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EVALUATION OF FERTILIZATION FOLLOWING OVULATORY DELAY WITH THIRAM IN THE LONG-EVANS HOODED RAT
Citation:
Stoker, T E., S C. Jeffay, AND S P. Darney. EVALUATION OF FERTILIZATION FOLLOWING OVULATORY DELAY WITH THIRAM IN THE LONG-EVANS HOODED RAT. Presented at Triangle Consortium for Reproductive Biology, RTP, NC, January 27, 2001.
Description:
Evaluation of fertilization following ovulatory delay with thiram in the Long-Evans Hooded Rat
1TE Stoker, 1* S Jeffay, and 1 SD Perreault.
1Gamete and Early Embryogenesis Biology Branch and 2 Endocrinology Branch, Reproductive Toxicology Division, NHEERL, US EPA, RTP, NC, USA.
Brief exposure to some pesticides, applied during a sensitive window for the neural regulation of ovulation, will block the preovulatory surge of LH, and thus delay ovulation. Thiram, a dithiocarbamate fungicide, suppresses dopamine beta hydroxylase activity thereby reducing the conversion of dopamine to norepinephrine, and hence release of GnRH. Previously, we have shown that a single i.p. injection of 50 mg/kg of thiram on proestrus (1300 h) suppresses the LH surge and delays ovulation for 24 h without altering the number of oocytes released. However, when bred, the treated dams had a decreased litter size and increased postimplantation loss. To test the hypothesis that the delayed oocytes had reduced fertilization potential, we dosed females with 50 mg/kg thiram on the day of vaginal proestrus and mated them with proven fertile males on the following evening. Zygotes were recovered the following afternoon and stained with acetalacmoid stain and Hoechst dye to access morphology and fertilization. A normal zygote was defined as having one male and one female pronuclei, the second polar body and a sperm tail inside. The percentage of normal zygotes from the delayed oocytes was significantly less than controls: 68.5 +/- 12.7 vs. 97.2 +/- 1.7, respectively. Summing all oocytes, 7.4% of the delayed oocytes were unfertilized (vs. 1% in controls) while 18.5% contained two male pronuclei with associated sperm tails (vs. 1% in controls). Interestingly, most delayed oocytes had additional sperm adhering to the zona pellucida (93.8% +/- 6.1 vs. 9.7% +/- 4.5 in controls), with an average of 2-9 extra sperm (vs. 1-2 in controls). Based on these results, we suggest that delayed oocytes have impaired ability to prevent polyspermy, but are otherwise normal. Such zygotes would yield triploid embryos that can implant but not develop to term, thus allowing for at least some of the pregnancy loss seen in the previous breeding study. An evaluation of the cortical granule release in such oocytes merits further study.