Extra-matrix Mg2+ limits Ca2+ uptake and modulates Ca2+ uptake–independent respiration and redox state in cardiac isolated mitochondria

Cardiac mitochondrial matrix (m) free Ca 2+ ([Ca 2+ ] m ) increases primarily by Ca 2+ uptake through the Ca 2+ uniporter (CU). Ca 2+ uptake via the CU is attenuated by extra-matrix (e) Mg 2+ ([Mg 2+ ] e ). How [Ca 2+ ] m is dynamically modulated by interacting physiological levels of [Ca 2+ ] e and [Mg 2+ ] e and how this interaction alters bioenergetics are not well understood. We postulated that as [Mg 2+ ] e modulates Ca 2+ uptake via the CU, it also alters bioenergetics in a matrix Ca 2+ –induced and matrix Ca 2+ –independent manner. To test this, we measured changes in [Ca 2+ ] e , [Ca 2+ ] m , [Mg 2+ ] e and [Mg 2+ ] m spectrofluorometrically in guinea pig cardiac mitochondria in response to added CaCl 2 (0–0.6 mM; 1 mM EGTA buffer) with/without added MgCl 2 (0–2 mM). In parallel, we assessed effects of added CaCl 2 and MgCl 2 on NADH, membrane potential (ΔΨ m ), and respiration. We found that > 0.125 mM MgCl 2 significantly attenuated CU-mediated Ca 2+ uptake and [Ca 2+ ] m . Incremental [Mg 2+ ] e did not reduce initial Ca 2+ uptake but attenuated the subsequent slower Ca 2+ uptake, so that [Ca 2+ ] m remained unaltered over time. Adding CaCl 2 without MgCl 2 to attain a [Ca 2+ ] m from 46 to 221 nM enhanced state 3 NADH oxidation and increased respiration by 15 %; up to 868 nM [Ca 2+ ] m did not additionally enhance NADH oxidation or respiration. Adding MgCl 2 did not increase [Mg 2+ ] m but it altered bioenergetics by its direct effect to decrease Ca 2+ uptake. However, at a given [Ca 2+ ] m , state 3 respiration was incrementally attenuated, and state 4 respiration enhanced, by higher [Mg 2+ ] e . Thus, [Mg 2+ ] e without a change in [Mg 2+ ] m can modulate bioenergetics independently of CU-mediated Ca 2+ transport.