Metabolic mapping of astrocytes and neurons in culture using stable isotopes and gas chromatography-mass spectrometry (GC-MS)
Research output: Contribution to journal › Journal article › Research › peer-review
Standard
Metabolic mapping of astrocytes and neurons in culture using stable isotopes and gas chromatography-mass spectrometry (GC-MS). / Walls, Anne B.; Bak, Lasse K.; Sonnewald, Ursula; Schousboe, Arne; Waagepetersen, Helle S.
In: Neuromethods, Vol. 90, 01.01.2014, p. 73-105.Research output: Contribution to journal › Journal article › Research › peer-review
Harvard
APA
Vancouver
Author
Bibtex
}
RIS
TY - JOUR
T1 - Metabolic mapping of astrocytes and neurons in culture using stable isotopes and gas chromatography-mass spectrometry (GC-MS)
AU - Walls, Anne B.
AU - Bak, Lasse K.
AU - Sonnewald, Ursula
AU - Schousboe, Arne
AU - Waagepetersen, Helle S.
PY - 2014/1/1
Y1 - 2014/1/1
N2 - The experimental procedure involved in metabolic mapping of astrocytes and neurons in culture using stable isotopes and gas chromatography-mass spectrometry (GC-MS) comprises several steps each of which has to be thoroughly considered in order to obtain the desired information. In this chapter we describe the different aspects to be considered when designing an incubation experiment to provide information about cellular metabolism in vitro, i.e., incubation time, incubation medium, and which isotope to employ. The labeling patterns obtained in several metabolites following metabolism of a variety of 13C or 15N labeled precursors through different metabolic pathways are depicted in order to provide suffi cient insight to enable the reader to select the best suited precursor. The cell extraction and sample preparation procedures required before GC-MS analysis are described as are the subsequent integration of chromatograms and the calculations needed to correct for natural abundance and to obtain percentage labeling of M, M + 1, M + 2, etc. in a compound. Also, the cell culturing procedure for preparing primary cultures of neurons and astrocytes and also co-cultures of these cell types isolated from either mouse cerebral cortex or cerebellum is described in detail.
AB - The experimental procedure involved in metabolic mapping of astrocytes and neurons in culture using stable isotopes and gas chromatography-mass spectrometry (GC-MS) comprises several steps each of which has to be thoroughly considered in order to obtain the desired information. In this chapter we describe the different aspects to be considered when designing an incubation experiment to provide information about cellular metabolism in vitro, i.e., incubation time, incubation medium, and which isotope to employ. The labeling patterns obtained in several metabolites following metabolism of a variety of 13C or 15N labeled precursors through different metabolic pathways are depicted in order to provide suffi cient insight to enable the reader to select the best suited precursor. The cell extraction and sample preparation procedures required before GC-MS analysis are described as are the subsequent integration of chromatograms and the calculations needed to correct for natural abundance and to obtain percentage labeling of M, M + 1, M + 2, etc. in a compound. Also, the cell culturing procedure for preparing primary cultures of neurons and astrocytes and also co-cultures of these cell types isolated from either mouse cerebral cortex or cerebellum is described in detail.
KW - C glucose/glutamate/glutamine
KW - N glutamate/glutamine
KW - Astrocytes
KW - Calculations of isotopic enrichment
KW - Co-cultures
KW - Glycogen
KW - Mass spectrometry
KW - Neurons
KW - Pentose phosphate pathway
UR - http://www.scopus.com/inward/record.url?scp=84921832164&partnerID=8YFLogxK
U2 - 10.1007/978-1-4939-1059-5_4
DO - 10.1007/978-1-4939-1059-5_4
M3 - Journal article
AN - SCOPUS:84921832164
VL - 90
SP - 73
EP - 105
JO - Neuromethods
JF - Neuromethods
SN - 0893-2336
ER -
ID: 203245511