Stimulation of glutamate receptors in cultured hippocampal neurons causes Ca2+-dependent mitochondrial contraction.
Cultured hippocampal neurons expressing mitochondrially-targeted enhanced yellow fluorescent protein (mito-eYFP) were used to quantitatively examine mitochondrial remodelling in response to excitotoxic glutamate. Mitochondrial morphology was evaluated using laser spinning-disk confocal microscopy followed by calibrated image processing and 3D image rendering. Glutamate triggered an increase in cytosolic Ca(2+) and mitochondrial depolarization accompanied by Ca(2+)-dependent morphological transformation of neuronal mitochondria from "thread-like" to rounded structures. The quantitative analysis of the mitochondrial remodelling revealed that exposure to glutamate resulted in a decrease in mitochondrial volume and surface area concurrent with an increase in sphericity of the organelles. NIM811, an inhibitor of the mitochondrial permeability transition, attenuated the glutamate-induced sustained increase in cytosolic Ca(2+) and suppressed mitochondrial remodelling in the majority of affected neurons, but it did not rescue mitochondrial membrane potential. Shortening, fragmentation, and formation of circular mitochondria with decreased volume and surface area accompanied mitochondrial depolarization with FCCP. However, FCCP-induced morphological alterations appeared to be distinctly different from mitochondrial remodelling caused by glutamate. Moreover, FCCP prevented glutamate-induced mitochondrial remodelling suggesting an important role of Ca(2+) influx into mitochondria in the morphological alterations. Consistent with this, in saponin-permeabilized neurons, Ca(2+) caused mitochondrial remodelling which could be prevented by Ru(360).
Brustovetsky
Tatiana
T
Department of Pharmacology and Toxicology, Indiana University School of Medicine, Indianapolis, IN 46202, USA. nbrous@iupui.edu
Li
Viacheslav
V
Brustovetsky
Nickolay
N
D000818:Animals; D002118:Calcium / Q000037:antagonists & inhibitors / Q000378:metabolism*; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone / Q000494:pharmacology; D002478:Cells, Cultured; D018698:Glutamic Acid / Q000494:pharmacology*; D049452:Green Fluorescent Proteins / Q000235:genetics / Q000378:metabolism; D006624:Hippocampus / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D008027:Light; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects; D008928:Mitochondria / Q000187:drug effects / Q000378:metabolism / Q000648:ultrastructure*; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism; D000083162:Mitochondrial Permeability Transition Pore; D008933:Mitochondrial Swelling / Q000187:drug effects; D009474:Neurons / Q000187:drug effects / Q000378:metabolism* / Q000648:ultrastructure; D010539:Permeability; D051381:Rats; D017470:Receptors, Glutamate / Q000378:metabolism; D017975:Ruthenium Compounds / Q000494:pharmacology; D014634:Valinomycin / Q000494:pharmacology
D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D017470:Receptors, Glutamate; C112020:Ru 360; D017975:Ruthenium Compounds; D049452:Green Fluorescent Proteins; D014634:Valinomycin; D002259:Carbonyl Cyanide p-Trifluoromethoxyphenylhydrazone; D018698:Glutamic Acid; D002118:Calcium