Apoptosis is a form of naturally occurring cell death that plays a fundamental role during development and is characterized by internucleosomal DNA fragmentation. In this study we used specific in situ labeling of DNA breaks (Gavrieli et al. [1992] J. Cell. Biol. 119:493-501) to analyze the distribution of apoptotic cells in rat cerebral cortex and thalamus at different developmental stages from embryonic day 16 to adulthood. Control experiments and electron microscopy confirmed that the reaction product was confined to the nucleus of selected cells. Plotting and counting of labeled nuclei in counterstained paraffin sections showed that apoptosis occurred mainly during the first postnatal week and was absent in embryonic and adult samples. In the cortex, the number of apoptotic cells progressively increased from birth to the first postnatal week, with a peak between postnatal (P) day 5 and P8, and subsequently decreased. At the time of maximal expression of apoptosis, labeled nuclei were present mainly in layer VIb and underlying white matter and at the border between cortical plate and layer I. Only a few apoptotic cells were found scattered in the thalamus, without a particular concentration in selected areas, but with a peak at P5. Differences in the number of apoptotic cells between cortex and thalamus suggest that apoptotic cell death may have a different functional significance in the two brain areas.