To investigate the regulatory mechanism of gonad inhibiting hormone (GIH) during gonadal development of Litopenaeus vannamei, expressions of gonadal development related genes(DMC1, VASA and VTG) were detected by real time fluorescent quantitative PCR after eyestalk ablation, or injection of the eyestalk extract. The results showed that DMC1 and VASA were only expressed in the testis and ovary in L. vannamei. DMC1 mRNA expression decreased significantly in testis and ovary after eyestalk ablation (P<0.05), while it increased significantly after eyestalk extract hormone injection (P<0.05). On the contrary, VTG expressions in ovaries, as well as VASA expressions in testis and ovaries increased significantly after eyestalk ablation, but decreased after eyestalk extract hormone injection. Results showed that eyestalk extract hormone affected the development of gonad by affecting the expression of the reproductive related genes.