Development expression of glial‐specific mRNAs in primary cultures of rat brain visualized by in situ hybridization

Abstract

The localization of mRNAs which encode the gilialspecific maker proteins, glial fibrillary acidic protein (GFAP), glycerol phosphate dehydrogenase (GPDH, EC 1.1.1.8), and myelin basic protein (MBP), was mapped by in situ hybridization in primary cultures of 1–2‐day‐old rat brain in serum‐supplemented medium. Developmental changes of these expressed mRNAs were examined after various times in culture ranging from 8 to 5 days and were correlated with the histological morphological and positional characteristics of the cell. By day 8, the culture stratified into a population of flat polygonal astrocytes covered by another populatioan of phase‐dark process‐bearing cells. When counterstained with May‐Grunwald histological stain, astrocytes appeared pale blue, whereas two subpopulations of phase‐dark cells stained differentially; one was dark blue while the other was red and smaller GFAP‐specific sequences were abundant at day 8, increased in the astrocyte bedlayer as the culture became confluent, and plateaued at approximately day 16. A minor proportion of blue phase‐dark cell contained GFAP mRNA although at a lower abundance. In contrast, GPDH mRNA positive blue phase‐dark cells were seem scattered throughout the upper layer of the culture and also around the perimeter of large clumps of red phase‐dark cells. These cells were infrequent at day 8 but increased in number at later time points. The expression of MBP mRNA differed from GPDH in that it was more abundant at early time points, plateaued between day 20 and day 24, and was predominantly localized in red phase‐dark cells. Initially small and round, the MBP mRNA‐positive cells extended long processes, which then appeared to make connections with other process‐bearing cells and reorganize into clumps that dominated the culture. By day 40, MBP mRNA was scattered over membranous expanses. Thus, in situ hybridization has revealed differential gene expression and abundance of glial‐specific mRNAs in morphologically and histochemically distinct cell populations in uninduced mixed glial cultures. In future studies, the hormonal regulation of glial‐specific mRNAs will be compared to the basal genes activation presented here.