Despite advances in clinical therapies and technologies, the prognosis for patients with malignant glioma is poor. Our previous research demonstrated that glioma stem cells (GSCs) were crucial for glioma malignancy and accelerated tumor migration and invasion. The migration and invasion of malignant glioma cells into the surrounding normal brain tissues cause the poor outcome. miR-145, a miRNA found to be expressed in neurons, was recently found to have reduced expression in glioblastoma multiforme tumors. And miR-145 loss in glioma cells led to increased cell proliferation and invasion. However, its function on the migration and invasion of GSCs was still unknown. In this study, we aimed to identify the effects and mechanisms of miR-145 on the migration and invasion of GSCs. Our investigations revealed that miR-145 was low expressed in malignant glioma tissues and their corresponding GSCs. Knockdown of miR-145 in vitro could enhance the migration and invasion of GSCs, while up-regulation of miR-145 had the opposite effects. Further investigation of the potential mechanism demonstrated that the function of miR-145 in regulating the migration and invasion of GSCs is mediated by its targeting of ABCG2 mRNA. ABCG2 is an ATP-binding cassette transporter protein, which was identified to be overexpressed in GSCs and higher-grade glioma tissues. We found that miR-145 was negative correlated with ABCG2 levels in GSCs, and reduction in ABCG2 expression decreased the cell migration and invasion of GSCs. Further, a luciferase reporter proved that ABCG2 was a direct target of miR-145 in GSCs. Thus, these findings underscore the potential of miR-145 to regulate the migration and invasion of GSCs through targeting ABCG2.