Glaucoma is a chronic optic neuropathy characterized by progressive degeneration of retinal ganglion cells (RGCs). Elevated intraocular pressure (IOP) and the resulting mechanical stress are classically considered the main causes of RGC death. However, RGC degeneration and ensuing vision loss often occur independent of IOP, indicating a multifactorial nature of glaucoma, with the likely contribution of glial and vascular function. The aim of the present study was to provide a comprehensive evaluation of the time course of neuro–glial–vascular changes associated with glaucoma progression. We used DBA/2J mice in the age range of 2–15 months as a spontaneous model of glaucoma with progressive IOP elevation and RGC loss typical of human open-angle glaucoma. We found that the onset of RGC degeneration at 10 months of age coincided with that of IOP elevation and vascular changes such as decreased density, increased lacunarity and decreased tight-junction protein zonula occludens (ZO)-1, while hypoxia-inducible factor (HIF)-1α and vascular endothelial growth factor (VEGF) were already significantly upregulated at 6 months of age together with the onset of Müller cell gliosis. Astrocytes, however, underwent significant gliosis at 10 months. These results indicate that Müller cell activation occurs well before IOP elevation, with probable inflammatory consequences, and represents an early event in the glaucomatous process. Early upregulation of HIF-1α and VEGF is likely to contribute to blood retinal barrier failure, facilitating RGC loss. The different time courses of neuro–glial–vascular changes during glaucoma progression provide further insight into the nature of the disease and suggest potential targets for the development of efficient therapeutic intervention aside from IOP lowering.