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Biomedical subjects

A Vernadakis

Publications and source records attributed to A Vernadakis.

At least 91 records · Page 5Linked to original sources
PubMed1986

Primary glial cells and brain fibroblasts: interactions in culture.

Primary glial-enriched cultures were prepared from newborn mouse cerebral hemispheres. The cultures were grown in Dulbecco's Modified Eagle Medium in which L-valine was substituted with D-valine; this medium selectively inhibits the growth of fibroblasts. Using glutamine synthetase and glial fibrillary acidic protein as immunocytochemical markers, cultures in D-valine medium were characterized as being over 80% astrocytic. However, these cultures exhibited a suppressed growth rate and lagged behind in their differentiation as assessed biochemically using DNA content and glutamine synthetase activity as markers for growth and differentiation. Growth was restored when D-valine cultures were grown in medium containing conditioned medium derived from brain fibroblast cultures when grown on matrix or killed substrata derived from brain fibroblast cultures. This in vitro approach offers the possibility of purifying factors and developing immunological probes to investigate the possible role of brain fibroblasts in influencing glial cell function.

Animals↗
PubMed1986

Effects of opiates on the growth of neuron-enriched cultures from chick embryonic brain.

Neuron-enriched cultures derived from 6-day-old chick embryo cerebral hemispheres were treated with morphine or methadone, 10(-5) M or 10(-6) M, on days 4-6 or 6-8 in culture and were evaluated morphologically and biochemically at day 9 using phase contrast microscopy and choline acetyltransferase activity (ChAT) as a cholinergic marker. The treatment of the cultures with morphine markedly affected their growth pattern; specifically, we observed an increased number of flat cells presumptively glia, and aggregates sided by flat cells and devoid of thick bundles of neuritic processes that normally characterize neuron-enriched cultures. These morphologic changes were reflected in a drastic decrease of ChAT activity in cultures treated from day 4 to day 6 but not from 6 to 8. In contrast to morphine, exposure to 10(-6) M methadone from day 4 to day 6 resulted in reduced ChAT activity but the growth pattern of the cultures remained morphologically intact. We suggest that morphine exerts a general neurotoxic effect whereas methadone may affect some specific cholinergic function.

Animals↗
PubMed1986

An unconventional response of adenylate cyclase to morphine and naloxone in the chicken during early development.

The developmental profile of basal, NaF- and forskolin-stimulated adenylate cyclase [ATP-pyrophosphatelyase (cyclizing), EC 4.6.1.1] activity was established throughout the 21-day embryonic age of the chicken. The highest activities were observed from day 6 to day 8. Morphine inhibited NaF- and forskolin-stimulated brain adenylate cyclase activities only at days 6-8. The inhibition was not reversed by the antagonist naloxone, which also inhibited the enzyme during the same embryonic period and had no inhibitory effect thereafter. Thus, this action of morphine is not mediated through the conventional opiate receptor-adenylate cyclase system. We propose that the temporal specificity of this effect of morphine may play a role in the development of prenatal opiate effects.

Adenylyl Cyclases↗
PubMed1985

Muscarinic receptor subclasses in the chick embryo retina: influence of corticosterone treatment.

The present study was performed on retinas of chick embryos receiving at day 8 of incubation an intracerebral injection of 0.02 microgram of corticosterone. We had previously shown with the use of [3H]quinuclidinylbenzilate [( 3H]QNB) that such treatment induced the appearance of two muscarinic binding sites in the treated retinas, whereas only one was detectable in the controls. In the present study we investigated muscarinic cholinergic receptor subclasses with agonist and antagonist binding. Agonist binding was studied by varying the concentrations of carbachol and acetylcholine (10(-9) M-10(-5) M) in the presence of a constant concentration (0.2 nM) of [3H]QNB. Two subpopulations of receptors were revealed, a high- and a low-affinity receptor, in both treated and control retinas. However, in the hormone-treated retinas, the two subpopulations significantly differed from the controls in their affinity and in their relative percentage among the total receptor population. Moreover, using pirenzepine, an antagonist known to have the capacity to distinguish between muscarinic cholinergic subclasses, two receptor subpopulations were found to be present in the hormone-treated retinas but a single one in the controls. It is suggested that hormone treatment can either induce the appearance of a new subclass of muscarinic cholinergic receptors or favor the maturation of a population of retinal cells having these receptors. Pirenzepine binding in retinas from intact embryos of 7, 9, and 11 days of incubation revealed one receptor subpopulation. Thus, these findings are more consistent with the hypothesis that corticosterone effects the target cells, either inducing changes in muscarinic receptor and/or modifying the receptor environment.

