The annual cycle of Erinaceus europaeus L. as a model for a further study of cytochemical heterogeneity in Purkinje neuron nuclei

Abstract

The cytochemical heterogeneity of nuclei in Purkinje cells, which reflects differences in metabolic states within the population, has been studied by comparing cerebellar hemispheres of the hedgehog during periods of activity and hibernation. Cerebellar granule cells and hepatocytes served as controls for diploid cells. Three different cytochemical stains (Feulgen reaction, Propidium iodide and Hoechst 33342 fluorochromes) were used in order to evaluate by microdensitometry and microfluorometry how much the heterogeneity is dependent on the variations in DNA content and degree of chromatin compaction. In the active period, Feulgen-DNA values of Purkinje cells were more widely dispersed than in control cells. Some values (18-40% of the population) exceeded (hyperdiploid) the diploid value. With Propidium iodide and Hoechst 33342 (under conditions for quantitative evaluation of DNA), a few values also exceeded the maximum value for control cells. We conclude that there is nuclear heterogeneity (diploid-hyperdiploid) in the Purkinje neuron population during normal activity. The percentage of hyperdiploid nuclei changes according to the technique used; in particular, the comparison of the different stains indicates that a different chromatin compaction is mainly responsible for hyperdiploid Feulgen-DNA values, but in a few hyperdiploid cells there is an actual surplus of DNA. In view of the modalities of detection of DNA by the different cytochemical approaches utilized, this may be mainly single-stranded DNA. During hibernation nuclear heterogeneity is still present in Purkinje neurons, but with a shift of the distribution towards lower values; hyperdiploid nuclei were not detected with all the procedures used. Moreover, when Propidium iodide is used at the concentration suitable for qualitative studies, the dye intercalation is lower than during activity. Data suggest that both decreased DNA and increased chromatin compaction occur in Purkinje cells, thus pointing to a lower metabolic activity of these neurons in hibernating animals. The changes in diploid granule cells were always less than those in Purkinje neurons and could be ascribed to a further increase in chromatin compaction only, not requiring changes in DNA content.