Multiple system atrophy (MSA) exhibits widespread astrogliosis together with α-synuclein (α-syn) glial cytoplasmic inclusions (GCIs) in mature oligodendrocytes. We quantified astrocyte activation by morphometric analysis of MSA cases, and investigated the correlation to GCI proximity. Using Imaris software, we obtained “skinned” three-dimensional models of GFAP-positive astrocytes in MSA and control tissue (n = 75) from confocal z-stacks and measured the astrocyte process length and thickness and radial distance to the GCI. Astrocytes proximal to GCI-containing oligodendrocytes (r < 25 μm) had significantly (p, 0.05) longer and thicker processes characteristic of activation than distal astrocytes (r > 25 μm), with a reciprocal linear correlation (m, 90 μm2) between mean process length and radial distance to the nearest GCI (R2, 0.7). In primary cell culture studies, α-syn addition caused ERK-dependent activation of rat astrocytes and perinuclear α-syn inclusions in mature (MOSP-positive) rat oligodendrocytes. Activated astrocytes were also observed in close proximity to α-syn deposits in a unilateral rotenone-lesion mouse model. Moreover, unilateral injection of MSA tissue-derived α-syn into the mouse medial forebrain bundle resulted in widespread neuroinflammation in the α-syn-injected, but not sham-injected hemisphere. Taken together, our data suggests that the action of localized concentrations of α-syn may underlie both astrocyte and oligodendrocyte MSA pathological features.