Morphology of dissociated hippocampal cultures from fetal mice

Peacock, John; Rush, Daphne F.; Mathers, Lawrence H.

doi:10.1016/0006-8993(79)91027-8

Cited by 52 publications

(19 citation statements)

References 14 publications

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“…Ultrastructural analysis of neurons in culture demonstrated their histotypic neuronal morphology (Figs. 4 and 5), in agreement with previous studies (22)(23)(24). The welldifferentiated neurons exhibited a histotypic pyramidal morphology, including a primary apical dendrite with multiple ramifications, finer caliber axons, and characteristic nuclear morphology.…”

supporting

confidence: 90%

Proliferation, differentiation, and long-term culture of primary hippocampal neurons.

Ray

Peterson

Schinstine

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

407

241

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Primary embryonic hippocampal neurons can develop morphologically and functionally in culture but do not survive more than a few weeks. It has been reported that basic fibroblast growth factor (bFGF) promotes the survival of and neurite elongation from fetal hippocampal neurons. We report that bFGF, in a dose-dependent manner, can induce the survival (50 pg to 1 ng/ml) and proliferation (10-20 ng/ml) of embryonic hippocampal progenitor neurons in vitro. In serumfree medium containing high concentrations of bFGF, neurons not only proliferated (4-day doubling time) and differentiated morphologically but also could be passaged and grown as continuous cell lines. The neuronal nature of the proliferating cells was positively established by immunostaining with several different neuron-specific markers and by detailed ultrastructural analyses. The proliferative effect of bFGF was used to generate nearly pure neuronal cell cultures that can be passaged, frozen, thawed, and cultured again. Neurons have been maintained >5 months in culture. The ability to establish long-term primary neuronal cultures offers the possibility that clonal lines of distinct neuronal cell types may be isolated from specific areas of the central nervous system. Such long-term neuronal cultures should prove valuable in studying neurons at the individual cell level and also in exploring interactions between neurons in vitro. The observed dose dependence raises the possibility that cell survival and proliferation in vivo may be influenced by different levels of bFGF.During embryonic development neurons and glia are generated from the neuroepithelial cells of the neural tube. Although the factors that control the growth and differentiation of these cells are largely unknown, the survival and growth of central nervous system neurons appear to be regulated by trophic factors. Nerve growth factor, the prototypical neurotrophic factor, supports the survival of cholinergic neurons of septum, striatum, and nucleus basalis (1-3). Basic fibroblast growth factor (bFGF) has a much broader range of effects and supports the survival of a variety of neurons from different regions of the brain (4-8); its most significant effect, however, is on fetal rat hippocampal neurons (4). Both neuronal survival and neurite elongation are increased in the presence of bFGF (4). Moreover, at concentrations >5 ng/ml, bFGF has short-term proliferative and morphodifferentiation effects on neuronal precursor cells from mesencephalon, telencephalon, cerebral hemispheres, and spinal cord (9-11). Proliferation and differentiation of primary neurons from both fetal and adult striatum in response to a combination of nerve growth factor and bFGF, only epidermal growth factor, or bFGF have been reported (12-14). Furthermore, long-term mixed cultures of neurons and glia have been established from fetal and neonatal rat and mouse brains (15, 16). However, in many cases these cell lines were transformed in vitro and caused tumors when injected into syngeneic animals (15, 16).Unavail...

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supporting

confidence: 90%

Proliferation, differentiation, and long-term culture of primary hippocampal neurons.

Ray

Peterson

Schinstine

et al. 1993

Proc. Natl. Acad. Sci. U.S.A.

407

241

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“…We must, however, stress the absence of the typical mossy fiber terminals observed in the hippocampus in situ (Amaral and Dent, 1981). The lack of this type of axonal endings connecting in situ granule cells with pyramidal neurons of the CA3-CA4 fields was, however, also noted previously in dissociated hippocampal cultures (Peacock et al, 1979;Bartlett and Banker, 1984). This may result from later differentiation of dentate gyrus neurons or from the absence in dissociated cultures of lamination typical for hippocampal formation in vivo.…”

Section: Discussionsupporting

confidence: 50%

Neurotoxic effect induced by quinolinic acid in dissociated cell culture of mouse hippocampus

