Activin A and Inhibin A Stimulated Zebrafish Oocyte Maturation
Previous studies in the zebrafish have shown that only follicles that reach certain sizes can undergo maturation in vitro in response to exogenous hormones. To determine which sizes of follicles are responsive to activin A, follicles were isolated from ovaries and grouped according to their sizes. Different groups of follicles were then incubated with control medium or activin A (100 ng/ml) for 18 h. As shown in Figure 2A, follicles within the category of 0.34-0.51 mm did not show any response to activin A while follicles of 0.52 mm or greater had an enhanced rate of GVBD after activin A treatment. Similarly, the rate of spontaneous maturation was increased with the increasing sizes of follicles, and spontaneous maturation did not occur in follicles smaller than 0.52 mm. Subsequent experiments were conducted using follicles greater than 0.52 mm.
When follicles were incubated with different concentrations of activin A for 18 or 24 h, a dose-dependent stimulation of GVBD was observed. Activin A, at 10-100 ng/ml, significantly increased the rate of GVBD compared to that of the control group (Fig. 2B). Similarly, inhibin A also induced oocyte maturation in a dose-dependent manner (Fig. 3). Follicles treated with 10-100 ng/ml of inhibin A had a significantly higher percentage of GVBD. Similar patterns were observed when follicles were incubated with inhibin A for 18 h and 24 h, although the rates of GVBD were slightly higher in both control and treated groups after 24-h incubation (Fig. 3B) than those observed at 18 h (Fig. 3A).
FIG. 2. Developmental stage-dependent (A), and dose- and time- dependent (B) effects of activin A on final oocyte maturation. A) Follicles were grouped according to their sizes and incubated with 100 ng/ml activin A for 18 h. Data are mean ± SEM of 3 wells. B) Follicles with sizes greater than 0.52 mm were treated with different concentrations of activin A for 18 and 24 h. Different letters denote statistical significance (P < 0.05). Data are combined from three experiments.
FIG. 3. Dose- and time-dependent effects of inhibin A on final oocyte maturation. Follicles were treated with different concentrations of inhibin A for 18 h (A) and 24 h (B), after which the percentage of GVBD was scored. Data are mean ± SEM of three experiments with three replicates in each experiment. Different letters denote statistical significance (P< 0.05).