Pyramidal neurons from the hippocampus undergo a well characterized programme of differentiation in vitro involving five distinct stages (1–5). While some important aspects of the dynamic organization of cell cytoplasmic structure that underlie neuronal polarization have been elucidated, little is known about corresponding changes in nuclear organization. Here we identify major changes affecting nuclear structure and gene expression during late stages of differentiation. At stage 4 a sustained increase in global transcriptional activity occurs. This is followed at stage 5 by proliferation of coiled bodies, i.e. subnuclear organelles containing splicing factors, which form a novel domain around the nucleolus that we refer to as the rosette. Both the morphology and timing of rosette formation are identical in neurons in vitro and in situ in the developing hippocampus in rat brain. Long-term synaptic inhibition in vitro or growth at low density does not prevent either nuclear reorganization, enhanced transcriptional activity or the formation of pre-synaptic specializations. These data indicate that stage-specific changes in nuclear structure and function, similar to distinct rearrangements of cytoplasmic components, are pre-programmed aspects of the neuronal differentiation pathway in the hippocampus.