In vivo vs. in vitro models for studying the effects of elevated temperature on the GV-stage oocyte, subsequent developmental competence and gene expression.
The ovarian pool of follicle-enclosed oocytes is highly susceptible to elevated ambient temperature. It is not clear, however, whether the model of using heat shock in vitro simulates the effects of heat stress that animals experience in vivo. The current study examined the reliability of in vitro models, relative to in vivo models, for studying the effects of elevated temperature on the germinal vesicle (GV)-stage oocyte with emphasis on the expression of genes involve in maturation and early embryonic development. Cumulus oocyte complexes (COCs) were aspirated from ovaries arbitrarily collected at the slaughterhouse from multiparous Holstein cows. In the in vivo model, COCs were collected during the hot (May-September) and cold (December-April) seasons and then subjected to in vitro embryo production (IVP) at 38.5°C. In the in vitro model, COCs were collected during the cold season, pre-cultured with 75μM 3-isobutyl-1-methylxanthine (IBMX) for 16h at 38.5 or 41.2°C, and then subjected to IVP. For both models, the relative abundance of C-MOS, GDF9, GAPDH, and POU5F1 transcripts was examined in MII-stage oocytes by real-time PCR. Cleavage and blastocyst developmental rates were higher during the cold vs. hot season. IBMX pre-culture at 38.5°C successfully blocked resumption of meiosis without compromising further embryonic development, and the proportion of cleaved and developed embryos did not differ from the cold season. Exposure of GV-stage oocytes to 41.2°C reduced the proportion of cleaved oocytes developing to blastocysts relative to controls. The most prominent finding was that the relative abundance of the examined genes' transcripts was similarly reduced in heat-stressed oocytes from both models. The in vitro model was reliable and might be relevant for other environmental stressors as well.
Department of Animal Sciences, Robert H. Smith, Food and Environment, The Hebrew University, Rehovot, Israel.
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Gendelman
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Roth
D000818:Animals; D002417:Cattle / Q000235:genetics / Q000502:physiology; D002478:Cells, Cultured; D046149:Embryo Culture Techniques / Q000662:veterinary; D047108:Embryonic Development / Q000235:genetics* / Q000502:physiology; D005260:Female; D015870:Gene Expression / Q000502:physiology; D006358:Hot Temperature / Q000009:adverse effects; D059471:In Vitro Oocyte Maturation Techniques / Q000662:veterinary; D008540:Meiosis / Q000187:drug effects / Q000235:genetics / Q000502:physiology; D008954:Models, Biological; D009865:Oocytes / Q000166:cytology / Q000378:metabolism / Q000502:physiology*; D009866:Oogenesis / Q000235:genetics / Q000502:physiology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D011687:Purines / Q000494:pharmacology; D000077546:Roscovitine; D012621:Seasons
D047428:Protein Kinase Inhibitors; D011687:Purines; D000077546:Roscovitine