1-Methyl-4-Phenylpyridinium Induces Autocrine Excitotoxicity, Protease Activation, and Neuronal Apoptosis
- Faculty of Biology, Chair of Molecular Toxicology, University of Konstanz, D-78457 Konstanz, Germany
Abstract
The pathogenesis of several neurodegenerative diseases may involve indirect excitotoxic mechanisms, where glutamate receptor overstimulation is a secondary consequence of initial functional defects of neurons (e.g., impairment of mitochondrial energy generation). The neurotoxin 1-methyl-4-phenylpyridinium (MPP+) and other mitochondrial inhibitors (e.g., rotenone or 3-nitropropionic acid) elicited apoptosis in cerebellar granule cell cultures via stimulation of autocrine excitotoxicity. Cell death, increase in intracellular Ca2+ concentration, release of cytochrome c, and all biochemical and morphological signs of apoptosis were prevented by blockade of theN-methyl-d-aspartate receptor with noncompetitive, glycine-site or glutamate-site inhibitors. In addition, MPP+-induced apoptosis was reduced by high Mg2+concentrations in the medium or by inhibiting exocytosis with clostridial neurotoxins. Two classes of cysteine proteases were involved in the execution of cell death: caspases and calpains. Inhibitors of either class of proteases prevented cell death, cleavage of intracellular proteins (i.e., fodrin), and the appearance of typical features of apoptosis such as phosphatidylserine translocation or DNA fragmentation. However, protease inhibitors did not interfere with the initial intracellular Ca2+ concentration increase. We suggest that MPP+ as well as other mitochondrial inhibitors trigger indirect excitotoxic processes, which lead to Ca2+overload, protease activation, and subsequent neuronal apoptosis.
Footnotes
-
Send reprint requests to: Dr P. Nicotera, University of Konstanz, Faculty of Biology, Department of Molecular Toxicology, Box X911, D-78457 Konstanz, Germany. E-mail:pierluigi.nicotera{at}uni-konstanz.de
-
This study was supported by the DFG Grants Ni519/1–1 and Ni519/2–1 and the EEC Grants ENV4-CT96–0169, BMH4CT97-2410, and 12029–97-06 F1ED ISP D.
- Abbreviations:
- [Ca2+]i
- intracellular Ca2+ concentration
- ΔΨ
- mitochondrial membrane potential
- AP5
- 5-aminophosphovalerat
- calp II
- acetyl-leucyl-leucyl-l-methional
- calp III
- z-Val-l-phenylalaninal
- CCCP
- carbonylcyanide-chlorophenylhydrazone
- CGC
- cerebellar granule cells
- CSS
- controlled salt solution
- DCK
- 5,7-dichlorokynurenate
- DIV
- daysin vitro
- EGTA
- ethylene glycol bis(β-aminoethyl ether)-N,N,N′,N′-tetraacetic acid
- HEPES
- 4-(2-hydroxyethyl)-1-piperazineethanesulfonic acid
- DNQX
- dinitroquinoxalinedione
- MK801
- (+)-5-methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine
- MPP+
- 1-methyl-4-phenylpyridinium
- MTT
- 3-(4,5-dimethylthiazole-2-yl)-2,5-diphenyltetrasodium bromide
- NMDA
- N-methyl-d-aspartate
- NMDA-R
- N-methyl-d-aspartate receptor
- NO
- nitric oxide
- 3-NP
- 3-nitropropionic acid
- PARP
- poly-(ADP-ribose)polymerase (E.C. 2.4.2.30)
- BoNT/C
- botulinum neurotoxin serotype C
- PS
- phosphatidylserine
- TMRE
- tetramethylrhodamine ethylester
- VDCC
- voltage-dependent calcium channel
- z-D-cbk
- z-aspartyl-2,6-dichlorobenzoyloxymethylketone
-
- Received April 1, 1998.
- Accepted August 11, 1998.
- The American Society for Pharmacology and Experimental Therapeutics
