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Confocal microscopy analysis of the activity of mitochondria contained within the ‘mitochondrial cloud’ during oogenesis in Xenopus laevis

Published online by Cambridge University Press:  28 November 2001

Martin Wilding
Affiliation:
Centre for Reproductive Biology, Clinica Villa del Sole, Via Manzoni, 15, 80126 Naples, Italy.
Rosa Carotenuto
Affiliation:
Department of Evolutionary and Comparative Anatomy, University of Naples ‘Federico II’, Via Mezzacanone, 8, 80134 Naples, Italy.
Vincenzo Infante
Affiliation:
Centre for Reproductive Biology, Clinica Villa del Sole, Via Manzoni, 15, 80126 Naples, Italy.
Brian Dale
Affiliation:
Centre for Reproductive Biology, Clinica Villa del Sole, Via Manzoni, 15, 80126 Naples, Italy.
Marcella Marino
Affiliation:
Centre for Reproductive Biology, Clinica Villa del Sole, Via Manzoni, 15, 80126 Naples, Italy.
Loredanna Di Matteo
Affiliation:
Centre for Reproductive Biology, Clinica Villa del Sole, Via Manzoni, 15, 80126 Naples, Italy.
Chiara Campanella
Affiliation:
Department of Evolutionary and Comparative Anatomy, University of Naples ‘Federico II’, Via Mezzacanone, 8, 80134 Naples, Italy.

Abstract

We have used ratiometric confocal microscopy and three fluorescence techniques to study the distribution and activity of mitochondria in frog oocytes during the early stages of oogenesis. Mitochondria in frog oocytes during oogenesis were characterised by a high ratio in the ‘mitochondrial cloud’ and peri-nuclear region and a low ratio in mitochondria freely dispersed within the cytoplasm. We tested whether the high ratio visualised by the three techniques represented mitochondrial membrane potential by perturbing the mitochondrial membrane potential. Carbonyl cyanide p-(trifluoromethyl)phenylhydrazone (FCCP) caused the immediate destruction of the membrane potential, and consequent loss of fluorescence from the membrane-potential-sensitive confocal channel. In contrast, nigericin caused an increase in membrane potential represented by a steady increase in fluorescence ratio. These data demonstrate that mitochondrial activity can be measured during oogenesis in frog oocytes, and suggest that the mitochondrial cloud and perinuclear regions are characterised by highly active mitochondria.

Type
Research Article
Copyright
2001 Cambridge University Press

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