Activation of astrocytes and abnormal synaptic glutamate metabolism are closely associated with the induction and maintenance of neuropathic pain (NP), but the exact mechanism underlying this association remains unclear. N-myc downstream-regulated gene 2 (NDRG2), a novel tumor-suppressor protein and stress-response gene, is involved in the pathogenesis of several neurodegenerative diseases. However, its role in nociceptive transduction has rarely been investigated. Here, we found that NDRG2, which was mainly expressed in the astrocytes in the central nervous system (CNS), was increased in the spinal cord of a spinal nerve ligation (SNL) rat model for NP. Suppression of NDRG2 by intrathecal injection of an NDRG2-RNAi-adenovirus significantly alleviated SNL-induced mechanical and thermal hypersensitivity, as well as elevated astrocytic glutamate transporter 1 (GLT-1) expression and downregulated pro-inflammatory cytokine levels, in the spinal dorsal horn of rats on Day 10 after SNL. Furthermore, in lipopolysaccharide (LPS)-stimulated primary astrocytic cultures derived from neonatal rats, inhibition of NDRG2 significantly reversed both the LPS-induced activation of astrocytes and decreased expression of GLT-1. By contrast, overexpression of NDRG2 by an adenoviral vector carrying NDRG2 resulted in astrocytic activation, aberrant glutamatergic neurotransmission, and spontaneous nociceptive responses in rats. Intrathecal injection of AG490, which is an inhibitor of the Janus tyrosine kinase and signal transducer and activator of the transcription 3 (JAK/STAT3) signaling pathway, significantly attenuated both mechanical and thermal hyperalgesia, as well as inhibited reactive astrocytes and restored normal expression levels of astrocytic GLT-1, in the spinal dorsal horn of NDRG2-overexpression rats. In conclusion, spinal astrocytic NDRG2 is critical in the maintenance of NP. Moreover, NDRG2 modulates astrocytic activation and inflammatory responses via regulating GLT-1 expression through the JAK/STAT3 signaling pathway. Our findings suggested that NDRG2 could be a novel therapeutic target for the treatment of NP.