Acetylcholine↗
PubMed1985

The aging brain.

Changes in neurons, glial cells, synapse morphologic and electrophysiologic appearance, and neurotransmission mechanisms during normal aging are considered for review in this article. These processes not only are fundamental in the neurobiology of normal aging but also are implicated in the pathogenesis of aging.

Aging↗
PubMed1984

Glial cells dissociated from newborn and aged mouse brain.

Changes occurring with days in culture and cell passage in cultured glial cells derived from newborn vs aged (18-mo) mouse cerebral hemispheres were compared. The activities of the enzymes glutamine synthetase (GS), an astrocyte marker, and 2',3'-cyclic nucleotide 3'- phosphohydrolase (CNP), an oligodendrocyte marker, were determined. In addition, glial fibrillary acidic protein (GFA) and glycerol phosphate dehydrogenase (GPDH) immunoreactivity was used to morphologically identify astrocytes and oligodendrocytes, respectively. In cultures derived from newborn mouse cerebral hemispheres, both GS and CNP activity and GFA-positive and GPDH-rhodamine-positive cells were present with cell passage. In general, GS activity did not change in early cell passage in cultures from either newborn or aged mouse; in passage 5, GS was high in both sources of cell populations. CNP activity increased with cell passage in cultures derived from newborn mouse; in cultures derived from aged mouse CNP was low in the primary cultures, increased with cell passages 2 and 3, and declined with passages 4 and 5. The survival of astrocytes as shown by GS and the decline in oligodendrocytes as shown by CNP was also supported by an increase in the proportions of GFA and GPDH immunoreactive cells. We interpret the increase in GS activity to parallel the astrogliosis observed in vivo in the aging brain. Moreover, the decline in oligodendrocytes in culture may represent a shift of balance between glial cell types that appears to be influenced by the age of brain tissue and time in culture.

2',3'-Cyclic Nucleotide 3'-Phosphodiesterase↗
PubMed1984

Glial cell growth in culture: influence of living cell substrata.

The role of the microenvironment in the growth of glial cells in culture has been the topic of ongoing research in this laboratory. Recently, we reported a study on the contribution of fibroblast cell substratum and extracellular matrix in glial cell growth. In the present study we report data concerning a) the influence of a neuronal-enriched living substratum from chick embryo on the growth of glial cells derived from chick embryonic brain and plated onto the substratum; b) the influence of dissociated cells derived from chick embryonic brain on the growth of established glial cells in culture, and c) the influence of dissociated cells derived from adult rat spinal cord on the growth of established glial cells from newborn rat in culture. The activities of glutamine synthetase (GS) and 2', 3'-cyclic nucleotide 3'-phosphohydrolase (CNP) were the biochemical probes determined for astrocytes and oligodendrocytes, respectively. We found that glial growth as assessed by both enzyme activities, was enhanced when a nervous tissue derived cell population was plated onto a glial-enriched substratum, whereas glial growth was inhibited when the neuronal-enriched population was the cell substratum.

Age Factors↗
PubMed1984

Effects of ethanol on cultured glial cells: proliferation and glutamine synthetase activity.

