Depolarization‐dependent survival of cultured mouse cerebellar granule neurons is strain‐restrained

Fujikawa, Naoto; Tominaga-Yoshino, Keiko; Okabe, Masaru; Ogura, Akihiko

doi:10.1046/j.1460-9568.2000.00072.x

Cited by 14 publications

(6 citation statements)

References 12 publications

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“…Consistently, Fujikawa et al (2000) reported that the KCldependency of cerebellar granule neurons is strain-dependent and that especially cerebellar granule neurons from C57Bl/6J mice require KCl for survival. Similar to the cell densitydependent survival, the effect of KCl was suggested to be mediated by autocrine neurotrophic substances released into the culture medium upon stimulation (Bessho et al, 1993;D'Mello et al, 1993).…”

Section: Seeding Density and Cultivation Of Hippocampal Cortical Andsupporting

confidence: 52%

Optimized protocols for the simultaneous preparation of primary neuronal cultures of the neocortex, hippocampus and cerebellum from individual newborn (P0.5) C57Bl/6J mice

Ahlemeyer

Baumgart-Vogt

2005

Journal of Neuroscience Methods

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Section: Seeding Density and Cultivation Of Hippocampal Cortical Andsupporting

confidence: 52%

Optimized protocols for the simultaneous preparation of primary neuronal cultures of the neocortex, hippocampus and cerebellum from individual newborn (P0.5) C57Bl/6J mice

Ahlemeyer

Baumgart-Vogt

2005

Journal of Neuroscience Methods

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“…We utilized our culture system to address this issue by subjecting these cells to chronic depolarization using a medium containing 31.5 mM potassium. This approach was chosen as it represents a well‐established and documented protocol to manipulate granule cell development and neurochemical maturation (Baader and Schilling, 1996; Mellor et al, 1998; Fujikawa et al, 2000). In low‐density cultures, this had no apparent effect on NeuN immunoreactivity.…”

Section: Resultsmentioning

confidence: 99%

Developmental and cell type‐specific expression of the neuronal marker NeuN in the murine cerebellum

Weyer

Schilling

2003

J of Neuroscience Research

247

195

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NeuN is a 46/48-kD nuclear protein antigen used widely to identify postmitotic neurons in both research and diagnostics. It is expressed by neurons throughout the nervous system of a variety of species, including birds, rodents, and man (Mullen et al. [1992] Development 116:201-211). When we sought to use NeuN to follow the developmental progression of murine cerebellar interneurons, we observed that expression of this antigen in the cerebellum was restricted to granule neurons and a small population of cells present in the lower molecular layer of the adult cerebellum. In an attempt to identify these cells, we combined immunostaining for NeuN with a panel of cell type-specific markers to unambiguously identify neurons that express NeuN in the adult and developing cerebellum. In contrast to postmitotic granule neurons, NeuN was not expressed by any other immunocytochemically identified cerebellar interneurons, which comprised basket and stellate cells, Golgi neurons, unipolar brush cells, and Lugaro cells. NeuN-positive cells in the molecular layer failed to express any cell type-specific markers tested. They may represent ectopic granule cells; alternatively, they may represent a hitherto unknown population of cerebellar cells. In vitro experiments suggest that NeuN expression is related closely to granule cell axogenesis. This approach also revealed that the level of NeuN expression could be modulated by chronically depolarizing these cells. Thus, whereas NeuN expression per se is a reliable marker of proliferative capacity, levels of NeuN expression may also be indicative of the physiological status of a postmitotic neuron.

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“…Cultures of cerebellar granular cells were prepared as described by Fujikawa et al (2000), with minor modifications. Briefly, cerebella obtained from the wild-type or GluR⑀1-KO mice at 6 -7 d of age were treated with 0.2% trypsin for 15 min at 37°C.…”

Section: Production Of the Glur⑀3 And Glur⑀1/⑀3 Knock-out Micementioning

confidence: 99%

NMDA Receptor GluRϵ/NR2 Subunits Are Essential for Postsynaptic Localization and Protein Stability of GluRζ1/NR1 Subunit

Abe¹,

Fukaya²,

Yagi³

et al. 2004

J. Neurosci.

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In NMDA receptors, GluR⑀/NR2 subunits strictly require the GluR1/NR1 subunit to exit from endoplasmic reticulum (ER) to the cell surface in vitro and to the postsynapse in vivo, whereas C terminus-dependent self-surface delivery has been demonstrated for the GluR1 subunit in vitro. To test whether this leads to C terminus-dependent self-postsynaptic expression in neurons in vivo, we investigated the GluR1 subunit in cerebellar granule cells lacking two major GluR⑀ subunits, GluR⑀1/NR2A and GluR⑀3/NR2C. In the mutant cerebellum, synaptic labeling for the GluR1 subunit containing the C2 (GluR1-C2) or C2Ј (GluR1-C2Ј) cassette was reduced at mossy fiber-granule cell synapses to the extrasynaptic level. The loss was not accompanied by decreased transcription and translation levels, increased extrasynaptic labeling, or ER accumulation. Quantitative immunoblot revealed substantial reductions in the mutant cerebellum of GluR1-C2 and GluR1-C2Ј. The most severe deficit was observed in the postsynaptic density (PSD) fraction: mutant levels relative to the wild-type level were 12.3 Ϯ 3.3% for GluR1-C2 and 17.0 Ϯ 4.6% for GluR1-C2Ј. The GluR1 subunit carrying the C1 cassette (GluR1-C1) was, although low in cerebellar content, also reduced to 12.7 Ϯ 3.5% in the mutant PSD fraction. Considering a trace amount of other GluR⑀ subunits in the mutant cerebellum, the severe reductions thus represent that the GluR1 subunit, by itself, is virtually unable to accumulate at postsynaptic sites, regardless of C-terminal forms. By protein turnover analysis, the degradation of the GluR1 subunit was accelerated in the mutant cerebellum, being particularly rapid for that carrying the C2 cassette. Therefore, accompanying expression of GluR⑀ subunits is essential for postsynaptic localization and protein stability of the GluR1 subunit.

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Depolarization‐dependent survival of cultured mouse cerebellar granule neurons is strain‐restrained

Cited by 14 publications

References 12 publications

Optimized protocols for the simultaneous preparation of primary neuronal cultures of the neocortex, hippocampus and cerebellum from individual newborn (P0.5) C57Bl/6J mice

Optimized protocols for the simultaneous preparation of primary neuronal cultures of the neocortex, hippocampus and cerebellum from individual newborn (P0.5) C57Bl/6J mice

Developmental and cell type‐specific expression of the neuronal marker NeuN in the murine cerebellum

NMDA Receptor GluRϵ/NR2 Subunits Are Essential for Postsynaptic Localization and Protein Stability of GluRζ1/NR1 Subunit

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