Mechanisms underlying the loss of mitochondrial membrane potential in glutamate excitotoxicity.
Glutamate excitotoxicity amplifies neuronal death following stroke. We have explored the mechanisms underlying the collapse of mitochondrial potential (Deltapsi(m)) and loss of [Ca(2+)](c) homeostasis in rat hippocampal neurons in culture following toxic glutamate exposure. The collapse of Deltapsi(m) is multiphasic and Ca(2+)-dependent. Glutamate induced a decrease in NADH autofluorescence which preceded the loss of Deltapsi(m). Both the decrease in NADH signal and the loss of Deltapsi(m) were suppressed by Ru360 and both were delayed by inhibition of PARP (by 3-AB or DPQ). During this period, addition of mitochondrial substrates (methyl succinate and TMPD-ascorbate) or buffering [Ca(2+)](i) (using BAPTA-AM or EGTA-AM), rescued Deltapsi(m). These data suggest that mitochondrial Ca(2+) uptake activates PARP which in turn depletes NADH, promoting the initial collapse of Deltapsi(m). After > approximately 20 min, buffering Ca(2+) or substrate addition failed to restore Deltapsi(m). In neurons from cyclophilin D-/- (cypD-/-) mice or in cells treated with cyclosporine A, removal of Ca(2+) restored Deltapsi(m) even after 20 min of glutamate exposure, suggesting involvement of the mPTP in the irreversible depolarisation seen in WT cells. Thus, mitochondrial depolarisation represents two consecutive but distinct processes driving cell death, the first of which is reversible while the second is not.
Abramov
Andrey Y
AY
Department of Physiology, University College London, Gower Street, London WC1E 6BT, UK. a.abramov@ucl.ac.uk
Duchen
Michael R
MR
D000818:Animals; D002545:Brain Ischemia / Q000503:physiopathology; D002118:Calcium / Q000378:metabolism / Q000502:physiology; D018698:Glutamic Acid / Q000633:toxicity*; D006624:Hippocampus / Q000502:physiology; D000860:Hypoxia / Q000503:physiopathology; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology*; D051336:Mitochondrial Membranes / Q000187:drug effects / Q000502:physiology*; D009243:NAD / Q000378:metabolism; D009474:Neurons / Q000502:physiology; D009498:Neurotoxins / Q000633:toxicity*; D051381:Rats; D017207:Rats, Sprague-Dawley
D009498:Neurotoxins; D009243:NAD; D018698:Glutamic Acid; D002118:Calcium