Drugs and the developing central nervous system.
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Biomedical subjects
Publications and source records attributed to A Vernadakis.
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Age-dependent decreases in the levels of ornithine decarboxylase activity were observed in the optic lobes, cerebral hemispheres, and midbrain-diencephalon of 6--17-day-old chick embryos. In dissociated cell cultures from chick embryonic brains a similar pattern of declining ornithine decarboxylase activity with time in culture was observed. Ornithine decarboxylase activity in the dissociated brain cell cultures was stimulated by changing the culture medium. The peak stimulatory effect was shown to occur 12 h after changing the medium. Although serum-free medium stimulated ornithine decarboxylase activity slightly, the presence of serum in the medium was the primary stimulatory factor. Both fetal calf serum and heat-inactivated fetal calf serum produced dose-dependent stimulation of ornithine decarboxylase activity. Dialyzed fetal calf sera stimulated ornithine decarboxylase, but to a lower level than that produced by nondialyzed sera. Insulin (0.5--10 microgram/ml) stimulated ornithine decarboxylase activity in a dose-dependent manner in serum free medium. In addition, 10(-2) M-L-asparagine stimulated ornithine decarboxylase activity in serum-free medium.
The uptake of [3H]norepinephrine ([3H]NE) was studied in dissociated brain cell cultures prepared from 8-day-old chick embryos using the whole brain (minus optic lobes). Uptake [3H]NE, 5 x 10(-9) m, 10 min incubation, in freshly dissociated noncultured embryonic chick brain cells, was detected in 6-day-old embryos; it was temperature and drug (cocaine, metanephrine) sensitive and increased with brain development. In cultured cells, which were assayed at various days in culture, the increase in [3H]NE accumulation per culture was less than that seen in freshly dissociated noncultured embryonic cells. When [H]NE uptake was expressed per mg protein, a decrease with days in culture was observed. reflecting perhaps a dilution of growth or proliferation of cells not accumulating NE. Metanephrine, 5 X 10(-6) M, an inhibitor of extraneuronal uptake, inhibited [3H]NE in 5-day-old cultures whereas desmethylimipramine, an inhibitor of neuronal uptake, inhibited [3H]NE uptake in 15- and 20-day-old cultures. Cocaine, another neuronal inhibitor, inhibited [3N]NE at 10 and 15 days only. We interpret these findings to suggest that during early growth in culture most neuroblasts accumulate NE nonspecifically and, as neuronal maturation proceeds, NE accumulation becomes specific.
The effect of various types of serum on morphological and biochemical changes in mouse neuroblastoma cells (clone NBP2) in culture was studied. The extent of spontaneous morphological differentiation varied markedly depending upon the type of serum and was maximal in agammaglobulin calf serum (CS). The extent of morphological differentiation after treatment of cells with cyclic AMP-stimulating agents was also dependent upon serum type and was least pronounced in fetal calf serum. The doubling time and extent of clumping varied with the type of serum. The activity of tyrosine hydroxylase (TH) in NB cells was dependent upon serum type and it was highest in newborn CS and agammaglobulin CS. Although elevation of intracellular levels of cyclic AMP in NBP2 clone invariably stimulates neurite formation and TH activity, these functions were increased in certain sera without a significant increase in the cellular cyclic AMP levels. The present study shows that neurite formation, growth rate and TH activity are regulated by more than one mode, one of which is mediated by cyclic AMP. The above changes are independently regulated in the sense that the expression of one can be increased in the absence of others.
Protein sythesis was studied in C-6 glial cells and neuroblastoma (NB) cells as a function of cell density and after differentiation with dibutyryl cyclic AMP and treatment with either norepinephrine (NE), dopamine or L-dopa. In both C-6 glial cells and NB cells, unincorporated 3H-leucine decreased, whereas incorporation of 3H-leucine into protein increased with increasing cell density, particularly at high cell densities. Exposure of C-6 glial cells of NE at various dose for 60 minutes stimulated the efficiency of 3H-leucine corporation into protein. This effect was not seen with L-dopa or dopamine. In contrast to the glial cells, in neuroblastoma cells, all three neurohumors caused a decrease in the incorporation of 3H-leucine into protein. The increase in protein synthesis by NE was also seen in DBcAMP-differentiated glial cells. These findings suggest that cellular activity as reflected by protein synthesis is cell density dependent. In addition, neurohumor substances may play a regulatory role in the cellular activity of glial cells.
