Microexplant cell cultures of fetal rat cerebellum contain essentially monolayer networks of Purkinje cells, occasional granule cells and neurons from the deep nuclei. The neurons and occasional filament-packed glial cells develop on top of a sheet of flattened, non-neuronal cells. In the absence of extrinsic input to the cerebellum and greatly reduced numbers of granule cells, the Purkinje cells develop a stunted and non-oriented dendritic arbor similar to that observed in agranular cerebella. The Purkinje cell dendritic branches, however, are spine-covered. Although the spines are not enveloped by glia and are only rarely contacted by a presynaptic bouton, most spines display a patch of electron-dense material resembling a postsynaptic membrane specialization. The Purkinje cells develop synaptic interactions among themselves and with granule cells. The ultrastructural morphology of boutons derived from both Purkinje cells and large neurons of the deep nuclei, identified after intracellular injection of horseradish peroxidase, is consistent with that observed in vivo. The present study indicates that cerebellar Purkinje cells survive and differentiate in a culture system in which individual neurons are accessible for electrophysiological and morphological analyses.