Insulin Diminishes Superoxide Increase in Cytosol and Mitochondria of Cultured Cortical Neurons Treated with Toxic Glutamate

Glutamate excitotoxicity is involved in the pathogenesis of many disorders, including stroke, traumatic brain injury, and Alzheimer's disease, for which central insulin resistance is a comorbid condition. Neurotoxicity of glutamate (Glu) is primarily associated with hyperactivation of the ionotropic N-methyl-D-aspartate receptors (NMDARs), causing a sustained increase in intracellular free calcium concentration ([Ca2+](i)) and synchronous mitochondrial depolarization and an increase in intracellular superoxide anion radical (O-2(-center dot)) production. Recently, we found that insulin protects neurons against excitotoxicity by decreasing the delayed calcium deregulation (DCD). However, the role of insulin in O-2(-center dot) production in excitotoxicity still needs to be clarified. The present study aims to investigate insulin's effects on glutamate-evoked O-2(-center dot) generation and DCD using the fluorescent indicators dihydroethidium, MitoSOX Red, and Fura-FF in cortical neurons. We found a linear correlation between [Ca2+](i) and [O-2(-center dot)] in primary cultures of the rat neuron exposed to Glu, with insulin significantly reducing the production of intracellular and mitochondrial O-2(-center dot) in the primary cultures of the rat neuron. MK 801, an inhibitor of NMDAR-gated Ca2+ influx, completely abrogated the glutamate effects in both the presence and absence of insulin. In experiments in sister cultures, insulin diminished neuronal death and O-2 consumption rate (OCR).

Automated summary

Insulin decreases the production of intracellular and mitochondrial superoxide anion (O-2(-center dot)) in glutamate-induced excitotoxicity and reduces the delayed calcium deregulation (DCD). The NMDAR inhibitor MK 801 completely eliminates the effects of glutamate.