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Transgenic Medaka Identify Embryonic Periods Sensitive to Disruption of Sex Determination
Petra Spirhanzlova1,2, Pauline Trébulle1, Justine Lallement1
1Laboratoire WatchFrog, Evry, Ile-de-France, France.
Abstract:
Gonadal development in medaka (Oryzias latipes) is dependent on the synergy between estrogens and androgens. Disruption of steroid hormone levels can lead to ovo-testis. To determine the sensitive windows for hormonally induced sex reversal in medaka, we developed a novel 42sp50-GFP_ChgH-GFP transgenic medaka line, allowing the identification of female gonadal tissue by fluorescence present in developing oocytes. Germinal transgenesis resulted in a stable line exhibiting a strong green fluorescent protein signal constitutively in the ovaries and in the liver in response to estrogens. The sensitivity of this line to disruption of sex determination following 16-d chronic exposures was in the nanograms per liter range. To identify the developmental period sensitive to exogenous agents, fry were exposed to 24-h pulses of high concentrations of 17β-estradiol (E2) or 5α-dihydrotestosterone (DHT) at various time points between days postfertilization (dpf) 0 and 12. Evaluation of phenotype followed by genotyping at 16 dpf revealed sensitivity to E2 between 1 and 8 dpf as well as 2 periods of susceptibility to DHT between 0 and 1 dpf and 4 and 8 dpf. No phenotypic sex reversal was detected after exposure to DHT or E2 on 11 or 12 dpf. The observed effects persisted to at least 24 dpf. The identified sensitive embryonic time periods for disruption of sex determination will aid future research on sex determination and the development of screening assays using early embryonic life stages. Environ Toxicol Chem 2020;39:842-851. © 2020 SETAC.
Insights
Medaka fish sex determination is sensitive to estrogen and androgen exposure during specific embryonic windows. This research identified key developmental periods for hormonally induced sex reversal, aiding future toxicity screening.
Area of Science:
- Endocrinology
- Developmental Biology
- Toxicology
Background:
- Medaka gonadal development relies on balanced estrogen and androgen levels; disruptions can cause intersex conditions (ovo-testis).
- Identifying sensitive developmental periods is crucial for understanding and assessing the impact of endocrine-disrupting chemicals.
Purpose of the Study:
- To determine the sensitive developmental windows for hormonally induced sex reversal in medaka (Oryzias latipes).
- To establish a transgenic medaka line for efficient identification of female gonadal tissue.
Main Methods:
- Developed a novel transgenic medaka line (42sp50-GFP_ChgH-GFP) for fluorescence-based identification of female gonadal tissue.
- Exposed medaka fry to short pulses of 17β-estradiol (E2) or 5α-dihydrotestosterone (DHT) during early development (days postfertilization 0-12).
- Evaluated phenotypic sex reversal and confirmed with genotyping at 16 days postfertilization.
Main Results:
- The transgenic line exhibited fluorescence in ovaries and liver in response to estrogens.
- Medaka were sensitive to E2 between days postfertilization 1 and 8.
- Susceptibility to DHT occurred during two periods: days 0-1 and days 4-8 postfertilization.
- No sex reversal was observed after exposure on days 11 or 12 postfertilization; effects persisted to at least day 24.
Conclusions:
- Specific embryonic time windows (1-8 dpf for E2, 0-1 dpf and 4-8 dpf for DHT) are sensitive to sex determination disruption in medaka.
- These findings support the use of early medaka embryonic stages for developing screening assays for endocrine disruptors.
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