This study has analyzed variations in the number of nucleoli and nuclear bodies, as well as in their ultrastructural and cytochemical organization, after the osmotically induced activation of supraoptic nucleus (SON) neurons of the rat. The number of nucleoli and nuclear bodies and also the nucleolar size were determined on smear preparations of previously block-impregnated SON. The mean number of nucleoli per cell was 1.35 +/- 0.6 (mean +/- SDM) in control rats. No significant variations in this value were registered either in dehydrated or rehydrated rats. The mean nucleolar volume and the total nucleolar volume per cell showed a significant increase in dehydrated rats with respect to the controls, whereas these two parameters tended to return to control values in rats rehydrated after dehydration. The mean number of nuclear bodies per cell increased significantly from 0.56 +/- 0.50 (mean +/- SDM) in control rats to 1.54 +/- 1.1 after 6 days of dehydration. By electron microscopy, SON neurons displayed a reticulated nucleolar configuration. After the osmotically induced neuronal activation, there was an increase in the proportion of the total nucleolar area occupied by the granular component, and also a reduction in the mean fibrillar-center area. The most characteristic nucleolar features in rehydrated rats were the tendency for the granular component to be segregated and the occurrence of intranucleolar vacuoles. Ultrastructural cytochemistry with a specific silver method revealed a selective silver reaction on the coiled threads of the nuclear bodies--identified as "coiled bodies"--and on the nucleolar fibrillar components in all animal groups studied. Since nucleoli play a major role in ribosome biogenesis, a relationship between these nucleolar changes and the level of cellular activity of SON neurons is proposed. Furthermore, the response of nuclear "coiled bodies" to neuronal activation suggests their participation in the processing and transport of rRNA precursors.

Nuclear pores were assessed on freeze-fracture replicas from different neuronal and glial cell types of the rat cerebellar cortex. Nuclear diameter and perimeter were measured on semithin sections, and nuclear surface area and volume were calculated from these data. The proportion of inner nuclear membrane in apposition to condensed chromatin was measured on thin sections. The values of nuclear pore numerical density (number/micron2) were as follows (mean +/- S.D.): Purkinje cells, 22 +/- 3; Golgi cells 17 +/- 3; granule cells, 6 +/- 4; stellate and basket cells, 6 +/- 1; protoplasmic astrocytes, 11 +/- 1; Bergmann glia, 10 +/- 1; oligodendrocytes, 6 +/- 1. The total number of nuclear pores per nucleus varied from 18,451 +/- 2,336 (Purkinje cells) to 621 +/- 394 (granule cells) among neurons, and from 1,782 +/- 162 (protoplasmic astrocytes) to 402 +/- 67 (oligodendrocytes) among glial cells. The number of nuclear pores per unit nuclear volume (number/micron3), a parameter related to nucleocytoplasmic transport capacity, varied from 15 +/- 2 in Purkinje cells to 6 +/- 4 in granule cells. The proportion of nuclear membrane free of condensed chromatin was significantly (P less than 0.01) correlated to pore numerical density and total number of pores per nucleus. Some nuclear pores were associated in clusters of two or more pores. The amount of pore clustering was measured by counting the proportion of pores associated in clusters. This proportion varied among the different cell types from 82% in Purkinje cells to 44% in stellate and basket cells. The amount of pore clustering showed a positive linear correlation to pore numerical density and pore number per nucleus. However, the proportion of pores in clusters was not significantly correlated with the amount of condensed chromatin applied against the inner nuclear membrane.

The increased GFAP and vimentin immunoreactivity and the morphometric and cytological changes in rat SON astrocytes may reflect a sustained upregulation of cellular activity with age, resulting in hypertrophy of glial perikarya and cell processes. Several factors that are known to influence the expression of the astrocytic phenotype, such as signals produced by degenerating neurons and activated microglia, as well as variations in neuronal activity are considered possible causes of the age-associated changes in SON astrocytes.

We present a cytological and biochemical study of the cell death of granule cell precursors in developing rat cerebellum following treatment with the cytotoxic agent methylazoxymethanol (MAM) during the first postnatal week. The density of apoptotic figures per square millimeter progressively increases after 6, 12, 24 and 44 h of treatment, whereas cells immunoreactive for proliferating cell nuclear antigen tend to disappear in the external granular layer (EGL). DNA migration on gel electrophoresis reveals a typical ladder pattern of internucleosomal cleavage following MAM treatment, whereas gel electrophoresis of rRNA shows a conspicuous degradation of both 28S and 18S rRNAs. Ultrastructural analysis has revealed the alterations of structures containing chromatin and ribonucleoprotein (RNP) in dying cells of the EGL. The typical granular beaded configuration of the condensed chromatin changes to a denser, more homogeneous texture suggesting nucleosomal disruption. The reorganization of RNP nuclear domains is reflected by the appearance of dispersed nucleoplasmic RNP particles and the formation of a coiled-body-like structure. However, typical nuclear domains involved in the splicing of RNAs, namely interchromatin granule clusters and typical "coiled bodies", are not found in apoptotic cells. Intranuclear bundles of filaments have also been detected. In the cytoplasm, the presence of dispersed single ribosomes is an initial sign of apoptosis. The massive dispersion and disruption of ribosomes detected after 24 h and 44 h of MAM treatment is reflected by the degradation of both 28S and 18s rRNAs. These results show that MAM treatment provides a useful experimental model for the study of apoptosis in the developing central nervous system. The organization of the cell nucleus in cells undergoing apoptosis clearly reflects a disruption of the nuclear compartments involved in transcription and the processing and transport of RNA and is related to the patterns of DNA and rRNA degradation.

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