Stromal cell-derived factor-1α (SDF-1α) plays an important role after injury. However, little is known regarding its temporal and spatial expression patterns or how it interacts with glial cells after optic nerve crush injury. We characterized the temporal and spatial expression pattern of SDF-1α in the retina and optic nerve following optic nerve crush and demonstrated that SDF-1α is localized to the glial cells that are distributed in the retina and optic nerve. CXCR4, the receptor for SDF-1α, is expressed along the ganglion cell layer (GCL). The relative expression levels of Sdf-1α mRNA and SDF-1α protein in the retina and optic nerve 1, 2, 3, 5, 7, 10 and 14 days after injury were determined using real-time polymerase chain reaction (PCR) and enzyme-linked immunosorbent assay, respectively, and the Cxcr4 mRNA expression was determined using real-time PCR. Immunofluorescence and immunohistochemical approaches were used to detect the localization of SDF-1α and CXCR4 after injury. The upregulation of Sdf-1α and Cxcr4 mRNA was detected as early as day one after injury in the retina and day two in the optic nerve, the expression peaks 5-7 days after injury. The expression of Sdf-1α and Cxcr4 mRNA was maintained for at least 14 days after the optic nerve crush injury. Furthermore, SDF-1α-positive zones were distributed locally in the reactive glial cells, which suggested potential autocrine stimulation. CXCR4 was mainly expressed in the GCL, which was also adjacent to the the glial cells. These findings suggest that following optic nerve crush, the levels of endogenous SDF-1α and CXCR4 increase in the retina and optic nerve, where activated glial cells may act as a source of increased SDF-1α protein.