Spontaneous mitochondrial depolarizations are independent of SR Ca2+ release

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

Research output: Contribution to journalArticle

19 Citations (Scopus)

Abstract

The mitochondrial membrane potential (ΔΨm) underlies many mitochondrial functions, including Ca2+ influx into the mitochondria, which allows them to serve as buffers of intracellular Ca2+. Spontaneous depolarizations of ΔΨm, flickers, have been observed in isolated mitochondria and intact cells using the fluorescent cationic lipophile tetramethylrhodamine ethyl ester (TMRE), which distributes across the inner mitochondrial membrane in accordance with the Nernst equation. Flickers in cardiomyocytes have been attributed to uptake of Ca2+ released from the sarcoplasmic reticulum (SR) via ryanodine receptors in focal transients called Ca2+ sparks. We have shown previously that an increase in global Ca2+ in smooth muscle cells causes an increase in mitochondrial Ca2+ and depolarization of ΔΨm. Here we sought to determine whether flickers in smooth muscle cells are caused by uptake of Ca2+ released focally in Ca2+ sparks. High-speed three-dimensional imaging was used to monitor ΔΨm in freshly dissociated myocytes from toad stomach that were simultaneously voltage clamped at 0 mV to ensure the cytosolic TMRE concentration was constant and equal to the low level in the bath (2.5 nM). This approach allows quantitative analysis of flickers as we have previously demonstrated. Depletion of SR Ca2+ not only failed to eliminate flickers but rather increased their magnitude and frequency somewhat. Flickers were not altered in magnitude or frequency by ryanodine or xestospongin C, inhibitors of intracellular Ca2+ release, or by cyclosporin A, an inhibitor of the permeability transition pore. Focal Ca2+ release from the SR does not cause flickers in the cells employed here.
LanguageEnglish
PagesC1139-51
Number of pages13
JournalAmerican Journal of Physiology - Cell Physiology
Volume286
Issue number5
DOIs
Publication statusPublished - 2004

Fingerprint

Sarcoplasmic Reticulum
Smooth Muscle Myocytes
Mitochondria
Esters
Ryanodine
Ryanodine Receptor Calcium Release Channel
Three-Dimensional Imaging
Mitochondrial Membrane Potential
Mitochondrial Membranes
Baths
Cardiac Myocytes
Anura
Muscle Cells
Cyclosporine
Permeability
Stomach
Buffers
tetramethylrhodamine

Keywords

  • mitochondrial depolarization
  • SR Ca2+ release
  • ion channels

Cite this

O'Reilly, C.M. ; Fogarty, K.E. ; Drummond, R.M. ; Tuft, R.A. ; Walsh, J.V. / Spontaneous mitochondrial depolarizations are independent of SR Ca2+ release. In: American Journal of Physiology - Cell Physiology. 2004 ; Vol. 286, No. 5. pp. C1139-51.
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Spontaneous mitochondrial depolarizations are independent of SR Ca2+ release. / O'Reilly, C.M.; Fogarty, K.E.; Drummond, R.M.; Tuft, R.A.; Walsh, J.V.

In: American Journal of Physiology - Cell Physiology, Vol. 286, No. 5, 2004, p. C1139-51.

Research output: Contribution to journalArticle

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T1 - Spontaneous mitochondrial depolarizations are independent of SR Ca2+ release

AU - O'Reilly, C.M.

AU - Fogarty, K.E.

AU - Drummond, R.M.

AU - Tuft, R.A.

AU - Walsh, J.V.

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AB - The mitochondrial membrane potential (ΔΨm) underlies many mitochondrial functions, including Ca2+ influx into the mitochondria, which allows them to serve as buffers of intracellular Ca2+. Spontaneous depolarizations of ΔΨm, flickers, have been observed in isolated mitochondria and intact cells using the fluorescent cationic lipophile tetramethylrhodamine ethyl ester (TMRE), which distributes across the inner mitochondrial membrane in accordance with the Nernst equation. Flickers in cardiomyocytes have been attributed to uptake of Ca2+ released from the sarcoplasmic reticulum (SR) via ryanodine receptors in focal transients called Ca2+ sparks. We have shown previously that an increase in global Ca2+ in smooth muscle cells causes an increase in mitochondrial Ca2+ and depolarization of ΔΨm. Here we sought to determine whether flickers in smooth muscle cells are caused by uptake of Ca2+ released focally in Ca2+ sparks. High-speed three-dimensional imaging was used to monitor ΔΨm in freshly dissociated myocytes from toad stomach that were simultaneously voltage clamped at 0 mV to ensure the cytosolic TMRE concentration was constant and equal to the low level in the bath (2.5 nM). This approach allows quantitative analysis of flickers as we have previously demonstrated. Depletion of SR Ca2+ not only failed to eliminate flickers but rather increased their magnitude and frequency somewhat. Flickers were not altered in magnitude or frequency by ryanodine or xestospongin C, inhibitors of intracellular Ca2+ release, or by cyclosporin A, an inhibitor of the permeability transition pore. Focal Ca2+ release from the SR does not cause flickers in the cells employed here.

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T2 - American Journal of Physiology - Cell Physiology

JF - American Journal of Physiology - Cell Physiology

SN - 0363-6143

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