Quantitative analysis of spontaneous mitochondrial depolarizations

C.M. O'Reilly, K.E. Fogarty, R.M. Drummond, R.A. Tuft, J.V. Walsh

Research output: Contribution to journalArticle

96 Citations (Scopus)

Abstract

Spontaneous transient depolarizations in mitochondrial membrane potential ({Delta}{Psi}m), mitochondrial flickers, have been observed in isolated mitochondria and intact cells using the fluorescent probe, tetramethylrhodamine ethyl ester (TMRE). In theory, the ratio of [TMRE] in cytosol and mitochondrion allows {Delta}{Psi}m to be calculated with the Nernst equation, but this has proven difficult in practice due to fluorescence quenching and binding of dye to mitochondrial membranes. We developed a new method to determine the amplitude of flickers in terms of millivolts of depolarization. TMRE fluorescence was monitored using high-speed, high-sensitivity three-dimensional imaging to track individual mitochondria in freshly dissociated smooth muscle cells. Resting mitochondrial fluorescence, an exponential function of resting {Delta}{Psi}m, varied among mitochondria and was approximately normally distributed. Spontaneous changes in mitochondrial fluorescence, indicating depolarizations and repolarizations in {Delta}{Psi}m, were observed. The depolarizations were reversible and did not result in permanent depolarization of the mitochondria. The magnitude of the flickers ranged from <10 mV to >100 mV with a mean of 17.6 ± 1.0 mV (n = 360) and a distribution skewed to smaller values. Nearly all mitochondria flickered, and they did so independently of one another, indicating that mitochondria function as independent units in the myocytes employed here.
Original languageEnglish
Pages (from-to)3350-3357
Number of pages7
JournalBiophysical Journal
Volume85
Issue number5
Publication statusPublished - 2003

Fingerprint

Mitochondria
Fluorescence
Esters
Three-Dimensional Imaging
Mitochondrial Membrane Potential
Mitochondrial Membranes
Fluorescent Dyes
Cytosol
Muscle Cells
Smooth Muscle Myocytes
Coloring Agents
tetramethylrhodamine

Keywords

  • mitochondrial depolarizations
  • biophysics
  • bioscience
  • mitochondria

Cite this

O'Reilly, C. M., Fogarty, K. E., Drummond, R. M., Tuft, R. A., & Walsh, J. V. (2003). Quantitative analysis of spontaneous mitochondrial depolarizations. Biophysical Journal, 85(5), 3350-3357.
O'Reilly, C.M. ; Fogarty, K.E. ; Drummond, R.M. ; Tuft, R.A. ; Walsh, J.V. / Quantitative analysis of spontaneous mitochondrial depolarizations. In: Biophysical Journal. 2003 ; Vol. 85, No. 5. pp. 3350-3357.
@article{bc1bb02fc7474e3398973868b511abd5,
title = "Quantitative analysis of spontaneous mitochondrial depolarizations",
abstract = "Spontaneous transient depolarizations in mitochondrial membrane potential ({Delta}{Psi}m), mitochondrial flickers, have been observed in isolated mitochondria and intact cells using the fluorescent probe, tetramethylrhodamine ethyl ester (TMRE). In theory, the ratio of [TMRE] in cytosol and mitochondrion allows {Delta}{Psi}m to be calculated with the Nernst equation, but this has proven difficult in practice due to fluorescence quenching and binding of dye to mitochondrial membranes. We developed a new method to determine the amplitude of flickers in terms of millivolts of depolarization. TMRE fluorescence was monitored using high-speed, high-sensitivity three-dimensional imaging to track individual mitochondria in freshly dissociated smooth muscle cells. Resting mitochondrial fluorescence, an exponential function of resting {Delta}{Psi}m, varied among mitochondria and was approximately normally distributed. Spontaneous changes in mitochondrial fluorescence, indicating depolarizations and repolarizations in {Delta}{Psi}m, were observed. The depolarizations were reversible and did not result in permanent depolarization of the mitochondria. The magnitude of the flickers ranged from <10 mV to >100 mV with a mean of 17.6 ± 1.0 mV (n = 360) and a distribution skewed to smaller values. Nearly all mitochondria flickered, and they did so independently of one another, indicating that mitochondria function as independent units in the myocytes employed here.",
keywords = "mitochondrial depolarizations, biophysics, bioscience, mitochondria",
author = "C.M. O'Reilly and K.E. Fogarty and R.M. Drummond and R.A. Tuft and J.V. Walsh",
year = "2003",
language = "English",
volume = "85",
pages = "3350--3357",
journal = "Biophysical Journal",
issn = "0006-3495",
number = "5",

}

O'Reilly, CM, Fogarty, KE, Drummond, RM, Tuft, RA & Walsh, JV 2003, 'Quantitative analysis of spontaneous mitochondrial depolarizations', Biophysical Journal, vol. 85, no. 5, pp. 3350-3357.