Studies of glial cells in neural tissue culture systems suggest that glial cells subserve different functions during development and aging of the central nervous system and that they may help modulate the neuronal environment by virtue of their responsiveness to hormones and other intrinsic factors. There is a marked proliferation of glial cells during early stages of brain development, probably reflecting the involvement of glial cells in myelination and other growth processes. Studies in culture suggest that proliferation of glial cells can be induced by steroid hormones. The migration rate of glial cells from cerebellar explants of embryonic chick brain grown in organotypic culture was measured in control and hormone-treated explants. Treatment with cortisol, corticosterone, estradiol, and progesterone significantly elevated glial cell migration from the tissue explants. The influence of steroid hormones on glial cells may be mediated via a steroid intracellular mechanism. In C-6 glioma cells and in chick embryo dissociated brain cell cultures consisting predominantly of glial cells, 3H-corticosterone was shown to accumulate by a saturable but non-specific retention mechanism. In contrast, the accumulation of 3H-corticosterone by predominantly neuronal cultures was both saturable and specific. Glial cells in culture exhibit certain age-related changes, including changes in resting membrane potentials and in cellular responses to hormone treatment, as measured by changes in incorporation of 3H-leucine into protein and incorporation of 3H-uridine into RNA. The possibility that glial cells in vivo may likewise exhibit differential responses to hormones throughout the lifespan suggests that hormones may markedly influence cellular aging.
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The development of the chick optic lobe was impaired following removal of the optic cup of the early embryo. Tectal cell number is reduced but cell size may be relatively normal. Ther was evidence of neuronal cell death and several neuron-associated proteins and enzymes (nerve-specific protein and acetylcholinesterase) showed selectively impaired maturation. However, other nerve-specific enzymes (choline acetyltransferase, tyrosine hydroxylase), develop normally on a per cell basis. The noninnervated optic lobe had a normal blood-brain barrier but a depressed ability to accumulate amino acids from plasma. Levels of 3':5'-cyclic GMP were also reduced in the nonafferented lobe.
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Urine was collected from 20 one-year-old infants from the time of awakening in the moring for 8 consecutive h. Free cortisol was measured by competitive protein-binding analysis in each urine sample. The mean cortisol excretion rate for the entire collection period was .228 +/- .019 mug/h/kg. The mean excretion rate of sleep related cortisol (1.86 +/- .28 mug/h) was significantly different from the mean rate for awake periods (2.61 +/- .28 mug/h). The difference could not be attributed to time of day. The results suggest a direct relationship between cortisol production and daytime sleep-wake states in the one-year-old.
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Integration of the central nervous system is an expression of cerebral homeostasis that is essential for the internal ability of the organism to adapt to its changing environment throughout life. It is generally accepted that neurons undergo no further division after differentiation, whereas glial cells continue to proliferate throughout life. The increase in glial cells with advanced age may reflect a compensatory process of the brain to overcome neuronal loss or neuronal functional changes that may occur with age. Therefore, these neuronal-glial interactions during development and aging may play a key role in the integrative capacity of the brain. One of the mechanisms contributing to brain stability is the blood-brain barrier, which regulates the neuronal-glial microenvironment in the mature organism. Neuronal intercommunication is mediated via neurotransmitter substances and a shift may occur from excitation to inhibition and vice versa in some CNS areas with aging. Studies of some aspects of cholinergic, monoaminergic and amino acid neurotransmission show that their maturational patterns are CNS-area specific and that some neurotransmitter processes decline with advanced age. Glial cells, besides participating in the regulation of extraneuronal environment, are also proposed to be involved in neurotransmission mechanisms in the adult and aging CNS and since they are the major CNS cellular compartment that changes with age they may thus contribute significantly to the maintenance of CNS integrative ability and adaptation with age.
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