Journal of Bioenergetics and Biomembranes

, Volume 44, Issue 2, pp 253–263

Quantitative imaging of mitochondrial and cytosolic free zinc levels in an in vitro model of ischemia/reperfusion

  • Bryan J. McCranor
  • Rebecca A. Bozym
  • Michele I. Vitolo
  • Carol A. Fierke
  • Linda Bambrick
  • Brian M. Polster
  • Gary Fiskum
  • Richard B. Thompson
Article

DOI: 10.1007/s10863-012-9427-2

Cite this article as:
McCranor, B.J., Bozym, R.A., Vitolo, M.I. et al. J Bioenerg Biomembr (2012) 44: 253. doi:10.1007/s10863-012-9427-2

Abstract

The role of zinc ion in cytotoxicity following ischemic stroke, prolonged status epilepticus, and traumatic brain injury remains controversial, but likely is the result of mitochondrial dysfunction. We describe an excitation ratiometric fluorescence biosensor based on human carbonic anhydrase II variants expressed in the mitochondrial matrix, permitting free zinc levels to be quantitatively imaged therein. We observed an average mitochondrial matrix free zinc concentration of 0.2 pM in the PC12 rat pheochromacytoma cell culture line. Cytoplasmic and mitochondrial free zinc levels were imaged in a cellular oxygen glucose deprivation (OGD) model of ischemia/reperfusion. We observed a significant increase in mitochondrial zinc 1 h following 3 h OGD, at a time point when cytosolic zinc levels were depressed. Following the increase, mitochondrial zinc levels returned to physiological levels, while cytosolic zinc increased gradually over a 24 h time period in viable cells. The increase in intramitochondrial zinc observed during reoxygenation after OGD may contribute to bioenergetic dysfunction and cell death that occurs with both in vitro and in vivo models of reperfusion.

Keywords

Zinc Oxidative stress Mitochondria Ischemia Stroke Fluorescence biosensor Ratiometric Carbonic anhydrase II 

Copyright information

© Springer Science+Business Media, LLC 2012

Authors and Affiliations

  • Bryan J. McCranor
    • 1
  • Rebecca A. Bozym
    • 1
  • Michele I. Vitolo
    • 1
    • 2
  • Carol A. Fierke
    • 3
  • Linda Bambrick
    • 4
  • Brian M. Polster
    • 4
  • Gary Fiskum
    • 4
  • Richard B. Thompson
    • 1
  1. 1.Department of Biochemistry and Molecular BiologyUniversity of Maryland School of MedicineBaltimoreUSA
  2. 2.University of Maryland Marlene and Stewart Greenebaum NIH Cancer CenterUniversity of MarylandBaltimoreUSA
  3. 3.Department of ChemistryUniversity of MichiganAnn ArborUSA
  4. 4.Department of Anesthesiology and the Center for ShockTrauma, and Anesthesiology Research, University of Maryland School of MedicineBaltimoreUSA

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