Alterations of intracellular calcium homeostasis and mitochondrial function are involved in ruthenium red neurotoxicity in primary cortical cultures.
Ruthenium red (RR) is a polycationic dye that induces neuronal death in vivo and in primary cultures. To characterize this neurotoxic action and to determine the mechanisms involved, we have analyzed the ultrastructural alterations induced by RR in rat cortical neuronal cultures and measured its effect on cytoplasmic Ca(2+) concentration ([Ca(2+)](i)) and on mitochondrial function. RR produced a dose-dependent, progressive disruption of neurites and plasma membrane of neuronal somata after 8-24 hr of incubation. RR caused also an elevation of both the basal [Ca(2+)](i) and its maximal levels after K(+) depolarization. Mitochondrial oxidative function, assessed by reduction of 3-(4,5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide and by changes in dihydrorhodamine-123 fluorescence, was significantly diminished after treatment with RR, both in cultured neurons and in isolated brain mitochondria. La(3+) did not prevent but rather potentiated RR-induced cell death. Glutamate receptor antagonists also failed to prevent RR neurotoxicity. Apoptotic electron microscope images were not observed, and protein synthesis inhibitors did not show any protective effect. It is concluded that RR penetrates neurons and that its neurotoxic damage probably is due to intracellular Ca(2+) dishomeostasis and disruption of mitochondrial oxidative function. These results enhance our understanding of the intracellular mechanisms underlying neuronal death.
Velasco
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I
Departamento de Neurociencias, Instituto de Fisiología Celular, Universidad Nacional Autónoma de México.
Tapia
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D000818:Animals; D017209:Apoptosis; D001667:Binding, Competitive / Q000187:drug effects; D002118:Calcium / Q000378:metabolism*; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000378:metabolism; D002462:Cell Membrane / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism*; D004305:Dose-Response Relationship, Drug; D018691:Excitatory Amino Acid Antagonists / Q000494:pharmacology; D006706:Homeostasis / Q000187:drug effects; D007424:Intracellular Fluid / Q000378:metabolism*; D008855:Microscopy, Electron, Scanning; D008928:Mitochondria / Q000378:metabolism*; D016501:Neurites / Q000187:drug effects / Q000378:metabolism; D009474:Neurons / Q000166:cytology / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure; D010084:Oxidation-Reduction / Q000187:drug effects; D051381:Rats; D012235:Rhodamines / Q000378:metabolism / Q000493:pharmacokinetics; D012430:Ruthenium Red / Q000633:toxicity*
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D018691:Excitatory Amino Acid Antagonists; D012235:Rhodamines; C058319:dihydrorhodamine 123; D012430:Ruthenium Red; D002118:Calcium