Mitochondrial membrane potential modulates regulation of mitochondrial Ca2+ in rat ventricular myocytes.
Although recent studies focused on the contribution of mitochondrial Ca2+ to the mechanisms of ischemia-reperfusion injury, the regulation of mitochondrial Ca2+ under pathophysiological conditions remains largely unclear. By using saponin-permeabilized rat myocytes, we measured mitochondrial membrane potential (DeltaPsi(m)) and mitochondrial Ca2+ concentration ([Ca2+](m)) at the physiological range of cytosolic Ca2+ concentration ([Ca2+](c); 300 nM) and investigated the regulation of [Ca2+](m) during both normal and dissipated DeltaPsi(m). When DeltaPsi(m) was partially depolarized by carbonyl cyanide p-(trifluoromethoxy)phenylhydrazone (FCCP, 0.01-0.1 microM), there were dose-dependent decreases in [Ca2+](m). When complete DeltaPsi(m) dissipation was achieved by FCCP (0.3-1 microM), [Ca2+](m) remained at one-half of the control level despite no Ca2+ influx via the Ca2+ uniporter. The DeltaPsi(m) dissipation by FCCP accelerated calcein leakage from mitochondria in a cyclosporin A (CsA)-sensitive manner, which indicates that DeltaPsi(m) dissipation opened the mitochondrial permeability transition pore (mPTP). After FCCP addition, inhibition of the mPTP by CsA caused further [Ca2+](m) reduction; however, inhibition of mitochondrial Na+/Ca2+ exchange (mitoNCX) by a Na+-free solution abolished this [Ca2+](m) reduction. Cytosolic Na(+) concentrations that yielded one-half maximal activity levels for mitoNCX were 3.6 mM at normal DeltaPsi(m) and 7.6 mM at DeltaPsi(m) dissipation. We conclude that 1) the mitochondrial Ca2+ uniporter accumulates Ca2+ in a manner that is dependent on DeltaPsi(m) at the physiological range of [Ca2+](c); 2) DeltaPsi(m) dissipation opens the mPTP and results in Ca2+ influx to mitochondria; and 3) although mitoNCX activity is impaired, mitoNCX extrudes Ca2+ from the matrix even after DeltaPsi(m) dissipation.
Saotome
Masao
M
Division of Cardiology, Internal Medicine III, Hamamatsu University School of Medicine, 1-20-1 Handayama, Hamamatsu 431-3192, Japan.
Katoh
Hideki
H
Satoh
Hiroshi
H
Nagasaka
Shiro
S
Yoshihara
Shu
S
Terada
Hajime
H
Hayashi
Hideharu
H
D000818:Animals; D002118:Calcium / Q000378:metabolism*; D015220:Calcium Channels; D002135:Calcium-Binding Proteins / Q000378:metabolism; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006352:Heart Ventricles / Q000166:cytology; D006575:Heterocyclic Compounds, 3-Ring; D007425:Intracellular Membranes / Q000502:physiology; D007473:Ion Channels / Q000378:metabolism; D008297:Male; D008564:Membrane Potentials / Q000502:physiology; D008856:Microscopy, Fluorescence / Q000379:methods; D008928:Mitochondria / Q000502:physiology*; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D032383:Myocytes, Cardiac / Q000502:physiology*; D051381:Rats; D017207:Rats, Sprague-Dawley; D019831:Sodium-Calcium Exchanger / Q000378:metabolism
D015220:Calcium Channels; D002135:Calcium-Binding Proteins; D005452:Fluoresceins; D005456:Fluorescent Dyes; D006575:Heterocyclic Compounds, 3-Ring; D007473:Ion Channels; D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D019831:Sodium-Calcium Exchanger; C034703:mitochondrial calcium uniporter; C068483:rhod-2; D002118:Calcium; C007740:fluorexon