Quantitative analysis of spontaneous mitochondrial depolarizations. / O'Reilly, C.M.; Fogarty, K.E.; Drummond, R.M.; Tuft, R.A.; Walsh, J.V.

In: Biophysical Journal, Vol. 85, No. 5, 2003, p. 3350-3357.

Research output: Contribution to journalArticle

TY - JOUR

T1 - Quantitative analysis of spontaneous mitochondrial depolarizations

AU - O'Reilly, C.M.

AU - Fogarty, K.E.

AU - Drummond, R.M.

AU - Tuft, R.A.

AU - Walsh, J.V.

PY - 2003

Y1 - 2003

N2 - Spontaneous transient depolarizations in mitochondrial membrane potential ({Delta}{Psi}m), mitochondrial flickers, have been observed in isolated mitochondria and intact cells using the fluorescent probe, tetramethylrhodamine ethyl ester (TMRE). In theory, the ratio of [TMRE] in cytosol and mitochondrion allows {Delta}{Psi}m to be calculated with the Nernst equation, but this has proven difficult in practice due to fluorescence quenching and binding of dye to mitochondrial membranes. We developed a new method to determine the amplitude of flickers in terms of millivolts of depolarization. TMRE fluorescence was monitored using high-speed, high-sensitivity three-dimensional imaging to track individual mitochondria in freshly dissociated smooth muscle cells. Resting mitochondrial fluorescence, an exponential function of resting {Delta}{Psi}m, varied among mitochondria and was approximately normally distributed. Spontaneous changes in mitochondrial fluorescence, indicating depolarizations and repolarizations in {Delta}{Psi}m, were observed. The depolarizations were reversible and did not result in permanent depolarization of the mitochondria. The magnitude of the flickers ranged from <10 mV to >100 mV with a mean of 17.6 ± 1.0 mV (n = 360) and a distribution skewed to smaller values. Nearly all mitochondria flickered, and they did so independently of one another, indicating that mitochondria function as independent units in the myocytes employed here.

AB - Spontaneous transient depolarizations in mitochondrial membrane potential ({Delta}{Psi}m), mitochondrial flickers, have been observed in isolated mitochondria and intact cells using the fluorescent probe, tetramethylrhodamine ethyl ester (TMRE). In theory, the ratio of [TMRE] in cytosol and mitochondrion allows {Delta}{Psi}m to be calculated with the Nernst equation, but this has proven difficult in practice due to fluorescence quenching and binding of dye to mitochondrial membranes. We developed a new method to determine the amplitude of flickers in terms of millivolts of depolarization. TMRE fluorescence was monitored using high-speed, high-sensitivity three-dimensional imaging to track individual mitochondria in freshly dissociated smooth muscle cells. Resting mitochondrial fluorescence, an exponential function of resting {Delta}{Psi}m, varied among mitochondria and was approximately normally distributed. Spontaneous changes in mitochondrial fluorescence, indicating depolarizations and repolarizations in {Delta}{Psi}m, were observed. The depolarizations were reversible and did not result in permanent depolarization of the mitochondria. The magnitude of the flickers ranged from <10 mV to >100 mV with a mean of 17.6 ± 1.0 mV (n = 360) and a distribution skewed to smaller values. Nearly all mitochondria flickered, and they did so independently of one another, indicating that mitochondria function as independent units in the myocytes employed here.

KW - mitochondrial depolarizations

KW - biophysics

KW - bioscience

KW - mitochondria

UR - http://www.biophysj.org/cgi/content/abstract/85/5/3350

M3 - Article

VL - 85

SP - 3350

EP - 3357

JO - Biophysical Journal

JF - Biophysical Journal

SN - 0006-3495

IS - 5

ER -

O'Reilly CM, Fogarty KE, Drummond RM, Tuft RA, Walsh JV. Quantitative analysis of spontaneous mitochondrial depolarizations. Biophysical Journal. 2003;85(5):3350-3357.