The development of glial-enriched cultures prepared from 15-day-old embryonic chick cerebral hemispheres was examined after exposure to ethanol. Between culture days 6 and 10, ethanol was added to the medium at 4 concentrations: 0.1%, 0.5%, 1.0% and 2.0% (w/v). During the drug exposure interval, morphologic maturation of the cultures was surveyed with phase microscopy; cell proliferation was assessed by cell counts and DNA content. Exposure to ethanol concentrations of 0.1% and 0.5% resulted in only minimal changes in the parameters investigated, whereas biosynthetic and enzymatic activities were altered at a threshold dose of 1.0% ethanol. A small number of cells resembling reactive astrocytes were encountered in cultures exposed to 1.0% and 2.0% ethanol. In these cultures the number of cells and DNA content were lower than in control cultures. Additionally, [14C]leucine incorporation into protein and glutamine synthetase activity were markedly diminished in cultures treated with 1.0% and 2.0% ethanol. The ethanol-related reduction in glutamine synthetase activity was not accompanied by an equivalent decrease in protein content, suggesting that glutamine synthetase or cells rich in this enzyme are selectively vulnerable to ethanol. Impairment in glutamate/glutamine metabolism may be a potential consequence of ethanol-induced changes in glutamine synthetase activity.

Animals↗
PubMed1984

Effects of corticosterone on chick embryonic retinal cells in culture.

Corticosterone has been shown to affect several patterns of glial cell and neuronal development. We have previously reported that exogenously administered corticosterone preferentially accumulated into the retinas of 8-day-old chick embryos. Moreover, we observed that it affects muscarinic cholinergic binding. Thus, we investigated the effect of different concentrations of corticosterone on retinal cells in culture. Retinas were dissected from 8-day-old embryos, dissociated and cells plated on salt-precipitated collagen. At day 5, cultures were treated with corticosterone (from 10(-9) M to 10(-7) M) for 24 h. Controls received either Dulbecco's Modified Eagle Medium (DMEM) plus 10% fetal calf serum (FCS) or DMEM only. Results show that the main effect of the hormone was inhibition of neuronal process outgrowth. Also cell aggregation, flat cell proliferation and confluency are altered in hormone-treated cultures. All these effects are reversible and can be attributed to hormone effect and not to serum deprivation.

Animals↗
PubMed1983

Muscarinic cholinergic binding in chick embryo retino-tectal system: effects of corticosterone.

Muscarinic binding sites were measured using the radioligand [3H]quinuclidinyl benzilate (QNB) in the retina and tectum of 11-day-old chick embryos, after intracerebral administration of 0.02 microgram of corticosterone at 8 days of incubation. This age was chosen because the hormone preferentially accumulates in retinas at 8 days of development. Hormone treatment significantly affected the affinity of 3H-QNB-binding sites in retinas and slightly affected the affinity in treated tecta, whereas the number of binding sites remained unchanged. The specific binding was determined with either atropine or unlabeled QNB. Scatchard plot analysis of specific 3H-QNB binding revealed the presence of nonsaturable binding at high 3H-QNB concentrations (6-11 nM) in the treated retinas, but not in controls. It can be concluded from these data that the hormone has a primary effect on retinal cells during early growth in the chick embryo. The possibility that the hormone delays maturation of specific populations of retinal cells is considered in the discussion.

Animals↗
PubMed1983

Effects of N-LAAM on [3H]etorphine binding in neuronal-enriched cell cultures.

Stereospecific [3H]etorphine binding sites are present in neuronal-enriched cell cultures dissociated from 7-day-old chick embryonic brain. Moreover, binding was regulated by both ions and GTP in a manner similar to that of in vivo brain tissue. When cultures were exposed to N-LAAM (10(-6) M) from day 6 to day 7 or 8 and assayed for binding at day 8, Bmax was decreased and KD was increased. These findings support our view that primary neuronal cultures are a suitable model with which to study interactions of drugs with opiate receptors.

Animals↗
PubMed1982

Effects of methadone on ornithine decarboxylase and cyclic nucleotide phosphohydrolase in neuronal and glial cell cultures.

Mixed neuronal and nonneuronal cell cultures were obtained from 8-day-old chick embryos cerebral hemispheres and glial-enriched cultures were obtained from fifteen-day-old chick embryo cerebral hemispheres. Cultures were exposed to methadone, a narcotic drug, from days four to six. The activity of ornithine decarboxylase (ODC) was determined at day eight and the activity of cyclic nucleotide phosphohydrolase (CNP) was determined at day fifteen. Both ODC and CNP activity were higher in mixed neuronal-nonneuronal cell cultures treated with methadone as compared to control. No effect was observed in the neuronal-enriched or glial-enriched cultures. These findings are interpreted to reflect that neuronal-glial interaction is important in the response of primary neural cells to methadone.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗
PubMed1982

Effects of corticosterone on brain cholinergic enzymes in chick embryos.

