title: Photo-induction and automated quantification of reversible mitochondrial permeability transition pore opening in primary mouse myotubes.
abstract: Opening of the mitochondrial permeability transition pore (mPTP) is involved in various cellular processes including apoptosis induction. Two distinct states of mPTP opening have been identified allowing the transfer of molecules with a molecular weight <1500 Da or <300 Da. The latter state is considered to be reversible and suggested to play a role in normal cell physiology. Here we present a strategy combining live-cell imaging and computer-assisted image processing allowing spatial visualization and quantitative analysis of reversible mPTP openings ("ΔΨ flickering") in primary mouse myotubes. The latter were stained with the photosensitive cation TMRM, which partitions between the cytosol and mitochondrial matrix as a function of mitochondrial membrane potential (ΔΨ). Controlled illumination of TMRM-stained primary mouse myotubes induced ΔΨ flickering in particular parts of the cell ("flickering domains"). A novel quantitative automated analysis was developed and validated to detect and quantify the frequency, size, and location of individual ΔΨ flickering events in myotubes.
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands; Institute for Molecules and Materials, Analytical Chemistry/Chemometrics, Radboud University Nijmegen, Nijmegen, The Netherlands; Centre for Systems Biology and Bioenergetics, Radboud University Medical Center, Nijmegen, The Netherlands.
authors.forename: Lionel
authors.initials: L
authors.lastname: Blanchet
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands; Centre for Systems Biology and Bioenergetics, Radboud University Medical Center, Nijmegen, The Netherlands.
authors.forename: Sander
authors.initials: S
authors.lastname: Grefte
authors.affiliation: Department of Paediatrics, Nijmegen Centre for Mitochondrial disorders, Radboud University Medical Center, Nijmegen, The Netherlands.
authors.forename: Jan A M
authors.initials: JA
authors.lastname: Smeitink
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands; Centre for Systems Biology and Bioenergetics, Radboud University Medical Center, Nijmegen, The Netherlands.
authors.forename: Peter H G M
authors.initials: PH
authors.lastname: Willems
authors.affiliation: Department of Biochemistry, Radboud Institute for Molecular Life Sciences, Radboud University Medical Center, Nijmegen, The Netherlands; Centre for Systems Biology and Bioenergetics, Radboud University Medical Center, Nijmegen, The Netherlands.
authors.forename: Werner J H
authors.initials: WJ
authors.lastname: Koopman
mesh_terms: D000818:Animals; D016572:Cyclosporine / Q000494:pharmacology; D005260:Female; D008027:Light; D051379:Mice; D008810:Mice, Inbred C57BL; D033681:Mitochondrial Membrane Transport Proteins / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects*; D000083162:Mitochondrial Permeability Transition Pore; D018485:Muscle Fibers, Skeletal / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects*; D009840:Oligomycins / Q000494:pharmacology
chemical_list: D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D009840:Oligomycins; C031004:oligomycin A; D016572:Cyclosporine