Regulation of mitochondrial free Ca2+ by metabolite and pH‐dependent Ca2+ buffering in the matrix: analysis by a computational model of mitochondrial Ca2+ handling

Abstract

Mitochondrial free Ca2+ (mfCa2+) is regulated by ion fluxes through Ca2+ uniporter (CU), Na+/Ca2+ exchanger (NCE), Na+/H+ exchanger (NHE), and Ca2+/H+ exchanger (CHE) as well as by mitochondrial protein buffering. Regulation of mfCa2+ by metabolite and pH‐dependent dynamic Ca2+ buffering in the matrix during transient respiration, however, is not well known. To gain a quantitative understanding of these phenomena, we developed a biophysical model of mitochondrial Ca2+ handling and bioenergetics by developing and integrating kinetic models of the CU, NCE, NHE, and CHE into our well‐validated model of mitochondrial bioenergetics. Experiments were performed to spectrofluorometrically measure mfCa2+, pHm, membrane potential, and NADH redox state in guinea pig heart mitochondria suspended in Na+ and Ca2+ free buffer with 2.5 mM EGTA and 0.5 mM pyruvic acid (PA). Dynamics were inferred with addition of CaCl2, ADP, and CCCP with and without ruthenium red (CU blocker). Model analysis of the data on a) decreased mfCa2+ with addition of Na+‐independent substrate PA, and b) transient increases of mfCa2+ with added ADP and CCCP, implicates metabolite (ATP, ADP, Pi and TCA cycle intermediates) and pH‐dependent dynamic Ca2+ buffering in the cardiac mitochondrial matrix. The model will be helpful to understand mechanisms by which mfCa2+ both regulates, and is modulated by, mitochondrial energy metabolism.