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Astrocyte-rich primary cultures (APCs) are frequently used as a model system for the investigation of properties of brain astrocytes. However, as APCs contain a substantial number of microglial and oligodendroglial cells, biochemical parameters determined for such cultures may at least in part reflect also the presence of the contaminating cell types. To lower the potential contributions of microglial and oligodendroglial cells on properties of the astrocytes in APCs we prepared rat astrocyte-rich secondary cultures (ASCs) by subculturing of APCs and compared these ASCs with APCs regarding basal metabolic parameters, specific enzyme activities and the accumulation of iron oxide nanoparticles. Immunocytochemical characterization revealed that ASCs contained only minute amounts of microglial and oligodendroglial cells. ASCs and APCs did not significantly differ in their specific glucose consumption and lactate production rates, in their specific iron and glutathione contents, in their specific activities of various enzymes involved in glucose and glutathione metabolism nor in their accumulation of iron oxide nanoparticles. Thus, the absence or presence of some contaminating microglial and oligodendroglial cells appears not to substantially modulate the investigated metabolic parameters of astrocyte cultures. [ABSTRACT FROM AUTHOR]

Astroglial cultures from newborn mouse cerebral cortex contain [I]insulin binding sites. Binding was specific reversible, time dependent and reached equilibrium after 45 min. Insulin analogues compete for this [I]Insulin binding. Incubation of cerebral cortex astroglial cultures with insulin induced a time-and dose-dependent inhibition of the [H]GABA high affinity uptake. A decrease in the V rather than, an effect on the K was observed. This effect was dose-dependent and effective at 10 M. Autoradiographic observations on the cell monolayer showed the presence of two groups of cells: one which strongly takes up [H]GABA and consist in smaller GFAP positive process-bearing cells and another group of much flatter and larger GFAP positive cells which uptake was lower. The smaller stellate cells were apparently the most sensitive to insulin effect. These results: 1) confirm the presence of insulin binding sites on astroglial primary cultures, 2) show an effect of insulin of [H]GABA high affinity uptake of these cells; this effect being optimal on a stellate-like population of astrocytes, and 3) indicate, that insulin may interfere in neuromodulation through astroglial signals. [ABSTRACT FROM AUTHOR]

The search for neuronotrophic factors addressing CNS neurons requires CNS neuronal cell cultures to quantitate putative effects on neuronal survival. Investigation of neurons dissociated from several embryonic CNS tissues have shown that their short-term survival requires supplementation of the culture medium with either pyruvate or the enzyme catalase. Pyruvate can be replaced with α-ketoglutarate or oxaloacetate, or with amino acids capable to transaminate to these three metabolites in the presence of exogenous α-ketoacid acceptors. Experiments were designed to evaluate the ability of cultured CNS neurons to utilized glucose as their primary source. We show that: (1) catalase requires the availability of glucose in the medium in order to exert its neuronal maintenance effect, (2) in the absence of catalase, the cells are unable to metabolize glucose through the tricarboxylic acid cycle, (3) catalase restores the neuronal ability to utilize glucose for oxydative metabolism, and renders redundant the use of other sources such as glutamate conversion to α-ketoglutarate, (4) graded concentrations of glucose in the medium affect in parallel these metabolic activities and the viability of the cultured neurons, and (5) anti-oxidant agents other than catalase mimic the catalase effects. We conclude that dissociated embryonic CNS neurons suffer from a block in glucose utilization which results from an imbalance between free radical attack and cellular defenses to it and speculate on a more general involvement of peroxidation damage in the trophic requirements for neuronal survival. [ABSTRACT FROM AUTHOR]

The presence and specificity of insulin receptors was investigated in cultured cells obtained from 15-16 days old embryonic mouse cerebra. Developmental studies suggested that the maximum insulin binding occurred at about 11 days in vitro (DIV). Scatchard analysis of binding data revealed two types of binding sites. One type of receptor was the high affinity type ( K=7.77×10 M; number of receptor sites, B=350 fmol/mg protein) while the other type was of low affinity type ( K=5.75×10 M; B=1150 fmol/mg protein). The specificity of receptors for insulin was also confirmed by showing that [I]insulin was displaced by non-radioactive insulin but not by glucagon or growth hormone. Insulin displayed a clear dose-dependent stimulation of thymidine incorporation. It also stimulated the activity of the enzyme 2′,3′-cyclic nucleotide phosphohydrolase (CNPase), which is specifically associated with myelin produced from oligodendroglia. Thus insulin has a positive influence on the proliferation and differentiation of brain cells. [ABSTRACT FROM AUTHOR]