The zebrafish is a popular model for studies of vertebrate development and toxicology. However, in vitro approaches with this organism have not been fully exploited because cell culture systems have been unavailable. We developed methods for the culture of cells from blastula-stage diploid and haploid zebrafish embryos, as well as cells from the caudal and pelvic fin, gill, liver, and viscera of adult fish. The haploid embryo-derived cells differentiated in culture to a pigmented phenotype and expressed, upon exposure to 2,3,7,8-tetrachlorodibenzo-p-dioxin, a protein that was immunologically and functionally similar to rainbow trout cytochrome P450IA1. Zebrafish cultures were grown in a complex basal nutrient medium supplemented with insulin, trout embryo extract, and low concentrations of trout and fetal bovine serum; they could not be maintained in conventional culture medium containing a high concentration of mammalian serum. Using calcium phosphate-mediated transfection, a plasmid constructed for use in mammalian cells was introduced into zebrafish embryo cell cultures and expressed in a stable manner. These results indicated that the transfection procedures utilized in mammalian systems can also be applied to zebrafish cell cultures, providing a means for in vitro alteration of the genotype and phenotype of the cells.

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PubMedDate not supplied

Preimplantation exposure to bisphenol A advances postnatal development.

Prenatal exposure to bisphenol A (BPA), an estrogenic compound, has been shown to alter postnatal development at an environmentally relevant exposure level. To elucidate these low dose effects of preimplatation exposure to BPA, two-cell mouse embryos were cultured with 1 nM BPA. More embryos exposed to 1 nM BPA than controls reached the blastocyst stage. When the blastocysts with or without BPA exposure were transferred to uterine horns of pseudopregnant recipient mice not treated with BPA, the number of pups per litter and body weight at birth did not differ. At weaning on postnatal day 21, however, pups treated with 1 nM BPA during the preimplantation period were significantly heavier than controls. These findings suggest that BPA may not only affect early embryonic development even at low, environmentally relevant doses, but also may exert late effects on postnatal development.

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PubMedDate not supplied

Fish embryo cell cultures for derivation of stem cells and transgenic chimeras.

It has been demonstrated in mammalian systems that techniques using embryonal stem cells provide advantages over conventional injection of DNA into embryos for generation of transgenic animals. We employed cell culture approaches in an attempt to develop this technology for fish transgenesis. Using a trout embryo-derived mitogenic preparation in a specialized culture medium, we initiated replication of zebrafish blastula-derived cell cultures and expressed marker genes introduced into the cells by plasmid transfection. Reintroduction of cells from the cultures into blastula-stage embryos indicated that the cultured cells survived and may contribute to the developing organism.

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