Zinc-dependent multi-conductance channel activity in mitochondria isolated from ischemic brain.
Transient global ischemia is a neuronal insult that induces delayed cell death. A hallmark event in the early post-ischemic period is enhanced permeability of mitochondrial membranes. The precise mechanisms by which mitochondrial function is disrupted are, as yet, unclear. Here we show that global ischemia promotes alterations in mitochondrial membrane contact points, a rise in intramitochondrial Zn2+, and activation of large, multi-conductance channels in mitochondrial outer membranes by 1 h after insult. Mitochondrial channel activity was associated with enhanced protease activity and proteolytic cleavage of BCL-xL to generate its pro-death counterpart, deltaN-BCL-xL. The findings implicate deltaN-BCL-xL in large, multi-conductance channel activity. Consistent with this, large channel activity was mimicked by introduction of recombinant deltaN-BCL-xL to control mitochondria and blocked by introduction of a functional BCL-xL antibody to post-ischemic mitochondria via the patch pipette. Channel activity was also inhibited by nicotinamide adenine dinucleotide, indicative of a role for the voltage-dependent anion channel (VDAC) of the outer mitochondrial membrane. In vivo administration of the membrane-impermeant Zn2+ chelator CaEDTA before ischemia or in vitro application of the membrane-permeant Zn2+ chelator tetrakis-(2-pyridylmethyl) ethylenediamine attenuated channel activity, suggesting a requirement for Zn2+. These findings reveal a novel mechanism by which ischemic insults disrupt the functional integrity of the outer mitochondrial membrane and implicate deltaN-BCL-xL and VDAC in the large, Zn2+-dependent mitochondrial channels observed in post-ischemic hippocampal mitochondria.
Bonanni
Laura
L
Department of Oncology and Neuroscience and Center for Excellence on Aging, d'Annunzio Foundation, Università G. d'Annunzio-Chieti, 66013 Chieti, Italy.
Chachar
Mushtaque
M
Jover-Mengual
Teresa
T
Li
Hongmei
H
Jones
Adrienne
A
Yokota
Hidenori
H
Ofengeim
Dimitry
D
Flannery
Richard J
RJ
Miyawaki
Takahiro
T
Cho
Chang-Hoon
CH
Polster
Brian M
BM
Pypaert
Marc
M
Hardwick
J Marie
JM
Sensi
Stefano L
SL
Zukin
R Suzanne
RS
Jonas
Elizabeth A
EA
D000818:Animals; D015153:Blotting, Western / Q000379:methods; D002545:Brain Ischemia / Q000473:pathology*; D020169:Caspases / Q000378:metabolism; D002614:Chelating Agents / Q000494:pharmacology; D003952:Diagnostic Imaging / Q000379:methods; D004305:Dose-Response Relationship, Drug; D005029:Ethylenediamines / Q000494:pharmacology; D015640:Ion Channel Gating / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D007473:Ion Channels / Q000145:classification / Q000502:physiology*; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology / Q000528:radiation effects; D046529:Microscopy, Electron, Transmission / Q000379:methods; D008928:Mitochondria / Q000187:drug effects / Q000473:pathology / Q000502:physiology* / Q000648:ultrastructure; D009243:NAD / Q000494:pharmacology; D018408:Patch-Clamp Techniques / Q000379:methods; D051381:Rats; D017207:Rats, Sprague-Dawley; D013574:Synaptosomes / Q000187:drug effects / Q000502:physiology / Q000648:ultrastructure; D014966:Xanthenes; D015032:Zinc / Q000378:metabolism*
D002614:Chelating Agents; D005029:Ethylenediamines; D007473:Ion Channels; C484817:RhodZin-3; D014966:Xanthenes; D009243:NAD; D020169:Caspases; D015032:Zinc; C044387:N,N,N',N'-tetrakis(2-pyridylmethyl)ethylenediamine