ABSTRACT
Glutamine synthetase (GS), the enzyme that catalyses glutamine synthesis from glutamate and ammonia, plays a central role in the detoxification of brain ammonia. In the central nervous system (CNS), GS also subserves additional important functions such as regulating glutamate, GABA and amino acid metabolism. Oligodendrocytes (OL) form the myelin sheath in the central nervous system (CNS) and are essential for efficient propagation of nerve impulses. In culture, OL arise from bipotential O-2A progenitor cells. These O-2A cells give rise to type-2 astrocytes in the presence of serum. GS is expressed in mature glial cells in vivo and in vitro, but it is unknown whether GS is present in glial progenitors. In addition, a comparison of the GS expression level among the various types of glial cells has never been done in vitro. The current study investigates in vitro GS expression levels in O-2A progenitors, astrocytes and OL. We demonstrate that the GS gene is expressed in O-2A progenitors and is expressed at different levels in each cultured glial cell type. GS also is stimulated during OL developmental maturation. Thus, the GS gene is expressed in O-2A cells and is regulated in a developmental and macroglial cell type-specific manner.
Subject(s)
Astrocytes/enzymology , Brain/enzymology , Gene Expression Regulation, Developmental , Glutamate-Ammonia Ligase/genetics , Oligodendroglia/enzymology , Stem Cells/enzymology , Animals , Animals, Newborn , Astrocytes/cytology , Brain/cytology , Cells, Cultured , Gene Expression Regulation, Enzymologic , Glutamate-Ammonia Ligase/analysis , Oligodendroglia/cytology , Rats , Rats, Sprague-Dawley , Stem Cells/cytologyABSTRACT
3,5,3'-Triiodo-L-thyronine (T3) acts at the genomic level by interacting with nuclear T3 receptors (T3Rs). We have used double immunostaining to follow the expression of T3Rs and oligodendrocytes (OL) lineage markers in rat secondary cultures consisting of 85-90% OL. Using antibodies against different synthetic peptides of T3Rs (alpha common: alpha 1 + alpha 2 and beta 1) we find that alpha-T3R is expressed in both O-2A progenitors and in mature OL, while beta 1-T3R is expressed only in mature OL. In cultured OL, beta 1-T3R mRNA is upregulated the most by T3. OL exhibit more numerous and longer processes when treated by T3.
Subject(s)
Brain/metabolism , Gene Expression/drug effects , Oligodendroglia/metabolism , Receptors, Thyroid Hormone/biosynthesis , Triiodothyronine/pharmacology , Animals , Animals, Newborn , Blotting, Northern , Cell Nucleus/metabolism , Cells, Cultured , Fluorescent Antibody Technique , Oligodendroglia/cytology , Oligodendroglia/drug effects , RNA, Messenger/biosynthesis , RNA, Messenger/isolation & purification , RatsABSTRACT
Previous results (Fressinaud, C., Sarliève, L.L., and Labourdette, G. J. J. Cell. Physiol., 141:667-674, 1989b) have shown that cerebroside sulfotransferase (CST; EC 2.8.2.11) is enriched in pure rat oligodendrocyte (OL) cultures and that its activity is increased by factors mitogenic for OL precursors and galactocerebroside (GC) expressing OL, such as basic fibroblast growth factor (bFGF), platelet-derived growth factor, and high insulin concentrations. In contrast, transforming growth factor beta or low insulin concentrations were found to be ineffective in this culture system. As bFGF mainly enhanced the proliferation of OL precursors (GC negative cells) rather than that of differentiated (GC+) cells, a relationship between OL precursor proliferation and CST increase was suggested. This hypothesis was first tested in 20-day-old OL cultures grown in chemically defined medium. The dose-response curve of [125I] Iododeoxyuridine ([125I]dUrd) incorporation toward bFGF was parallel to that of CST specific activity, and maximal stimulation was reached at 5 ng/ml bFGF for both. In contrast, 2',3'-cyclic nucleotide 3'-phosphodiesterase (CNP; EC 3.1.4.37) specific activity decreased after bFGF treatment. To determine if CST increase was linked to the proliferation of OL precursors induced by bFGF, cell proliferation was blocked by cytosine arabinoside (ARA-C). From 10(-8) to 10(-5) M ARA-C there was a dose-dependent inhibition of cell proliferation and a decrease in CST specific activity, whereas CNP specific activity was enhanced. When the cells were treated with bFGF and 10(-6) M ARA-C together, the proliferation was completely blocked and CST activity decreased by 72% below control values, whereas CNP activity was not significantly decreased. Immunocytochemical studies showed that the number of sulfatide-expressing cells and the number of cycling cells were increased after bFGF treatment, but that there was no overlapping between these two populations. Taken together these results suggest that CST activity and sulfatide expression appear shortly after the arrest of OL precursor division.
