Cyclosporin A Increases Mitochondrial Buffering of Calcium: An Additional Mechanism in Delaying Mitochondrial Permeability Transition Pore Opening

Regulation of mitochondrial free Ca(2+) is critically important for cellular homeostasis. An increase in mitochondrial matrix free Ca(2+) concentration ([Ca(2+)](m)) predisposes mitochondria to opening of the permeability transition pore (mPTP). Opening of the pore can be delayed by cyclosporin A (CsA), possibly by inhibiting cyclophilin D (Cyp D), a key regulator of mPTP. Here, we report on a novel mechanism by which CsA delays mPTP opening by enhanced sequestration of matrix free Ca(2+). Cardiac-isolated mitochondria were challenged with repetitive CaCl(2) boluses under Na(+)-free buffer conditions with and without CsA. CsA significantly delayed mPTP opening primarily by promoting matrix Ca(2+) sequestration, leading to sustained basal [Ca(2+)](m) levels for an extended period. The preservation of basal [Ca(2+)](m) during the CaCl(2) pulse challenge was associated with normalized NADH, matrix pH (pH(m)), and mitochondrial membrane potential (ΔΨ(m)). Notably, we found that in PO(4)(3−) (P(i))-free buffer condition, the CsA-mediated buffering of [Ca(2+)](m) was abrogated, and mitochondrial bioenergetics variables were concurrently compromised. In the presence of CsA, addition of P(i) just before pore opening in the P(i)-depleted condition reinstated the Ca(2+) buffering system and rescued mitochondria from mPTP opening. This study shows that CsA promotes P(i)-dependent mitochondrial Ca(2+) sequestration to delay mPTP opening and, concomitantly, maintains mitochondrial function.