Mitochondrial inner membrane electrophysiology assessed by rhodamine-123 transport and fluorescence. Ann Biomed Eng 2007 Jul;35(7):1276-85
Date
03/21/2007Pubmed ID
17372838Pubmed Central ID
PMC3508792DOI
10.1007/s10439-007-9265-2Scopus ID
2-s2.0-34250736372 (requires institutional sign-in at Scopus site) 71 CitationsAbstract
Rhodamine-123 is widely used to make dynamic measurements of mitochondrial membrane potential both in vitro and in situ. Yet data interpretation is difficult due to a lack of quantitative understanding of how membrane potential and measured fluorescence are related. To develop such understanding, a model for dye transport across the mitochondrial inner membrane and partition into the membrane was developed. The model accounts for experimentally measured dye self-quenching and was integrated into a model of mitochondrial electrophysiology to estimate transients in mitochondrial membrane potential from kinetic fluorescence measurements. Our analysis indicates that (i) R123 fluorescence peaks at concentrations near 50 microM due to self-quenching; (ii) measured fluorescence intensity and membrane potential are related by a non-linear calibration curve sensitive to certain experimental details, including total concentration of dye and mitochondria in suspensions; and (iii) the time courses of membrane potential and electron transport fluxes following a perturbation (i.e. addition of ADP) significantly differ from observed transients in fluorescence intensity. These findings are consistent with the model predictions that mitochondria display a characteristic time of response to changes in substrate concentration of less than 0.1 s, corresponding to the time scale over which the rate of ATP synthesis changes to meet changes in ADP concentration.
Author List
Huang M, Camara AK, Stowe DF, Qi F, Beard DAAuthor
David F. Stowe MD, PhD Professor in the Anesthesiology department at Medical College of WisconsinMESH terms used to index this publication - Major topics in bold
Adenosine DiphosphateAdenosine Triphosphate
Animals
Biological Transport
Female
Guinea Pigs
Male
Membrane Potential, Mitochondrial
Mitochondria, Heart
Mitochondrial Membranes
Models, Biological
Rhodamine 123
