Beta-carbolines induce apoptosis in cultured cerebellar granule neurons via the mitochondrial pathway.
N-butyl-beta-carboline-3-carboxylate (betaCCB) is, together with 2-methyl-norharmanium and 2,9-dimethylnorharmanium ions, an endogenously occurring beta-carboline. Due to their structural similarities with the synthetic neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), harman and norharman compounds have been proposed to be involved in the pathogenesis of Parkinson's disease. While also structurally related, betaCCB has received much less interest in that respect although we had previously demonstrated that it induces the apoptotic cell death of cultured cerebellar granule neurons (CGNs). Herein, we have investigated the molecular events leading to CGN apoptosis upon betaCCB treatment. We first demonstrated that betaCCB-induced apoptosis occurs in neurons only, most likely as a consequence of a specific neuronal uptake as shown using binding/uptake experiments. Then we observed that, in betaCCB-treated CGNs, caspases 9, 3 and 8 were successively activated, suggesting an activation of the mitochondrial pathway. Consistently, betaCCB also induced the release from the mitochondrial intermembrane space of two pro-apoptotic factors, i.e. cytochrome c and apotptosis inducing factor (AIF). Interestingly, no mitochondrial membrane depolarisation was associated with this release, suggesting a mitochondrial permeability transition pore-independent mechanism. The absence of any neuroprotective effect provided by two mPTP inhibitors, i.e. cyclosporine A and bongkrekic acid, further supported this hypothesis. Together, these results show that betaCCB is specifically taken up by neuronal cells where it triggers a specific permeabilization of the outer mitochondrial membrane and a subsequent apoptotic cell death.
Hans
Grégory
G
Centre of Cellular and Molecular Neurobiology, Université de Liège, 17 Place Delcour, 4020 Liège 2, Belgium. g.hans@ulg.ac.be
Malgrange
Brigitte
B
Lallemend
François
F
Crommen
Jessica
J
Wislet-Gendebien
Sabine
S
Belachew
Shibeshih
S
Robe
Pierre
P
Rogister
Bernard
B
Moonen
Gustave
G
Rigo
Jean-Michel
JM
D048868:Adaptor Proteins, Signal Transducing / Q000378:metabolism; D000704:Analysis of Variance; D000818:Animals; D000831:Animals, Newborn; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D051033:Apoptosis Inducing Factor; D001253:Astrocytes / Q000187:drug effects; D001562:Benzimidazoles / Q000378:metabolism; D001665:Binding Sites / Q000187:drug effects; D015153:Blotting, Western / Q000379:methods; D002232:Carbocyanines / Q000378:metabolism; D002243:Carbolines / Q000378:metabolism / Q000494:pharmacology*; D061945:Caspase Inhibitors; D020169:Caspases / Q000378:metabolism; D002452:Cell Count / Q000379:methods; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002531:Cerebellum / Q000166:cytology*; D045304:Cytochromes c / Q000494:pharmacology; D004347:Drug Interactions; D004996:Ethidium; D005420:Flavoproteins / Q000378:metabolism; D005434:Flow Cytometry / Q000379:methods; D005455:Fluorescent Antibody Technique / Q000379:methods; D005562:Formazans; D008565:Membrane Proteins / Q000378:metabolism; D008928:Mitochondria / Q000187:drug effects*; D009474:Neurons / Q000166:cytology / Q000187:drug effects*; D051381:Rats; D013778:Tetrazolium Salts; D013997:Time Factors; D014316:Tritium / Q000378:metabolism
D048868:Adaptor Proteins, Signal Transducing; C494831:Aifm1 protein, rat; D051033:Apoptosis Inducing Factor; D001562:Benzimidazoles; D002232:Carbocyanines; D002243:Carbolines; D061945:Caspase Inhibitors; D005420:Flavoproteins; D005562:Formazans; D008565:Membrane Proteins; D013778:Tetrazolium Salts; D014316:Tritium; C068624:5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine; C079782:MTT formazan; C049504:N-butyl beta-carboline-3-carboxylate; D045304:Cytochromes c; D020169:Caspases; D004996:Ethidium