Ca2+ Dysregulation Induces Mitochondrial Depolarization and Apoptosis

Abstract

We previously reported that constitutively activated Gαq (Q209L) expression in cardiomyocytes induces apoptosis through opening of the mitochondrial permeability transition pore. We assessed the hypothesis that disturbances in Ca2+ handling linked Gαq activity to apoptosis because resting Ca2+ levels were significantly increased prior to development of apoptosis. Treating cells with EGTA lowered Ca 2+ and blocked both loss of mitochondrial membrane potential (an indicator of permeability transition pore opening) and apoptosis (assessed by DNA fragmentation). When cytosolic Ca2+ and mitochondrial membrane potential were simultaneously measured by confocal microscopy, sarcoplasmic reticulum (SR)-driven slow Ca2+ oscillations (time-to-peak ∼4 s) were observed in Q209L-expressing cells. These oscillations were seen to transition into sustained increases in cytosolic Ca2+, directly paralleled by loss of mitochondrial membrane potential. Ca2+ transients generated by caffeine-induced release of SR Ca2+ were greatly prolonged in Q209L-expressing cells, suggesting a decreased ability to extrude Ca2+. Indeed, the Na+/Ca2+ exchanger (NCX), which removes Ca2+ from the cell, was markedly down-regulated at the mRNA and protein levels. Adenoviral NCX expression normalized cytosolic Ca2+ levels and prevented DNA fragmentation in cells expressing Q209L. Interestingly, constitutively activated Akt, which rescues cells from Q209L-induced apoptosis, prevented the decrease in NCX expression, normalized cytosolic Ca2+ levels and spontaneous Ca2+ oscillations, shortened caffeine-induced Ca2+ transients, and prevented loss of the mitochondrial membrane potential. Our findings demonstrate that NCX down-regulation and consequent increases in cytosolic and SR Ca2+ can lead to Ca2+ overloading-induced loss of mitochondrial membrane potential and suggest that recovery of Ca2+ dysregulation is a target of Akt-mediated protection. © 2005 by The American Society for Biochemistry and Molecular Biology, Inc.