Comparative analysis of cytosolic and mitochondrial ATP synthesis in embryonic and postnatal hippocampal neuronal cultures.

ATP in neurons is commonly believed to be synthesized mostly by mitochondria via oxidative phosphorylation. Neuronal mitochondria have been studied primarily in culture, i.e.

Store-operated Orai1 and IP3 receptor-operated TRPC1 channel.

Store-operated channels (SOC) are known to be physiologically activated following agonist-induced IP3 production and depletion of Ca2+ stores. Here we present molecular,biophysical and mechanistic evidence that two ubiquitously expressed plasma membrane channels may be responsible for creating a complex and sometimes controversial SOC image: one being a real SOC encoded by Orai1 and activated exclusively upon depletion of Ca2+ stores (via iPLA2beta -dependent pathway), while the second one is an IP3 receptor-operated channel (IP3ROC) encoded by TRPC1 and activated via its conformational coupling with IP3 receptor. In RBL-2H3 cells endogenously expressing Orai1 and TRPC1, we unmasked and characterized whole-cell current through IP3ROC channels that was hiding behind some familiar fingerprints of ICRAC, a current through the classical Ca2+-selective SOC (CRAC) channels.

Measurements of mitochondrial pH in cultured cortical neurons clarify contribution of mitochondrial pore to the mechanism of glutamate-induced delayed Ca2+ deregulation.

To clarify the role of the mitochondrial permeability transition pore (MPT) in the mechanism of the glutamate-induced delayed calcium deregulation (DCD) and mitochondrial depolarization (MD), we studied changes in cytosolic (pH(c)) and mitochondrial pH (pH(m)) induced by glutamate in cultured cortical neurons expressing pH-sensitive fluorescent proteins. We found that DCD and MD were associated with a prominent pH(m) decrease which presumably resulted from MPT opening. This pH(m) decrease occurred with some delay after the onset of DCD and MD.

Mitochondrial Ca2+ cycle mediated by the palmitate-activated cyclosporin A-insensitive pore.

Earlier we found that in isolated rat liver mitochondria the reversible opening of the mitochondrial cyclosporin A-insensitive pore induced by low concentrations of palmitic acid (Pal) plus Ca(2+) results in the brief loss of Deltapsi [Mironova et al., J Bioenerg Biomembr (2004), 36:171-178]. Now we report that Pal and Ca(2+), increased to 30 and 70 nmol/mg protein respectively, induce a stable and prolonged (10 min) partial depolarization of the mitochondrial membrane, the release of Ca(2+) and the swelling of mitochondria.