Exposure to estrogenic contaminants may pose a substantial threat to freshwater ecosystems worldwide by eliciting physiological and morphological effects in fish and other organisms that may contribute to population-level declines, yet more information regarding adverse outcome pathways is needed to assess risks. Evidence suggests that reproductive effects associated with estrogen-induced feminization of male fish may result in reproductive failure an alarming prospect, considering the near-ubiquitous existence of estrogenic contaminants in the environment and increasing awareness of the widespread occurrence of feminized fish around the world. Estrogen exposure in aquatic environments is commonly linked to the widespread occurrence of testicular oocytes, an intersex condition characterized by the development of oocytes in the testis, and adverse reproductive effects associated with intersex have raised concerns about population-level effects. Furthermore, effects of global climate change may exacerbate existing effects of estrogens (or vice versa) via interactions with toxicodynamic and toxicokinetic processes, which could heighten risks for the foreseeable future. Despite concerns, large gaps exist in our understanding of adverse outcome pathways. Key events leading from estrogen exposure to potential population-level effects in wild fish are poorly understood, precluding our ability to assess risk, identify areas of concern, and monitor ecosystem health. To better understand relationships between environmental factors, estrogen exposure, and physiological effects in fish, and to inform adverse outcome pathways for feminized fish, in the present work, a series of experiments investigating biotic and abiotic factors associated with the feminization of male fish were conducted.