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The present study sought to optimize an in vitro test system using pituitary cells isolated from previtellogenic female coho salmon and rainbow trout, focusing on fshb and lhb subunit gene expression. Initially, we optimized culture conditions for duration and benefits of culturing with and without addition of endogenous sex steroids (17\u03b2-estradiol [E2] or 11-ketotestosterone) or gonadotropin-releasing hormone (GnRH). The results suggest that culturing with and without E2 was valuable because it could mimic the (+) feedback effects on Lh that are observed from in vivo studies. After optimizing assay conditions, a suite of 12 contaminants and other hormones was evaluated for their effects on fshb and lhb gene expression. Each chemical was tested at four to five different concentrations up to solubility limitations in cell culture media. The results indicate that more chemicals alter lhb synthesis than fshb. The more potent chemicals were estrogens (E2 and 17\u03b1-ethynylestradiol) and the aromatizable androgen testosterone, which induced lhb. The estrogen antagonists 4-OH-tamoxifen and prochloraz decreased the E2-stimulated expression of lhb. Among several selective serotonin reuptake inhibitors tested, the sertraline metabolite norsertraline was notable for both increasing fshb synthesis and decreasing the E2 stimulation of lhb. These results indicate that diverse types of chemicals can alter gonadotropin production in fish. Furthermore, we have shown that pituitary cell culture is useful for screening chemicals with potential endocrine-disrupting activity and can support the development of quantitative adverse outcome pathways in fish. Environ Toxicol Chem 2023;42:1730\u20131742. \u00a9 2023 SETAC<\/jats:p>","DOI":"10.1002\/etc.5644","type":"journal-article","created":{"date-parts":[[2023,5,3]],"date-time":"2023-05-03T11:31:41Z","timestamp":1683113501000},"page":"1730-1742","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":3,"title":["Salmonid Pituitary Cells as a Test System for Identifying Endocrine-Disrupting Compounds"],"prefix":"10.1093","volume":"42","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-2645-8274","authenticated-orcid":false,"given":"Louisa","family":"Harding","sequence":"first","affiliation":[{"name":"School of Aquatic and Fishery Sciences University of Washington Seattle Washington USA"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Irvin R.","family":"Schultz","sequence":"additional","affiliation":[{"name":"Northwest Fisheries Science Center National Oceanic and Atmospheric Administration Fisheries Seattle Washington 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