Mitochondrial free Ca²? levels and their effects on energy metabolism in Drosophila motor nerve terminals.

ABSTRACT: Mitochondrial Ca²? uptake exerts dual effects on mitochondria. Ca²? accumulation in the mitochondrial matrix dissipates membrane potential (??m), but Ca²? binding of the intramitochondrial enzymes accelerates oxidative phosphorylation, leading to mitochondrial hyperpolarization. The levels of matrix free Ca²? ([Ca²?]m) that trigger these metabolic responses in mitochondria in nerve terminals have not been determined. Here, we estimated [Ca²?]m in motor neuron terminals of Drosophila larvae using two methods: the relative responses of two chemical Ca²? indicators with a 20-fold difference in Ca²? affinity (rhod-FF and rhod-5N), and the response of a low-affinity, genetically encoded ratiometric Ca²? indicator (D4cpv) calibrated against known Ca²? levels. Matrix pH (pHm) and ??m were monitored using ratiometric pericam and tetramethylrhodamine ethyl ester probe, respectively, to determine when mitochondrial energy metabolism was elevated. At rest, [Ca²?]m was 0.22 ± 0.04 ?M, but it rose to ~26 ?M (24.3 ± 3.4 ?M with rhod-FF/rhod-5N and 27.0 ± 2.6 ?M with D4cpv) when the axon fired close to its endogenous frequency for only 2 s. This elevation in [Ca²?]m coincided with a rapid elevation in pHm and was followed by an after-stimulus ??m hyperpolarization. However, pHm decreased and no ??m hyperpolarization was observed in response to lower levels of [Ca²?]m, up to 13.1 ?M. These data indicate that surprisingly high levels of [Ca²?]m are required to stimulate presynaptic mitochondrial energy metabolism.

SUBMITTER: Ivannikov MV 

PROVIDER: S-EPMC3672877 | biostudies-other | 2013 Jun

REPOSITORIES: biostudies-other