Abstract
The introduction of microplate-based assays that measure extracellular fluxes in intact, living cells has revolutionized the field of cellular bioenergetics. Here, we describe a method for real time assessment of mitochondrial oxygen consumption rates in primary mouse cortical neurons and astrocytes. This method requires the Extracellular Flux Analyzer Instrument (XF24, Seahorse Biosciences), which uses fluorescent oxygen sensors in a microplate assay format.
This is a preview of subscription content, log in via an institution to check access.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
References
Stanley IA, Ribeiro SM, Giménez-Cassina A et al (2013) Changing appetites: the adaptive advantages of fuel choice. Trends Cell Biol 24:118–127
Pivovarova NB, Andrews SB (2010) Calcium-dependent mitochondrial function and dysfunction in neurons. FEBS J 277(18):3622–3636
Mattson MP, Gleichmann M, Cheng A (2008) Mitochondria in neuroplasticity and neurological disorders. Neuron 60(5):748–766
Ronald ZH, Zielke CL, Baab PJ (2009) Direct measurement of oxidative metabolism in the living brain by microdialysis: a review. J Neurochem 109:24–29
Gimenez-Cassina A, Martinez-Francois JR, Fisher JK et al (2012) BAD-dependent regulation of fuel metabolism and K(ATP) channel activity confers resistance to epileptic seizures. Neuron 74(4):719–730
Brand MD, Nicholls DG (2011) Assessing mitochondrial dysfunction in cells. Biochem J 435(2):297–312
Lange M, Zeng Y, Knight A et al (2012) Comprehensive method for culturing embryonic dorsal root ganglion neurons for seahorse extracellular flux XF24 analysis. Front Neurol 3:175
Sauerbeck A, Pandya J, Singh I et al (2011) Analysis of regional brain mitochondrial bioenergetics and susceptibility to mitochondrial inhibition utilizing a microplate based system. J Neurosci Methods 198(1):36–43
Clerc P, Polster BM (2012) Investigation of mitochondrial dysfunction by sequential microplate-based respiration measurements from intact and permeabilized neurons. PLoS One 7(4):e34465
Wu M, Neilson A, Swift AL et al (2007) Multiparameter metabolic analysis reveals a close link between attenuated mitochondrial bioenergetic function and enhanced glycolysis dependency in human tumor cells. Am J Physiol Cell Physiol 292(1):C125–C136
Gerencser AA, Neilson A, Choi SW et al (2009) Quantitative microplate-based respirometry with correction for oxygen diffusion. Anal Chem 81(16):6868–6878
Ferrick DA, Neilson A, Beeson C (2008) Advances in measuring cellular bioenergetics using extracellular flux. Drug Discov Today 13(5–6):268–274
Acknowledgments
The authors acknowledge support from the US National Institute of Health grants R56 NS072142 and the Portuguese Foundation for Science and Technology (FCT) grant SFRH/BD/51200/2010. We thank the members of the Danial laboratory for helpful discussions.
Author information
Authors and Affiliations
Corresponding author
Editor information
Editors and Affiliations
Rights and permissions
Copyright information
© 2015 Springer Science+Business Media, New York
About this protocol
Cite this protocol
Ribeiro, S.M., Giménez-Cassina, A., Danial, N.N. (2015). Measurement of Mitochondrial Oxygen Consumption Rates in Mouse Primary Neurons and Astrocytes. In: Palmeira, C., Rolo, A. (eds) Mitochondrial Regulation. Methods in Molecular Biology, vol 1241. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-1875-1_6
Download citation
DOI: https://doi.org/10.1007/978-1-4939-1875-1_6
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-1874-4
Online ISBN: 978-1-4939-1875-1
eBook Packages: Springer Protocols