Strong calcium entry activates mitochondrial superoxide generation, upregulating kinase signaling in hippocampal neurons

Large increases in cytosolic free Ca2+ ([Ca2+](i)) activate several kinases that are important for neuronal plasticity, including Ca2+/calmodulin-dependent kinase II (CaMKII), protein kinase A (PKA), and protein kinase C (PKC). Because it is also known, mainly in non-neuronal systems, that superoxide radicals (O-2(-)) activate these (and other) kinases and because O-2(-) generation by mitochondria is in part [Ca2+](i) dependent, we examined in hippocampal neurons the relationship between Ca2+ entry, O-2(-) production, and kinase activity. We found that, after large stimulus-induced [Ca2+](i) increases, O-2(-) selectively produced by mitochondria near plasmalemmal sites of Ca2+ entry acts as a modulator to upregulate the two kinases, namely, CaMKII and PKA, whose activities are directly or indirectly phosphorylation dependent. The common mechanism involves O-2(-) inhibition of inactivating protein phosphatases. Conversely, because small [Ca2+](i) increases do not promote mitochondrial respiration and O-2(-) generation, weak stimuli favor enhanced phosphatase activity, which therefore leads to suppressed kinase activity. Enhanced O-2(-) production also promoted PKC activity but by a phosphatase-independent pathway. These results suggest that Ca2+-dependent upregulation of mitochondrial O-2(-) production may be a general mechanism for linking Ca2+ entry to enhanced kinase activity and therefore to synaptic plasticity. This mechanism also represents yet another way that mitochondria, acting as calcium sensors, can play a role in neuronal signal transduction.