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dc.contributor.authorFuente Pérez, Sergio de la
dc.contributor.authorFonteriz García, Rosalba Inés 
dc.contributor.authorMontero Zoccola, María Teresa 
dc.contributor.authorÁlvarez Martín, Javier 
dc.date.accessioned2014-09-16T15:45:48Z
dc.date.available2014-09-16T15:45:48Z
dc.date.issued2012
dc.identifier.citationCell Calcium, 2012, vol. 51, p. 65-71es
dc.identifier.issn0143-4160es
dc.identifier.urihttp://uvadoc.uva.es/handle/10324/5993
dc.descriptionProducción Científicaes
dc.description.abstractAvailable methods to measure mitochondrial [Ca2+] ([Ca2+]M) include both targeted proteins and fluorescent dyes. Targeted proteins usually report much higher [Ca2+]M values than fluorescent dyes, up to two orders of magnitude. However, we show here that the low-Ca2+-affinity dye rhod-5N provides [Ca2+]M values similar to those reported by targeted aequorin, suggesting that the discrepancies are mainly due to the higher Ca2+-affinity of the fluorescent dyes used. We find rhod-5N has an apparent in situ intramitochondrial Kd around 0.5 mM. Addition of Ca2+ buffers containing between 4.5 and 10 M [Ca2+] to permeabilized cells loaded with rhod-5N induced increases in calibrated [Ca2+]M up to the 100 M–1 mM range, which were dependent on mitochondrial membrane potential. Ca2+ release from mitochondria was largely dependent on [Na+]. We have then used rhod-5N loaded cells to investigate the [Ca2+]M response to agonist stimulation at the single-cell and subcellular level.The [Ca2+]M peaks induced by histamine varied by nearly 10-fold among different cells, with a mean about 25 M. In the presence of the Ca2+ uniporter stimulator kaempferol, the [Ca2+]M peaks induced by histamine were also highly variable, and the mean [Ca2+]M peak was 3-fold higher. Simultaneous measurement of cytosolic and mitochondrial [Ca2+] peaks showed little correlation among the heights of the peaks in both compartments. Studying the [Ca2+]M peaks at the subcellular level, we found significant heterogeneities among regions in the same cell. In particular, the [Ca2+]M increase in mitochondrial regions close to the nucleus was more than double that of mitochondrial regions far from the nucleus.es
dc.format.mimetypeapplication/pdfes
dc.language.isoenges
dc.publisherElsevier Ltd.es
dc.rights.accessRightsinfo:eu-repo/semantics/openAccesses
dc.rights.urihttp://creativecommons.org/licenses/by-nc-nd/4.0/
dc.subjectMitocondriaes
dc.titleDynamics of mitochondrial [Ca2+] measured with the low-Ca2+-affinity dye rhod-5Nes
dc.typeinfo:eu-repo/semantics/articlees
dc.identifier.doi10.1016/j.ceca.2011.10.007es
dc.identifier.publicationfirstpage65es
dc.identifier.publicationlastpage71es
dc.identifier.publicationtitleCell Calciumes
dc.identifier.publicationvolume51es
dc.peerreviewedSIes
dc.rightsAttribution-NonCommercial-NoDerivatives 4.0 International


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