Corticotropin-releasing hormone (CRH) plays a key regulatory role in coordinating the regulation of endocrine, autonomic nervous activity, immunity, osmoregulation and food intake in teleost. However, a direct effect of CRH on steroidogenesis in follicles of zebrafish remains unkown. Here, the expression of CRH receptors (CRHRs) and genes related to steroidogenesis in cultured zebrafish ovarian follicles were analyzed by real-time quantitative PCR. The contents of estradiol and progesterone in culture medium were analyzed by radioimmunoassay, respectively. The mRNA levels of CRHR1, but not CRHR2, were significantly up-regulated in both CRHα and CRHβ treatments compared with control group. Treatment with CRHβ stimulated the expression of cyp19a1a. However, other genes related to steroidogenesis had no apparent changes after being treated with CRHα and CRHβ. The protein levels of estradiol were significantly decreased after CRHα or CRHβ treatments, but no effect was found on progesterone. Taken together, these results suggested that CRH might reduce the synthesis of estradiol by regulating the expression of genes related to steroidogenesis through CRHR1.