N-methyl-D-aspartate receptor-mediated mitochondrial Ca(2+) overload in acute excitotoxic motor neuron death: a mechanism distinct from chronic neurotoxicity after Ca(2+) influx.
Mitochondrial uptake of Ca(2+) has recently been found to play an important role in glutamate-induced neurotoxicity (GNT) as well as in the activation of Ca(2+)-dependent molecules, such as calmodulin and neuronal nitric oxide synthase (nNOS), in the cytoplasm. Prolonged exposure to glutamate injures motor neurons predominantly through the activation of Ca(2+)/calmodulin-nNOS, as previously reported, and is, in part, associated with the pathogenesis of amyotrophic lateral sclerosis (ALS). In the present study, we investigated how mitochondrial uptake of Ca(2+) is involved in GNT in spinal motor neurons. Acute excitotoxicity induced by exposure to 0.5 mM glutamate for 5 min was found in both motor and nonmotor neurons in cultured spinal cords from rat embryos and was dependent on extracellular Ca(2+) and on N-methyl-D-aspartate (NMDA) receptor activation. Mitochondrial uncouplers markedly blocked acute excitotoxicity, and membrane-permeable superoxide dismutase mimics attenuated acute excitotoxicity induced by glutamate and NMDA but not by alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) or kainate. Fluorimetric analysis showed that mitochondrial Ca(2+) was elevated promptly with subsequent accumulation of reactive oxygen species (ROS) in the mitochondria. An NMDA receptor antagonist and a mitochondrial uncoupler eliminated the increase in fluorescence of mitochondrial Ca(2+) and ROS indicators. These data indicate that acute excitotoxicity in spinal neurons is mediated by mitochondrial Ca(2+) overload and ROS generation through the activation of NMDA receptors. This mechanism is different from that of chronic GNT.
Department of Neurology, Graduate School of Medicine, Kyoto University, Sakyoku, Kyoto, Japan.
M
M
Urushitani
T
T
Nakamizo
R
R
Inoue
H
H
Sawada
T
T
Kihara
K
K
Honda
A
A
Akaike
S
S
Shimohama
D019297:2,4-Dinitrophenol / Q000494:pharmacology; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione / Q000494:pharmacology; D000818:Animals; D002118:Calcium / Q000378:metabolism*; D020013:Calcium Signaling / Q000502:physiology*; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002478:Cells, Cultured / Q000187:drug effects; D016572:Cyclosporine / Q000494:pharmacology; D003992:Dibucaine / Q000494:pharmacology; D016291:Dizocilpine Maleate / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D018690:Excitatory Amino Acid Agonists / Q000494:pharmacology; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D005452:Fluoresceins; D005456:Fluorescent Dyes; D018698:Glutamic Acid / Q000494:pharmacology / Q000633:toxicity*; D006575:Heterocyclic Compounds, 3-Ring; D007093:Imidazoles / Q000494:pharmacology; D007608:Kainic Acid / Q000494:pharmacology; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism*; D016472:Motor Neuron Disease / Q000378:metabolism; D009046:Motor Neurons / Q000187:drug effects* / Q000473:pathology; D016202:N-Methylaspartate / Q000494:pharmacology; D009419:Nerve Tissue Proteins / Q000378:metabolism; D009474:Neurons / Q000187:drug effects; D009498:Neurotoxins / Q000494:pharmacology / Q000633:toxicity*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052248:Nitric Oxide Synthase Type I; D018384:Oxidative Stress; D051381:Rats; D017208:Rats, Wistar; D017382:Reactive Oxygen Species; D018091:Receptors, AMPA / Q000187:drug effects; D018092:Kainic Acid Receptors / Q000187:drug effects; D016194:Receptors, N-Methyl-D-Aspartate / Q000502:physiology*; D012235:Rhodamines; D013116:Spinal Cord / Q000166:cytology / Q000196:embryology; D013482:Superoxide Dismutase / Q000378:metabolism; D014475:Uncoupling Agents / Q000494:pharmacology; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid / Q000494:pharmacology
D019297:2,4-Dinitrophenol; D018750:6-Cyano-7-nitroquinoxaline-2,3-dione; D002118:Calcium; D054732:Calcium-Calmodulin-Dependent Protein Kinase Type 2; D017871:Calcium-Calmodulin-Dependent Protein Kinases; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D016572:Cyclosporine; D003992:Dibucaine; D016291:Dizocilpine Maleate; D004791:Enzyme Inhibitors; D018690:Excitatory Amino Acid Agonists; D018691:Excitatory Amino Acid Antagonists; D005452:Fluoresceins; D005456:Fluorescent Dyes; D018698:Glutamic Acid; D006575:Heterocyclic Compounds, 3-Ring; D007093:Imidazoles; D007608:Kainic Acid; D016202:N-Methylaspartate; D009419:Nerve Tissue Proteins; D009498:Neurotoxins; D019001:Nitric Oxide Synthase; D052248:Nitric Oxide Synthase Type I; D017382:Reactive Oxygen Species; D018091:Receptors, AMPA; D018092:Kainic Acid Receptors; D016194:Receptors, N-Methyl-D-Aspartate; D012235:Rhodamines; D013482:Superoxide Dismutase; D014475:Uncoupling Agents; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; C109098:2,7-dihydrodichlorofluorescein diacetate; C058319:dihydrorhodamine 123; C068483:rhod-2; C031938:calmidazolium; C496322:Nos1 protein, rat