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Growth factorCell cultureGlial cell

We compared the effects of three growth factors, acidic fibroblast growth factor (aFGF), epidermal growth factor (EGF), and thrombin, on rat astroblast proliferation, morphology, glutamine synthetase-specific activity, and phenotypic expression of proteins. In vitroexperiments were made on 20-day-old primary cultures. Astroblast proliferation was stimulated transiently (after 48 h treatment) by the three growth factors, while the cell glutamine synthetase activity began to increase significantly only after 3 days of treatment. Acidic FGF and EGF, but not thrombin, modified the cell morphology. The effects on phenotypic expression were first determined after 5 days of treatment to minimize the mitogenic effect of the factors. Proteins synthesized during the last 18 h of the treatments were separated by two-dimensional polyacrylamide gel electrophoresis. About 600 spots were compared, 54 were modulated by the various treatments, 13 were altered similarly by all three factors, 28 by aFGF and EGF, 7 by only aFGF, 3 by only EGF, and 3 by only thrombin.

Embryonic and neonatal neurons require specific trophic supplements for their survival and the induction of transmitter-synthesizing enzymes in vivo and in vitro. Acidic and basic fibroblast growth factor (aFGF, bFGF) and the closely related astroglial growth factors AGF-1 and AGF-2 were studied for putative neurotrophic functions using dissociated, highly neuron-enriched cultures from chick and rat peripheral ganglia and central nervous system tissues. Embryonic chick ciliary ganglion neurons were the only peripheral neurons that responded to bFGF and AGF-2 by enhanced survival equivalent to that obtained with ciliary neurotrophic factor. Half-maximal effects were achieved with bFGF at 360 pg/ml or AGF-2 at 3 ng/ml. Small effects seen with aFGF could be potentiated by adding heparin at 1 microgram/ml. bFGF, but not ciliary neurotropic factor, also promoted neuron survival after the factor was bound to polyornithine and laminin. Both AGF-2 and ciliary neurotropic factor induced choline acetyltransferase activity during 48 hr. AGFs and FGFs also enhanced the long-term survival of embryonic chick spinal cord neurons, including motoneurons that had been retrogradely labeled with rhodamine isothiocyanate. These results demonstrate the potency of a class of mitogenic growth factors as neurotrophic agents for embryonic ciliary ganglion and spinal cord neurons--adding to the emerging evidence that mitogenic and neuronal growth factors are not strictly separate entities.

The relative synthesis of the brain-specific s100 protein increased as clonal rat glial cells, C6, progressed from logarithmic to stationary growth in monolayer culture. Drugs that disrupt microtubules, such as colchicine, vinblastine, Colcemid, and podophyllotoxin, inhibited the relative synthesis of S100 protein in stationary cultures. Colchicine (0.1 muM) caused a 50% inhibition of the relative synthesis of S100 protein whereas lumicolchicine,an isomer of colchicine that does not disrupt the microtubular system, had no effect. Succinylated concanavalin A (500 mug/ml) increased relative synthesis in logarithmic but not stationary cultures. These results suggest that signals inducing an increase in the relative synthesis of S100 protein in stationary cultures are transmitted intracellularly from the cell membrane by the microtubular network.

Modifications of the morphology, the proliferation and the synthesis of carbonic anhydrase of glial cells in primary cultures maintained in defined medium have been investigated under the action of basic fibroblast growth factor. Cultures contained essentially three cell types: astrocytes which expressed glial fibrillary acidic protein, oligodendrocytes which were characterized by the presence of carbonic anhydrase and precursor cells in which these two proteins were detected by immunocytochemistry. In the presence of basic fibroblast growth factor astrocytes and oligodendrocytes underwent morphological changes, characterized by a fibrous aspect; astroglial cells acquired essentially several long processes and oligodendroglial cells formed generally two long processes. The factor increased the proliferation of these two cell types. The quantity of carbonic anhydrase per oligodendrocyte was enhanced in treated cultures. The double‐stained precursor cells were present between days 7 and 11 of culture in defined medium, while in the presence of fibroblast growth factor these cells were more numerous and were still present after 14 days. The basic fibroblast growth factor stimulated the proliferation of these young glial cells and modified their morphology. But the differentiation of precursor cells towards one glial cell type appeared to be delayed.

In the rat liver homogenate, maximal protein kinase C activity was found at two calcium concentrations (1.75 and 3.5 mM). Subcellular fractionation of the liver homogenate revealed that the protein kinase C activity requiring 1.75 mMcalcium was present only in the cytosolic and particulate subcellular fractions. The protein kinase C activity requiring 3.5 mMcalcium concentration was mainly located in the rat liver nuclei preparation. About 19% of the liver homogenate protein kinase C activity requiring 3.5 mMcalcium was present in the nuclei. Goat anti-rat brain protein kinase C antibodies revealed a single immunoreactive band at 80–82 kDa in the rat liver nuclear, particulate, or cytosolic fractions. Based on the ratio of plasma membrane marker enzyme activity determined in the nuclear preparation, the purity of the isolated nuclei was ascertained.