Delayed Mitochondrial Dysfunction in Excitotoxic Neuron Death: CytochromecRelease and a Secondary Increase in Superoxide Production

Abstract

An increased production of superoxide has been shown to mediate glutamate-induced neuron death. We monitored intracellular superoxide production of hippocampal neurons during and after exposure to the glutamate receptor agonist NMDA (300 microm). During a 30 min NMDA exposure, intracellular superoxide production increased significantly and remained elevated for several hours after wash-out of NMDA. After a 5 min exposure, superoxide production remained elevated for 10 min, but then rapidly returned to baseline. Mitochondrial membrane potential also recovered after wash-out of NMDA. However, recovery of mitochondria was transient and followed by delayed mitochondrial depolarization, loss of cytochrome c, and a secondary rise in superoxide production 4-8 hr after NMDA exposure. Treatment with a superoxide dismutase mimetic before the secondary rise conferred the same protection against cell death as a treatment before the first. The secondary rise could be inhibited by the complex I inhibitor rotenone (in combination with oligomycin) and mimicked by the complex III inhibitor antimycin A. To investigate the relationship between cytochrome c release and superoxide production, human D283 medulloblastoma cells deficient in mitochondrial respiration (rho(-) cells) were exposed to the apoptosis-inducing agent staurosporine. Treatment with staurosporine induced mitochondrial release of cytochrome c, caspase activation, and cell death in control and rho(-) cells. However, a delayed increase in superoxide production was only observed in control cells. Our data suggest that the delayed superoxide production in excitotoxicity and apoptosis occurs secondary to a defect in mitochondrial electron transport and that mitochondrial cytochrome c release occurs upstream of this defect.

AMP kinase–mediated activation of the BH3-only protein Bim couples energy depletion to stress-induced apoptosis

“…Conversely, if the ATP demand is less severe and the capacity of cells to generate ATP is sufficient, neurons restore their ionic gradients, and only a transient decrease in mitochondrial membrane potential ( m ) or cellular ATP levels can be detected. Nevertheless, many of these neurons proceed to undergo apoptotic death over the following hours Bonfoco et al, 1995;Luetjens et al, 2000;Ward et al, 2007).…”

“…Stimulation of glutamate receptors in cerebellar granular neurons (CGNs) with 100 µM glutamate/10 µM Gly for 10 min resulted in a delayed excitotoxic apoptosis within a 4-24-h time frame, which was characterized by cell shrinkage and nuclear pyknosis ( outer membrane permeabilization in excitotoxic apoptosis Budd et al, 2000;Luetjens et al, 2000;Ward et al, 2000Ward et al, , 2006Wang et al, 2004;Cheung et al, 2005). We next investigated whether the loss of Bim expression modulated these events.…”

“…Neurons attempt to restore their cellular ion homeostasis via the activity of ATP-consuming ion pumps, including Ca 2+ ATPases and the plasma membrane Na + K + ATPase, the driving force for Na + -Ca 2+ and Na + -H + exchangers (Brines and Robbins, 1992;Mironov, 1995). Importantly, the extent of ATP depletion and recovery of cellular bioenergetics critically determines the type of cell death that occurs Bonfoco et al, 1995;Luetjens et al, 2000;Ward et al, 2000). If the extent of ionic disturbances exceeds the capacity of the ATP-dependent ion pumps to restore ionic gradients, rapid cell swelling and necrosis occurs E xcitotoxicity after glutamate receptor overactivation induces disturbances in cellular ion gradients, resulting in necrosis or apoptosis.…”

Pharmacological inhibition of neuronal NADPH oxidase protects against 1-methyl-4-phenylpyridinium (MPP+)-induced oxidative stress and apoptosis in mesencephalic dopaminergic neuronal cells

“…SOD was used to demonstrate specificity of MPP + -induced ROS, which can be scavenged by SOD treatment. We and others have shown addition of exogenous SOD or the cell permeable SOD mimetic MnTBAP attenuated both generation of ROS and apoptosis in neuronal cells (Drukarch et al, 1998;Choi et al, 1999;Luetjens et al, 2000;Kitazawa et al, 2001;Anantharam et al, 2002;Kaul et al, 2003); hence, we used SOD not as an NADPH oxidase inhibitor but as a ROS inhibitor. Using flow cytometric analysis with the ROS-sensitive fluorescence probe hydroethidine, we show that the NADPH oxidase inhibitor AEBSF significantly blocked MPP + -induced increases in ROS production.…”

Ischemic Cardiomyopathy Following Seizure Induction by Domoic Acid

“…14 Overstimulation of ionotropic glutamate receptors (iGluR), namely N-methyl-D-aspartate (NMDA) receptors, provokes pathophysiological increases in intracellular Ca 2ϩ and consequently leads to the activation of several overlapping Ca 2ϩ -dependent processes, such as the generation of mitochondrial reactive oxygen species and loss of cellular viability. [15][16][17][18][19][20] DOM has been shown to be 8 to 10 times more potent than kainic acid in rodents, producing a similar reproducible pattern of behavioral toxicity culminating in seizures. 8,21,22 Studies conducted in rats, mice, and cynomolgus monkeys have shown a consistent pattern of DOM-induced damage to the hippocampal pyramidal neurons, as well as to the thalamic, amygdalar, entorhinal, cortical, and septal neurons after DOM administration (0.22 to 4.4 mg/kg i.p.).…”