Estrogens are steroid hormones that have been implicated in a variety of cellular and physiological processes in the development of diseases such as cancer and are also known to be associated with the effects of endocrine disrupting chemicals. Here we show that 17beta-estradiol (E(2)) alters microRNA (miRNA) expression profiles in the adult zebrafish (Danio rerio). An association between E(2) and the expression of 25 miRNAs was found 12 h after treatment. Among the most up-regulated miRNAs were miR-196b and let-7h, and the most down-regulated miRNAs included miR-130c and miR-101a. Tissue-specific changes in the transcripts levels of estrogen receptors (Esr1, Esr2a, and Esr2b) and miRNAs were found after hormone treatment. The most up-regulated miR-196b and its precursors are highly expressed in the skin and showed similar tissue-specific expression patterns after treatment, indicating a common pattern of regulation by E(2). MiR-196b was shown to fine-tune the expression of its target gene Hoxb8a after treatment in whole-body homogenates. Taken together, our results suggest a novel pathway for the multifunctional and pleiotropic effects of estrogens and open new directions for future investigations of their association with miRNAs involved in estrogen-regulated physiological processes and diseases.