The Cas-Br-E murine leukemia virus (MuLV) induces a progressive hindlimb paralysis accompanied by a spongiform myeloencephalopathy in susceptible mice. In order to better understand the pathological process leading to these neurodegenerative lesions, we have investigated the nature of the cell type(s) infected by the virus during the course of the disease in CFW/D and SWR/J mice. For this purpose, we used in situ hybridization with virus-specific probes in combination with cell-type-specific histochemical (lectin) and immunological markers as well as morphological assessment. In the early stage of infection, endothelial cells represented the main cell type expressing viral RNA in the central nervous system (CNS). With disease progression and the appearance of lesions, microglial cells became the major cell type infected, accounting for up to 65% of the total infected cell population in diseased areas. Morphologically, these cells appeared activated and were frequently found in clusters. Infection and activation of microglial cells were almost exclusively restricted to diseased regions of the CNS. Neurons in diseased regions were not discernibly infected with virus at either early or late times of disease progression. Similarly, the proportion of infected astrocytes was typically < 1%. Although some endothelial cells and oligodendrocytes were infected by the virus, their infection was not limited to diseased CNS regions. These results are consistent with a model of indirect motor neuron degeneration, subsequent to the infection of nonneuronal CNS cells and especially of microglial cells. Infected microglial cells may play a role in the disease process by releasing not only virions or viral env-gene-encoded gp70 proteins but also other factors which may be directly or indirectly toxic to neurons. Parallels between microglial cell infection by MuLV and by lentiviruses, and specifically by human immunodeficiency virus, are discussed.