Abstract
Intracellular Ca2+ is involved in control of a large variety of cell functions including apoptosis and neuron cell death . For example, intracellular Ca2+ overload is critical in neuron cell death induced by excitotoxicity . Thus, single cell monitoring of intracellular Ca2+ concentration ([Ca2+]cyt ) in neurons concurrently with apoptosis and neuron cell death is widely required.
Procedures for culture and preparation of primary cultures of hippocampal rat neurons and fluorescence imaging of cytosolic Ca2+ concentration in Fura2/AM -loaded neurons are described. We also describe a method for apoptosis detection by immunofluorescence imaging . Finally, a simple method for concurrent measurements of [Ca2+]cyt and apoptosis in the same neurons is described.
Access this chapter
Tax calculation will be finalised at checkout
Purchases are for personal use only
Similar content being viewed by others
References
Berridge MJ (2012) Calcium signalling remodelling and disease. Biochem Soc Trans 40:297–309
Sattler R, Tymianski M (2001) Molecular mechanisms of glutamate receptor-mediated excitotoxic neuronal cell death. Mol Neurobiol 24:107–129
Liu Y, Wong TP, Aarts M et al (2007) NMDA receptor subunits have differential roles in mediating excitotoxic neuronal death both in vitro and in vivo. J Neurosci 27:2846–2857
Sanz-Blasco S, Valero RA, Rodríguez-Crespo I et al (2008) Mitochondrial Ca2+ overload underlies Aβ oligomers neurotoxicity providing an unexpected mechanism of neuroprotection by NSAIDs. PLoS One 3:e2718
Brustovetsky T, Bolshakov A, Brustovetsky N (2010) Calpain activation and Na+/Ca2+ exchanger degradation occur downstream of calcium deregulation in hippocampal neurons exposed to excitotoxic glutamate. J Neurosci Res 88:1317–1328
Pivovarova NB, Nguyen HV, Winters CA et al (2004) Excitotoxic calcium overload in a subpopulation of mitochondria triggers delayed death in hippocampal neurons. J Neurosci 24:5611–5622
Jordán J, Ceña V, Prehn JH (2003) Mitochondrial control of neuron death and its role in neurodegenerative disorders. J Physiol Biochem 59:129–141
Villalobos C, Caballero E, Sanz-Blasco S et al (2012) Study of neurotoxic intracellular calcium signalling triggered by amyloids. Meth Mol Biol 849:289–302
Pérez-Otaño I, Luján R, Tavalin SJ et al (2006) Endocytosis and synaptic removal of NR3A-containing NMDA receptors by PACSIN1/syndapin1. Nat Neurosci 9:611–621
Grynkiewicz G, Poenie M, Tsien RY (1985) A new generation of Ca2+ indicators with greatly improved fluorescence properties. J Biol Chem 260:3440–3450
Zhou M, Baudry M (2006) Developmental changes in NMDA neurotoxicity reflect developmental changes in subunit composition of NMDA receptors. J Neurosci 26:2956–2963
Acknowledgments
This work was supported by Ministerio de Economía y Competi-tividad (Spain) grants BFU2009-08967 and BFU2012-37146 and Regional Government Junta de Castilla y León (Spain) grants BIO103/VA45/11, VA145U13, and BIO/VA33/13. María Calvo was supported by a predoctoral fellowship from Regional Government Junta de Castilla y León (Spain) and the European Social Fund.
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
Calvo, M., Villalobos, C., Núñez, L. (2015). Calcium Imaging in Neuron Cell Death. In: Lossi, L., Merighi, A. (eds) Neuronal Cell Death. Methods in Molecular Biology, vol 1254. Humana Press, New York, NY. https://doi.org/10.1007/978-1-4939-2152-2_6
Download citation
DOI: https://doi.org/10.1007/978-1-4939-2152-2_6
Published:
Publisher Name: Humana Press, New York, NY
Print ISBN: 978-1-4939-2151-5
Online ISBN: 978-1-4939-2152-2
eBook Packages: Springer Protocols