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Brain (Re)Aggregate Cultures (Brain Spheroid
Cultures) P. Honegger, M.-G. Zurich, F.
Tschudi-Monnet Department of Physiology
University of Lausanne Lausanne, Switzerland
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R
R.R. Felder and U.C. Gupta, BIOforum Europe 5/2005
R. Felder U.C. Gupta
pp
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Aggregate formation from a single cell suspension
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Brain cell aggregates 17 h and 34 days in culture
Scale bar 100 mm
Scale bar 5 mm
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Brain cell aggregate histotypic structures
2 mm
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Neuron-specific enzyme activities
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Synaptogenesis in brain cell aggregates
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Maturation of macroglial cells
Days in vitro
Days in vitro
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Myelination in aggregated brain cell cultures
20 mm
2 mm
2 mm
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Effect of OTA on inflammation markers
Exposure 24 and 48 h
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• Characteristics of aggregating brain cell
  cultures and advantages for DNT
• - Spontaneous aggregation, a characteristic of
  immature cells of any species
• - Aggregation is organ-specific (e.g., no
  fibroblast contamination of brain cell cultures)
• - 3-D cell culture provides optimal cell-to-cell
  interactions
• Histotypic maturation and expression of the
  correct cellular phenotypes
• Morphogenetic events (synaptogenesis,
  myelination) according to the in vivo time
  schedule
• - Formation of neuronal networks with spontaneous
  electrical activity
• Pathogenic cascades (including glial reactivity)
  in response to primary (chemical) injuries
• - Simple methodology for culture preparation and
  maintenance
• - Suspension culture, robust, easy to handle, and
  adaptable to automation
• - Cultures develop and function in chemically
  defined medium (serum-free)
• - High yield and excellent reproducibility
• - DNT tests in both acute and chronic exposure
  schemes possible
• - Applicable to multidisciplinary assays, HT and
  HC testing
• Amenable to transcriptomics, proteomics and
  metabolomics
• Major disadvantages

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Environmental signals activating diverse
intracellular pathways and ERG networks, causing
phenotypic diversity
J. de Vellis E. Carpenter, Basic
Neurochemistry, Chapter 27
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Global changes in gene expression approach
Three independent experiments giving each
1 sample control (pool of 4 culture flasks) 1
sample OTA 10 nM, 24 h (pool of 4 culture flasks)
Checked all the samples by real time RT-PCR
Affymetrix analyses performed by P2
Criteria used for the selection of probes
p 0.0001 fold change 2
631 probes (204 EST)
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Global changes in gene expression results
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Markers of general cytotoxicity
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Effect of OTA on astrocytic cytoskeletal proteins
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Effect of OTA on astrocytic cytoskeletal proteins
control
OTA 15 nM
vimentin
GFAP
Vimentin GFAP
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Immunocytochemical characterization of aggregate
cultures
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Ammonium-induced impairment of axonal growth
Prevention by creatine
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In vitro models for brain research