DIFFERENTIAL EXPRESSION OF HEAT-SHOCK-70 PROTEINS IN PRIMARY CULTURESFROM RAT CEREBELLUM

While a number of studies have described the heat shock response in es tablished cell lines and in primary cultures of cells derived from the nervous system, there has been no systematic analysis comparing expre ssion and localization of the inducible heat shock 70 (hsp70) proteins and the constitutively synthesized members of the family (hsc70) in n eurons and glia. In the present communication, we utilized specific pr obes to compare the expression of hsp70 and hsc70 mRNAs and proteins i n two types of primary cultures, astroglial and neuro-astroglial, from postnatal rat cerebellum. Conditions were adjusted to maintain physio logical numbers of microglia in both types of culture, and cultures we re analyzed at a number of different time points following a precisely defined heat shock. The northern, in situ hybridization and immunohis tochemical analyses resulted in a number of novel observations concern ing the nature of the heat shock response in these neuronal and glial cells. In postnatal day 4-5 cultures, hsp70 mRNA levels were elevated for at least 10 h in both types of culture, but in situ hybridization analysis showed no evidence for hsp70 mRNAs in neurons. Microglia were the only cell type in which hsp70 was detected in non-stressed cultur es and this cell type contained the highest concentrations of hsp70 pr oteins in stressed cultures. Hsc70 mRNA levels were also increased aft er heat shock, but the increase was more transient. Hsc70 mRNAs and pr oteins were present in all cell types, again with the highest concentr ations being present in microglia. Hsc70 mRNAs and proteins were local ized in the cytoplasm at all time points examined, with hsc70 protein also being localized in nucleoli. Hsp70 mRNAs and proteins were diffus ely localized over nuclei of astrocytes, as well as of most microglia. Hsp70, but not hsc70, was localized on chromosomes in glia once they had resumed cell division after heat shock, suggesting a role for hsp7 0 either in targeting damaged chromosomal proteins or in cell division . Some cytoplasmic hsp70 was observed in astrocytes of the mixed neuro -astroglial cultures and a delayed hsp70 immunoreactivity was observed in granule neurons in these cultures, suggesting either that translat ion of low levels of hsp70 mRNAs was more efficient in neurons, or tha t glial-neuronal translocation of hsp70 proteins had taken place. Thes e results suggest that metabolism and functions of different heat shoc k protein family members may not always be identical and that care mus t be taken in extrapolation of results from one cell type to another.