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Further, histological analysis was conducted on all animals for which the sex was ambiguous when determined by gross morphology.
Eggs were manually stripped from the female and fertilized in 0.3 × modified mammalian Ringer's solution by using the sperm obtained from the dissected testes of the three males. After 4 days, the larvae were all mixed and netted into tanks 5 at a time repeatedly, until all tanks contained 30 larvae.
Larvae were reared in 4 liters of aerated 10% Holtfreter's solution (15) and fed a solution of ground Purina rabbit chow daily.
Food levels were adjusted as the animals grew to maximize growth. 1, we exposed larvae to atrazine at nominal concentrations of 0.01, 0.1, 1.0, 10.0, and 25 parts per billion (ppb), whereas the second experiment used 0.1, 0.4, 0.8, 1.0, 25, and 200 ppb atrazine.
Histologically, the ovary is distinguished by the ovarian vesicle (hole in the center) along its entire length and the internal ring of connective tissue (in blue).
Note the melanin granules (black) in the connective tissue in Serial transverse histological sectioning was conducted on the larynges of 10 males and 10 females from each replicate from all treatments in both experiments. To estimate the size of the larynx, the was measured.
Water was changed and treatments were renewed once every 72 h.