The tiger puffer (), the most valuable teraodontiformes fish, is widely cultured in northern China. Because the testes of male tiger puffers are regarded as a delicacy, males usually have a much higher value than females. Therefore, the production of an all-male population holds considerable potential to improve the economic benefit of tiger puffers. The first step to product an all-male tiger puffer population is to induce sex reversal of males by exogenous estrogens and obtain pseudo-females. However, previous studies have indicated that the ovarian development of pseudo-females is delayed after the exposure, with small number and sizes of oocytes. To explore the regulation mechanisms underlying delayed ovarian development in pseudo-females, in this study, tiger puffers were exposed to 10 µg/L E2 for 2 h once a day from 20 to 90 days posting hatching (dph), and genetic male fish from the treatment group (10 µg/L E2) and genetic female fish from the control group (0 µg/L E2) were collected at 90, 180 and 330 dph, respectively. Then, changes in the histological and morphological features of gonads, hormones (FSH, LH, E2, and 17α, 20βOH-PROG), and genes () on the hypothalamus-pituitary-gonad axis, and genes () involved in lipid accumulation were monitored. The results showed that 10 µg/L E2 was able to induce sex reversal in genetic males and obtain pseudo-females, and those pseudo-females were not reconverted into males or intersex at 330 dph. However, the gonadosomatic index, the oocyte number, and the area of previtellogenic oocyte in pseudo-females was significantly smaller than that of the control. Moreover, compared with control females at 90 dph, lower expression levels of a, 20βOH-PROG, as well as higher expression levels of , were detected in pseudo-females. At 180 dph, only the expression levels of were significantly lower in pseudo-females than in the control. At 330 dph, there was no significant difference between pseudo-females and control females among the hormones and genes. The results indicated that concentrations that are high enough of E2 were able to induce and maintain ovarian development in pseudo-females. However, the high concentrations of E2 might affect lipid accumulation in previtellogenic oocytes by suppressingexpression, resulting in delayed oocyte growth in pseudo-females. Furthermore, the high concentrations of E2 might also suppress meiosis initiation, leading to a decreased number of oocytes in pseudo-females.