The effects of corticosterone on the cholinergic enzymes, choline acetyltransferase (ChAT) and acetylcholinesterase (AChE) were studied in the chick embryonic brain. Chick embryos received either 0.25, 0.5, or 1.0 microgram of corticosterone via the air sac daily for three days during either embryonic days 6 through 8 (E6-E8), of cerebral neurogenesis, or days 10 through 12 (E10-E12), a period of cerebellar neurogenesis. Enzyme activities were determined in cerebral hemispheres, optic lobes, cerebellum and remaining brain at 10, 15, and 20 days of incubation. In embryos treated from E6 to E8, ChAT activity was generally higher at day 10 in cerebral hemispheres and optic lobes (cerebellum was not determined) while AChE activity was not affected. At day 20 ChAT activity of treated chick embryos was lower in the cerebral hemispheres and optic lobes, but not in the cerebellum; AChE activity was higher in the cerebral hemispheres, lower in the optic lobes, and not changed in the cerebellum as compared to controls. However, in embryos treated from E10 to E12 both cerebellar ChAT and AChE activities were higher at day 15 in comparison to controls. These data show that the hormonal effects were most prominent only in the brain areas undergoing neurogenesis during the period of hormonal treatment. Since AChE activity is also present in nonneuronal cells, the observed alterations caused by corticosterone may reflect glial cell responses to the hormone. Whether the hormone affects the final number and/or maturation of cholinergic neurons and/or glial cells remain to be investigated.

Acetylcholinesterase↗
PubMed1982

[3H]Etorphine binding activity in early chick embryos: brain and body tissue.

Stereospecific [3H]etorphine binding has been detected in chick embryos as early as day 4 of incubation in both brain and body tissue. By day 10 of incubation [3H]etorphine stereospecific binding activity is not detectable in nonneuronal tissue. The ubiquitous opiate binding sites early in embryogenesis are high affinity and respond to ion and GTP regulation in a manner similar to adult brain tissue. We interpret our observations to indicate all embryonic cells prior to cell differentiation contain opiate receptors. Therefore, we propose that opiate receptors play a dual role; one function early in embryogenesis not associated with neurotransmitter regulation, and another function later in embryonic development and in the adult: the classical neurotransmitter regulatory function.

Animals↗
PubMed1981

Preferential accumulation of [3H] corticosterone in chick brain during embryonic development.

In the present study, we examined the distribution of [3H]corticosterone ([3H]B) in chick embryonic brain during development using two different routes of administration: intracerebral and intraocular. After injection of 1 microCi into the brain of 8-day embryos, [3H]B was preferentially accumulated in the retinas, whereas regions such as cerebral hemispheres, optic tecta, and midbrain showed lower amounts of [3H]B. In 14-day embryos, a slightly higher amount of [3H]B was found in retinas and midbrain in comparison with other regions of the brain. After injection into the eye, [3H]B seemed to easily diffuse to brain regions and toe preferentially accumulate in the opposite eye and very slowly diffused to other brain areas. The accumulation of the hormone in the retina parallels the presence of hormone receptors reported by others. A correlation between the preferential accumulation of hormone and its action is proposed.

Animals↗
PubMed1980

"Transdifferentiation" of C6 glial cells in culture.

The activities of cyclic nucleotide phosphohydrolase, an enzyme marker for oligodendrocytes, and glutamine synthetase, an enzyme marker for astrocytes, were studied at early (21 to 26) and late (82 to 88) cell passages. The activity of cyclic nucleotide phosphohydrolase was markedly high and that of glutamine synthetase was low in the early passages, but this relation was reversed in the late passages. These findings suggest a "transdifferentiation" of C6 glial cells with passage in culture.

2',3'-Cyclic-Nucleotide Phosphodiesterases↗