Figure 11. Isolated mitochondrial studies III — calcium handling.

Mitochondrial calcium handling: Mitochondria were isolated in parallel from mutant and littermate control hearts. Extra- and intra-mitochondrial [Ca⁺²] were monitored simultaneously by fluorometry at room temperature, in freshly isolated, energized mitochondria (state 4 respiration with 5 mM glutamate/malate), using 0.1 μM Calcium Green-5N (λ_exc:505, λ_em:535 nm) and 20 μM Fura-FF (λ_exc:340 and 380 nm, λ_em:510 nm), respectively. Repeated CaCl₂ (5 μM) additions were performed until permeability transition pore (PTP) activation was detected. Mutant data were normalized to corresponding littermate control results. MyHC mutants demonstrated higher matrix intramitochondrial [Ca²⁺]_free, but PTP activation occurred at [Ca²⁺] similar to that of controls. TnT mutants had lower matrix [Ca²⁺]_free and PTP activation at lower [Ca²⁺] than controls. Data are presented as mean ± SD. n = 8 experiments from 5 mitochondrial preparations/10 mice in each group for control-M/MyHC, and n = 5 experiments from 5 mitochondrial preparations/10 mice in each group for control-T/TnT. *P < 0.05, using 2-sided 1-sample t test.