significant differences in comparison with control mice. It could be concluded that marked changes in preimplantation embryo development were detected in outbred ICR mice after streptozotocin administration and this process was partly reversible by insulin treatment. Furthermore, it was shown that the process of fertilization was negatively influenced and that during in vitro cultivation the delayed effects of impaired insulin secretion resulted in an increase of embryo degeneration at the time following the third mitotic cleavage.
Plant essential oils (EOs) have been reported to have health benefit properties and their preventive and therapeutic use in animals is expected to increase in the future. We evaluated the influence of five essential oils obtained from plant species which are known to have positive antimicrobial, antioxidative and anti-inflammatory effects – sage EO from Salvia officinalis L. (Lamiaceae), oregano EO from Origanum vulgare L. (Lamiaceae), thyme EO from Thymus vulgaris L. (Lamiaceae), clove EO from Syzygium aromaticum L. (Myrtaceae) and cinnamon EO from Cinnamomum zeylanicum Blume (Lauraceae) on the growth and development of mouse preimplantation embryos in vivo. Essential oils were added to commercial diet at concentrations of 0.25 % for sage EO, thyme EO, clove EO, cinnamon EO and 0.1 % for oregano EO, and fed to ICR female mice for 2 weeks ad libitum. Females were then mated with males of the same strain. Embryos obtained on Day 4 of pregnancy at the blastocyst stage were stained by morphological triple staining (Hoechst, PI, Calcein-AM) and evaluated using fluorescent microscopy. The effects of essential oils were estimated by the viability of embryos, number of nuclei and distribution of embryos according to nucleus number. Cinnamon EO significantly decreased the number of nuclei and the distribution of embryos according to nucleus number was significantly altered. Sage EO negatively influenced the distribution of embryos according to nucleus number. Clove and oregano EOs induced a significantly increased rate of cell death. Only thyme EO had no detectable effects on embryo development. In conclusion, none of the essential oils had any positive effect on embryo development, but some of them reduced the number of cells and increased the incidence of cell death., M. Domaracký, P. Rehák, Š. Juhás, J. Koppel., and Obsahuje bibliografii a bibliografické odkazy
Serotonin receptors have been found in several reproductive organs as well as in the central nervous system. Serotonin-binding sites have been demonstrated in duck ovarian follicles and the testis, hamster ovaries, human granulosa cells and mouse placenta. Local production of serotonin by the rat ovary, oviduct, uterus and testis has also been reported. We analyzed the expression of three types of serotonin receptors: 5-HT1B, 5-HT2C and 5-HT1D by reverse transcription-polymerase chain reaction in mouse unfertilized oocytes and preimplantation embryos from zygotes to the blastocyst stage in vivo. Transcripts for 5-HT1B and 5-HT2C serotonin receptors were detected neither in unfertilized oocytes nor at any stages of in vivo developing preimplantation embryos. Serotonin 5-HT1D receptor mRNA was present in unfertilized oocytes, zygotes, 2-cell embryos, compacted morulae and in vivo produced expanded blatocysts. The expression of the mRNA 5-HT1D serotonin receptor was also detected in blastocysts cultured in vitro. When added to the culture medium, specific serotonin 5-HT1D agonist sumatriptan (1 μM) significantly inhibited the development of mouse embryos cultured in vitro. Demonstration of the expression of 5-HT1D serotonin receptor in mouse oocytes and preimplantation embryos supports the idea of a functional serotonin (5-HT1D) receptor in early mammalian development., J. Veselá, P. Rehák, J. Mihalik, S. Czikková, J. Pokorný, J. Koppel., and Obsahuje bibliografii
To investigate the significance of impaired insulin secretion on preimplantation embryo development, outbred ICR female mice received a single injection of streptozotocin 130 mg (low) and 160 mg (subdiabetic) kg-1, 14-17 days before fertilization. Preimplantation embryos were collected on day 3 of pregnancy, four to eight-cell embryos were cultured in vitro 48 h (day 5) and their cell number was estimated. After spontaneous ovulation, the significantly different distribution pattern in comparison with the controls was detected only in preimplantation embryos isolated from subdiabetic (160 mg.kg-1 streptozotocin) mice. Furthermore, the incidence of degenerated embryos was significantly increased after 48 h in vitro cultivation. The analysis of cell number distribution in embryos after cultivation in vitro indicated a significant delay in cell proliferation in both experimental groups (130 and 160 mg.kg-1 streptozotocin) in comparison with control mice. After superovulation, the only significant difference was foTund in the distribution pattern of embryos isolated on day 3 of pregnancy from subdiabetic (160 mg.kg-1 streptozotocin) mice. No significant differences were found after embryo cultivation in vitro. It could be concluded th at, in outbred ICR mice, lower streptozotocin treatment (130 mg.kg-1) influenced only cell distribution of in vitro cultured embryos after spontaneous ovulation. In ICR mice, marked changes in preimplantation embryo development were detected only after subdiabetic (160 mg.kg-1) streptozotocin treatment. During in vitro cultivation delayed effects of impaired insulin secretion resulted in an increase of embryo degeneration at the time after the third mitotic cleavage. Our results indicate that the effects of impaired maternal insulin secretion on preimplantation embryo development in mice are marked and consistent after spontaneous ovulation. Suiperovulation apparently disguises subtle changes in preimplantation embryo development after low and subdiabetic streptozotocin treatment.
To estimate the significance of insulin in the regulation of preimplantation embryo growth, female mice received a single subdiabetogenic dose of streptozocin (65 mg/kg intraperitoneally) 8-11 days or 14-17 days before fertilization. Mean glycaemia levels and the number of embryos per mouse did not differ significantly between the streptozocin-treated and control groups. Morphological analysis of preimplantation embryos collected on day 3 of pregnancy revealed significant changes in the distribution pattern of preimplantation embryo stages recovered from streptozocin-treated females. Continuous insulin treatment of streptozocin-treated mice improved the impaired development of preimplantation embryos only in short-lasting experiments. After a long subdiabetic period (14-17 days) the incidence of degenerated embryos was increased in both streptozocin-treated groups. It can be concluded that the subdiabetic state in female mice impairs preimplantation embryo development which could partly be prevented by insulin treatment.
To further investigate the role of insulin during preimplantation embryo development, we compared the effects of insulin on the development of mouse and bovine preimplantation embryos and on cell proliferation during culture in vitro in simplex media. The influence of insulin on the development of mouse zygotes was determined during cultivation in mSOF medium, alone or supplemented with glucose. Similarly, the effects of insulin on the bovine preimplantation embryo development were studied in mSOF medium. The addition of insulin into mSOF medium enhanced significantly the number of cells per mouse blastocyst. Moreover, when mSOF medium was supplemented with insulin and 0.2 mmol.l-1 glucose, the percentage of hatched blastocysts and the mean cell number of mouse blastocysts were significantly higher. Insulin had no significant effect on the development of bovine embryos, produced by in vitro fertilization of in vitro matured oocytes. Neither the rates of developing embryos nor the mean number of cells in blastocysts were different in comparison with control embryos. Our results suggest that the in vitro development of mouse embryos could be enhanced by the addition of insulin to the culture medium and is further improved by the addition of glucose. In contrast to this our results indicate that insulin has no detectable beneficial effect on the preimplantation development of bovine embryos in mSOF medium., J. Mihalik, P. Rehák, J. Koppel., and Obsahuje bibliografii