dizocilpine-maleate and calmidazolium

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.

Topics: 2,4-Dinitrophenol; 6-Cyano-7-nitroquinoxaline-2,3-dione; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; Animals; Calcium; Calcium Signaling; Calcium-Calmodulin-Dependent Protein Kinase Type 2; Calcium-Calmodulin-Dependent Protein Kinases; Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; Cells, Cultured; Cyclosporine; Dibucaine; Dizocilpine Maleate; Enzyme Inhibitors; Excitatory Amino Acid Agonists; Excitatory Amino Acid Antagonists; Fluoresceins; Fluorescent Dyes; Glutamic Acid; Heterocyclic Compounds, 3-Ring; Imidazoles; Kainic Acid; Mitochondria; Motor Neuron Disease; Motor Neurons; N-Methylaspartate; Nerve Tissue Proteins; Neurons; Neurotoxins; Nitric Oxide Synthase; Nitric Oxide Synthase Type I; Oxidative Stress; Rats; Rats, Wistar; Reactive Oxygen Species; Receptors, AMPA; Receptors, Kainic Acid; Receptors, N-Methyl-D-Aspartate; Rhodamines; Spinal Cord; Superoxide Dismutase; Uncoupling Agents

Topics: Adenosine Triphosphate; Animals; Calmodulin; Dizocilpine Maleate; Energy Metabolism; Hippocampus; Imidazoles; In Vitro Techniques; Ischemic Attack, Transient; Ketamine; Kinetics; Rats

Excessive Ca2+ influx through NMDA receptor-coupled channels has been linked to neuronal cell death. Using an in vitro model of transient brain ischemia, we investigated possible protective effects of NMDA receptor antagonists ketamine or MK-801 and of calmidazolium, an inhibitor of intracellular Ca2(+)-activated proteins. Brain ischemia/recovery was simulated in isolated hippocampal slices and injury monitored by measurement of ATP levels. Omission of both glucose and oxygen (but not oxygen alone) for 20 min led to persistent ATP deficits after 4 h recovery. Addition of ketamine or MK-801 at 1 microM permitted ATP to recover within 1 h, as did addition of calmidazolium at 10 microM. Our findings are consistent with other reports that NMDA receptor antagonists can protect neuronal tissue from ischemic damage. The role of inappropriately activated Ca2(+)-mediated signaling processes in the mechanism(s) of such injury is suggested by the protection also seen with calmidazolium, an inhibitor of calmodulin and other structurally related proteins such as calpain(s) and protein kinase C. The inhibition of intracellular Ca2+ target proteins may be an alternative for protection of the brain against injury due to insults that activate NMDA receptors.

Topics: Adenosine Triphosphate; Animals; Brain Ischemia; Dizocilpine Maleate; Hippocampus; Imidazoles; In Vitro Techniques; Ketamine; Rats; Receptors, N-Methyl-D-Aspartate; Time Factors