Ca2+ uptake in mitochondria occurs via the reverse action of the Na+/Ca2+ exchanger in metabolically inhibited MDCK cells.
In ischemic or hypoxic tissues, elevated Ca2+ levels have emerged as one of the main damaging agents among other Ca2+-independent mechanisms of cellular injury. Because mitochondria, besides the endoplasmic reticulum, play a key role in the maintainance of cellular Ca2+ homeostasis, alterations in the mitochondrial Ca2+ content ([Ca2+]m) were monitored in addition to changes in cytosolic Ca2+ concentration ([Ca2+]i) during metabolic inhibition (MI) in renal epithelial Madin-Darby canine kidney (MDCK) cells. [Ca2+]i and [Ca2+]m were monitored via, respectively, fura 2 and rhod 2 measurements. MI induced an increase in [Ca2+]i reaching 631+/-78 nM in approximately 20 min, followed by a decrease to 118+/-9 nM in the next approximately 25 min. A pronounced drop in cellular ATP levels and a rapid increase in intracellular Na+ concentrations in the first 20 min of MI excluded Ca2+ efflux in the second phase via plasma membrane ATPases or Na+/Ca2+ exchangers (NCE). Mitochondrial rhod 2 intensities increased to 434+/-46% of the control value during MI, indicating that mitochondria sequester Ca2+ during MI. The mitochondrial potential (deltapsim) was lost in 20 min of MI, excluding mitochondrial Ca2+ uptake via the deltapsim-dependent mitochondrial Ca2+ uniporter after 20 min of MI. Under Na+-free conditions, or when CGP-37157, a specific inhibitor of the mitochondrial NCE, was used, no drop in [Ca2+]i was seen during MI, whereas the MI-induced increase in mitochondrial rhod 2 fluorescence was strongly reduced. To our knowledge, this study is the first to report that in metabolically inhibited renal epithelial cells mitochondria take up Ca2+ via the NCE acting in the reverse mode.
Smets
Ilse
I
MBW-Dept. of Physiology, Limburgs Universitair Centrum/Transnationale Universiteit Limburg, Biomedisch Onderzoeksinstituut, Universitaire Campus Gebouw D, B-3590 Diepenbeek, Belgium. ilse.smets@luc.ac.be
Caplanusi
Adrian
A
Despa
Sanda
S
Molnar
Zsolt
Z
Radu
Mihai
M
VandeVen
Martin
M
Ameloot
Marcel
M
Steels
Paul
P
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D002460:Cell Line; D003600:Cytosol / Q000378:metabolism; D004285:Dogs; D004734:Energy Metabolism / Q000502:physiology*; D004847:Epithelial Cells / Q000166:cytology / Q000378:metabolism; D007684:Kidney Tubules / Q000166:cytology / Q000378:metabolism*; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000378:metabolism*; D019831:Sodium-Calcium Exchanger / Q000378:metabolism*
D019831:Sodium-Calcium Exchanger; D002118:Calcium