MicroRNAs (miRNAs, miRs) are small noncoding RNAs that regulate gene expression at the posttranscriptional level. We have previously identified a group of novel miRNAs in proximal sciatic nerve after sciatic nerve transection by Solexa sequencing, and miR-sc3 is a member of the group. In this study, we aimed to investigate the effects of miR-sc3 on phenotypic modulation of Schwann cells (SCs). miR-sc3 was highly expressed in the injured nerve after sciatic nerve transection. An increased and decreased expression of miR-sc3 promoted and reduced the proliferation and migration of primary SCs, respectively. miR-sc3 directly targeted astrotactin 1 (Astn1) and led to translational suppression of Astn1. There was an inverse association between the time-dependent expressions of miR-sc3 and Astn1 in proximal sciatic nerve after sciatic nerve transection. Overall, miR-sc3 affected SC proliferation and migration by targeting Astn1, thus playing the regulatory role in peripheral nerve regeneration.