Androgens can affect the singing behavior via regulating the song control system. In the present study, the effect of androgen on the synaptic plasticity of high vocal center (HVC)-robust nucleus of the arcopallium (RA) pathway was investigated through electrophysiological recording in vivo. We divided the adult male zebra finches into control, castration and castration plus testosterone implantation groups. The changes of long-term depression (LTD) and the paired-pulse facilitation in HVC-RA pathway induced by high-frequency (400 Hz, 2 s) stimulation of HVC were recorded, respectively. The results showed that high-frequency stimulation could effectively induce LTD in control group, but only evoke short-term depression in the castration group. In castration plus testosterone implantation group, LTD was restored. The paired-pulse facilitation was not obvious in the castration group, whereas it was significantly improved in the control and castration plus testosterone implantation groups. These results suggest that androgens may maintain the stability of song by influencing the level of LTD in HVC-RA pathway in adult male zebra finches, and androgens can affect the short-term synaptic plasticity of HVC-RA pathway.