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Identification and investigation of mitochondria lacking cytochrome c oxidase activity in axons

Lookup NU author(s): Jessica Zambonin, Dr Sarah Engeham, Graham Campbell, Dr Iryna Ziabreva, Nina Beadle, Professor Robert Taylor, Dr Don Mahad

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Abstract

Mitochondrial defects have been implicated in the degeneration of axons in a number of CNS disorders, including multiple sclerosis. Uniquely, mitochondria harbor the only non-nuclear DNA (mitochondrial DNA or mtDNA), which encodes functionally important subunits of the respiratory chain. The pattern of mitochondrial respiratory chain subunit expression provides important clues to the underlying mechanism of mitochondrial injury. In snap frozen tissue mitochondrial respiratory chain complex IV or cytochrome c oxidase (COX) activity may be determined using a well-established histochemical technique. COX histochemistry. Lack of COX activity may be the result of mtDNA mutations, degradation of transcripts of subunits, modification of subunits or inhibition of complexes. Mitochondria lacking complex IV activity, however, have not been further explored within axons in CNS disorders. By combining COX histochemistry with immunofluorescent labeling of mitochondrial proteins we describe a method to identify mitochondria lacking complex IV activity in CNS tissue and locate inactive mitochondria to axons using confocal microscopy. Inactive axonal mitochondria may then be further investigated using confocal microscopy to define the pattern of mitochondrial respiratory chain complex subunit expression. Our technique may be used to gain important clues to the underlying mechanisms of mitochondrial injury within axons in a number of CNS disorders and relevant animal models. (C) 2010 Elsevier B.V. All rights reserved.

Publication metadata

Author(s): Zambonin J, Engeham S, Campbell GR, Ziabreva I, Beadle NE, Taylor RW, Mahad DJ

Publication type: Article

Publication status: Published

Journal: Journal of Neuroscience Methods

Year: 2010

Volume: 192

Issue: 1

Pages: 115-120

Print publication date: 24/07/2010

ISSN (print): 0165-0270

ISSN (electronic): 1872-678X

Publisher: Elsevier BV

URL: http://dx.doi.org/10.1016/j.jneumeth.2010.07.022

DOI: 10.1016/j.jneumeth.2010.07.022

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