Mitochondrial modulation of Ca2+ sparks and transient KCa currents in smooth muscle cells of rat cerebral arteries

Mitochondria sequester and release calcium (Ca2+) and regulate intracellular Ca2+ concentration ([Ca2+]i) in eukaryotic cells. However, the regulation of different Ca2+ signalling modalities by mitochondria in smooth muscle cells is poorly understood. Here, we investigated the regulation of Ca2+ sparks, Ca2+ waves and global [Ca2+]i by mitochondria in cerebral artery smooth muscle cells. CCCP (a protonophore; 1 μm) and rotenone (an electron transport chain complex I inhibitor; 10 μm) depolarized mitochondria, reduced Ca2+ spark and wave frequency, and elevated global [Ca2+]i in smooth muscle cells of intact arteries. In voltage-clamped (−40 mV) cells, mitochondrial depolarization elevated global [Ca2+]i, reduced Ca2+ spark amplitude, spatial spread and the effective coupling of sparks to large-conductance Ca2+-activated potassium (KCa) channels, and decreased transient KCa current frequency and amplitude. Inhibition of Ca2+ sparks and transient KCa currents by mitochondrial depolarization could not be explained by a decrease in intracellular ATP or a reduction in sarcoplasmic reticulum Ca2+ load, and occurred in the presence of diltiazem, a voltage-dependent Ca2+ channel blocker. Ru360 (10 μm), a mitochondrial Ca2+ uptake blocker, and lonidamine (100 μm), a permeability transition pore (PTP) opener, inhibited transient KCa currents similarly to mitochondrial depolarization. In contrast, CGP37157 (10 μm), a mitochondrial Na+–Ca2+ exchange blocker, activated these events. The PTP blockers bongkrekic acid and cyclosporin A both reduced inhibition of transient KCa currents by mitochondrial depolarization. These results indicate that mitochondrial depolarization leads to a voltage-independent elevation in global [Ca2+]i and Ca2+ spark and transient KCa current inhibition. Data also suggest that mitochondrial depolarization inhibits Ca2+ sparks and transient KCa currents via PTP opening and a decrease in intramitochondrial [Ca2+].