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Caspases Mediate 6‐Hydroxydopamine‐Induced Apoptosis but Not Necrosis in PC12 Cells
Author(s) -
Ochu Erinma E.,
Rothwell Nancy J.,
Waters Catherine M.
Publication year - 1998
Publication title -
journal of neurochemistry
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 1.75
H-Index - 229
eISSN - 1471-4159
pISSN - 0022-3042
DOI - 10.1046/j.1471-4159.1998.70062637.x
Subject(s) - apoptosis , caspase , programmed cell death , hydroxydopamine , intrinsic apoptosis , microbiology and biotechnology , viability assay , neurotoxin , caspase 3 , necrosis , biology , xiap , proteases , chemistry , biochemistry , endocrinology , enzyme , genetics , dopaminergic , dopamine
The neurotoxin 6‐hydroxydopamine (6‐OHDA) induces apoptosis in the rat phaeochromocytoma cell line PC12. 6‐OHDA‐induced apoptosis is morphologically indistinguishable from serum deprivation‐induced apoptosis. Exposure of PC12 cells to a low concentration of 6‐OHDA (25 µ M ) results in apoptosis, whereas an increased concentration (50 µ M ) results in a mixture of apoptosis and necrosis. We investigated the involvement of caspases in the apoptotic death of PC12 cells induced by 6‐OHDA, using a general caspase inhibitor, benzyloxycarbonyl‐Val‐Ala‐Asp‐fluoromethyl ketone (zVAD‐fmk), and compared this with serum deprivation‐induced apoptosis, which is known to involve caspases. We show that zVAD‐fmk (100 µ M ) completely prevented the apoptotic morphology of chromatin condensation induced by exposure to either 6‐OHDA (25 and 50 µ M ) or serum deprivation. Furthermore, cell lysates from 6‐OHDA‐treated cultures showed cleavage of a fluorogenic substrate for caspase‐3‐like proteases (caspase‐2, 3, and 7), acetyl‐Asp‐Glu‐Val‐Asp‐aminomethylcoumarin, and this was inhibited by zVAD‐fmk. However, although zVAD‐fmk restored total cell viability to serum‐deprived cells or cells exposed to 25 µ M 6‐OHDA, the inhibitor did not restore viability to cells exposed to 50 µ M 6‐OHDA. These data show the involvement of a caspase‐3‐like protease in 6‐OHDA‐induced apoptosis and that caspase inhibition is sufficient to rescue PC12 cells from the apoptotic but not the necrotic component of 6‐OHDA neurotoxicity.