Хаспеков

Kida

Victorov

et al. 1989

J of Neuroscience Research

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Quinolinic acid (QUIN), an endogenous convulsant that is a product of tryptophan metabolism, is suggested to belong to the pathogenic factors in some neurodegenerative disorders of the central nervous system (CNS). The aim of the present study was to evaluate the effect of QUIN on hippocampal nerve cells in dissociated cell culture at different periods of its development in vitro. The neurodegenerative effect of QUIN in vitro was found 1.5 to 2 hr after exposure to QUIN (500 microM) in differentiated hippocampal culture 20-21 days in vitro (DIV). The cytotoxic action depended on the nature of the interneuronal relations established in culture and on the period of neuronal development in vitro. Neurons in 11-DIV cultures were undamaged even 10 days after exposure to QUIN; nor were the individual, isolated neurons probably not connected synaptically with the other ones in 20-21 DIV cultures. Ultrastructural analysis of 20-21 DIV cultures exposed to QUIN showed acute swelling and destruction of postsynaptic elements and severe degeneration of individual nerve cells located in cellular aggregates, whereas presynaptic axon terminals remained intact. The results emphasize the utility of QUIN as a tool to model degenerative changes of CNS and confirm the participation of mature synaptic junctions in QUIN neurotoxicity.

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“…Such matings result, on average, in one of three embryos with three copies of chromosome 16 (Gropp et al, 1975). Primary hippocampal cultures were prepared at gestation day 16 from Ts16 embryos and their diploid littermates (Banker and Cowan, 1977;Peacock et al, 1979). For this procedure, embryos were removed into ice-cold GBSS solution [Gey's balanced salt solution, containing (in mM): NaC l 136, KC l 5, MgSO 4 0.3, NaH 2 PO 4 1, CaCl 2 1.5, NaHC O 3 2.7, K H 2 PO 4 0.22, MgC l 2 1, glucose 5, pH 7.4] after the maternal animal was decapitated under deep ether anesthesia.…”

Section: Preparation Of Cellsmentioning

confidence: 99%

Altered Ca²⁺Signaling and Mitochondrial Deficiencies in Hippocampal Neurons of Trisomy 16 Mice: A Model of Down’s Syndrome

1998

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It has been suggested that augmented nerve cell death in neurodegenerative diseases might result from an impairment of mitochondrial function. To test this hypothesis, we investigated age-dependent changes in neuronal survival and glutamate effects on Ca 2ϩ homeostasis and mitochondrial energy metabolism in cultured hippocampal neurons from diploid and trisomy 16 (Ts16) mice, a model of Down's syndrome. Microfluorometric techniques were used to measure survival rate, [Ca 2ϩ ] i level, mitochondrial membrane potential, and NAD(P)H autofluorescence. We found that Ts16 neurons die more than twice as fast as diploid neurons under otherwise identical culture conditions. Basal [Ca 2ϩ ] i levels were elevated in Ts16 neurons. Moreover, in comparison to diploid neurons, Ts16 neurons showed a prolonged recovery of [Ca 2ϩ ] i and mitochondrial membrane potential after brief glutamate application. Glutamate evoked an initial NAD(P)H decrease that was found to be extended in Ts16 neurons in comparison to diploid neurons. Furthermore, for all age groups tested, glutamate failed to cause a subsequent NAD(P)H overshoot in Ts16 cultures in contrast to diploid cultures. In the presence of cyclosporin A, an inhibitor of the mitochondrial membrane permeability transition, NAD(P)H increase was observed in both diploid and Ts16 neurons. The results support the hypothesis that Ca 2ϩ impairs mitochondrial energy metabolism and may play a role in the pathogenesis of neurodegenerative changes in neurons from Ts16 mice.

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Morphology of dissociated hippocampal cultures from fetal mice

Cited by 52 publications

References 14 publications

Proliferation, differentiation, and long-term culture of primary hippocampal neurons.

Proliferation, differentiation, and long-term culture of primary hippocampal neurons.

Neurotoxic effect induced by quinolinic acid in dissociated cell culture of mouse hippocampus

Altered Ca²⁺Signaling and Mitochondrial Deficiencies in Hippocampal Neurons of Trisomy 16 Mice: A Model of Down’s Syndrome

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Morphology of dissociated hippocampal cultures from fetal mice

Cited by 52 publications

References 14 publications

Proliferation, differentiation, and long-term culture of primary hippocampal neurons.

Proliferation, differentiation, and long-term culture of primary hippocampal neurons.

Neurotoxic effect induced by quinolinic acid in dissociated cell culture of mouse hippocampus

Altered Ca2+Signaling and Mitochondrial Deficiencies in Hippocampal Neurons of Trisomy 16 Mice: A Model of Down’s Syndrome

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Product

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Altered Ca²⁺Signaling and Mitochondrial Deficiencies in Hippocampal Neurons of Trisomy 16 Mice: A Model of Down’s Syndrome