Observations on rat cerebellar cells in vitro: influence of substratum, potassium concentration and relationship between neurones and astrocytes

Thangnipon, Wipawan; Kingsbury, Ann; Webb, M.; Balázs, R.

doi:10.1016/0165-3806(83)90215-8

Cited by 206 publications

(141 citation statements)

References 27 publications

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“…Primary cultures from 8-day-old rat cerebellum are greatly enriched in neurons. Contamination by non-neuronal cells (mainly astrocytes), as determined by immunological and morphological criteria, is below 5% (37,38). The replication of non-neuronal cells was prevented by adding 10 M cytosine arabinoside to the culture medium.…”

Section: Methodsmentioning

confidence: 97%

Changes in the Lipid Turnover, Composition, and Organization, as Sphingolipid-enriched Membrane Domains, in Rat Cerebellar Granule Cells Developing in Vitro

Prinetti

Chigorno

Prioni

et al. 2001

Journal of Biological Chemistry

169

154

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In the present paper, we report on the properties of sphingolipid-enriched domains of rat cerebellar granule cells in culture at different stages of neuronal development. The major lipid components of these domains were glycerophospholipids and cholesterol. Glycerophospholipids were 45-75% and cholesterol 15-45% of total lipids of the domains. This corresponded to 5-17% of total cell glycerophospholipids and 15-45% of total cell cholesterol. Phosphatidylcholine, mainly dipalmitoylphosphatidylcholine, was 66 -85% of all the glycerophospholipids associated with these domains. Consequently, the palmitoyl residue was significantly enriched in the domains. The surface occupied by these structures increased during development. 40 -70% of cell sphingolipids segregated in sphingolipid-enriched membrane domains, with the maximum ganglioside density in fully differentiated neurons. A high content of ceramide was found in the domains of aging neurons. Then, the sphingolipid/glycerophospholipid molar ratio was more than doubled during the initial stage of development, whereas the cholesterol/glycerophospholipid molar ratio gradually decreased during in vitro differentiation. Phosphorylated phosphoinositides, which were scant in the domains of undifferentiated cells, dramatically increased during differentiation and aging in culture. Proteins were minor components of the domains (0.1-2.8% of all domain components). Phosphotyrosine-containing proteins were selectively recovered in the sphingolipid-enriched domain. Among these, Src family protein-tyrosine kinases, known to participate to the process of neuronal differentiation, were associated with the sphingolipid-enriched domains in a way specific for the type of kinase and for the developmental stage of the cell. Proteins belonging to other signaling pathways, such as phosphoinositide 3-kinase and its downstream target, Akt, were not associated with the domains.

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Section: Methodsmentioning

confidence: 97%

Changes in the Lipid Turnover, Composition, and Organization, as Sphingolipid-enriched Membrane Domains, in Rat Cerebellar Granule Cells Developing in Vitro

Prinetti

Chigorno

Prioni

et al. 2001

Journal of Biological Chemistry

169

154

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“…Cerebellar granule neurons depend on chronic depolarization for continued survival in culture (10). This dependence is proposed to mimic a trophic requirement of cells in the cerebellar granule layer for innervation by the mossy fibers (2,51). The removal of trophic support results in upregulation of transcription factor c-Jun expression, and this is considered a requisite, early event in the cascade of signals leading to apoptosis in neurons (4,8,55,56).…”

Section: Resultsmentioning

confidence: 99%

“…The existence of physiological protein inhibitors of GSK-3 such as FRAT1 confers a further level of regulation to this kinase. FRAT1 selectively inhibits GSK-3 phosphorylation of specific targets (3,13,51). FRAT1 is a proto-oncogene first characterized as a promoter of lymphogenesis (32) and subsequently shown to play a role in development (59).…”

Section: Discussionmentioning

confidence: 99%

Lithium Blocks the c-Jun Stress Response and Protects Neurons via Its Action on Glycogen Synthase Kinase 3

Hongisto

Smeds

Brecht

et al. 2003

Molecular and Cellular Biology

131

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Lithium has been used as an effective mood-stabilizing drug for the treatment of manic episodes and depression for 50 years. More recently, lithium has been found to protect neurons from death induced by a wide array of neurotoxic insults. However, the molecular basis for the prophylactic effects of lithium have remained obscure. A target of lithium, glycogen synthase kinase 3 (GSK-3), is implicated in neuronal death after trophic deprivation. The mechanism whereby GSK-3 exerts its neurotoxic effects is also unknown. Here we show that lithium blocks the canonical c-Jun apoptotic pathway in cerebellar granule neurons deprived of trophic support. This effect is mimicked by the structurally independent inhibitors of GSK-3, FRAT1, and indirubin. Like lithium, these prevent the stress induced c-Jun protein increase and subsequent apoptosis. These events are downstream of c-Jun transactivation, since GSK-3 inhibitors block neuronal death induced by constitutively active c-Jun (Ser/Thr3Asp) and FRAT1 expression inhibits AP1 reporter activity. Consistent with this, AP1-dependent expression of proapoptotic Bim requires GSK-3-like activity. These data suggest that a GSK-3-like kinase acts in tandem with c-Jun N-terminal kinase to coordinate the full execution of the c-Jun stress response and neuronal death in response to trophic deprivation.

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“…Granule cells were obtained from the cerebellum of 8 day old Harlan Sprague-Dawley rats, plated in 100 mm dishes at a density of 9 3 10 6 cells/dish, and cultured with basal modified Eagle's medium containing 10% fetal calf serum for 8 days, the time necessary for their differentiation (19)(20)(21). Normal human skin fibroblasts were obtained by the punch technique, cultured, and propagated as described (22) in 100 mm dishes (z0.35 mg of protein/dish), using Eagle Minimum Essential Medium supplemented with 10% fetal calf serum.…”

Section: Cell Culturesmentioning

confidence: 99%

Ceramide and sphingomyelin species of fibroblasts and neurons in culture

Valsecchi

Mauri

Casellato

et al. 2007

Journal of Lipid Research

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The ceramide (Cer) and sphingomyelin (SM) species of cultured differentiated rat cerebellar granule cells and human fibroblasts were characterized by electrospray ionization-mass spectrometry. We identified 35 different species of Cer and 18 species of SM in human fibroblasts, and 35 different species of Cer and 9 species of SM were characterized in rat neurons. The main Cer species of rat cerebellar granule cells contained d18:1 sphingosine linked with palmitic, stearic, or nervonic fatty acid, and the two main SM species were d18:1,16:0 and d18:1,18:0. Both sphingolipids were enriched in detergent-resistant membranes (DRMs; or lipid rafts), and significant differences were found in the sphingolipid patterns of DRMs and of detergentsoluble fractions (DSF) from these cells. In human fibroblasts, the main Cer species were d18:1,16:0, d18:2,16:0, d18:1,24:0, d18:2,24:0, d18:1,24:1, and d18:2,24:1; the most represented species of SM were d18:1,16:0, d18:1,24:0, and d18:1,24:1. In these cells, SM was highly enriched in DRMs and Cer was mainly associated with DSF, and the species found in DRMs were markedly different from those found in DSF.-Valsecchi, M., L. Mauri, R. Casellato, S. Prioni, N. Loberto, A. Prinetti, V. Chigorno, and S. Sonnino. Ceramide and sphingomyelin species of fibroblasts and neurons in culture. J. Lipid Res. 2007. 48: 417-424.

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Observations on rat cerebellar cells in vitro: influence of substratum, potassium concentration and relationship between neurones and astrocytes

Cited by 206 publications

References 27 publications

Changes in the Lipid Turnover, Composition, and Organization, as Sphingolipid-enriched Membrane Domains, in Rat Cerebellar Granule Cells Developing in Vitro

Changes in the Lipid Turnover, Composition, and Organization, as Sphingolipid-enriched Membrane Domains, in Rat Cerebellar Granule Cells Developing in Vitro

Lithium Blocks the c-Jun Stress Response and Protects Neurons via Its Action on Glycogen Synthase Kinase 3

Ceramide and sphingomyelin species of fibroblasts and neurons in culture

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