Hierarchical recruitment by AMPA but not staurosporine of pro-apoptotic mitochondrial signaling in cultured cortical neurons: evidence for caspase-dependent/independent cross-talk

• Published in: Journal of neurochemistry 2007-12, Vol.103 (6), p.2408-2427
• Main Authors: Beart, Philip M, Lim, Maria L.R, Chen, Baohong, Diwakarla, Shanti, Mercer, Linda D, Cheung, Nam Sang, Nagley, Phillip
• Format: Article
• Language: English
• Subjects: (S)-α-amino-3-hydroxy-5-methylisoxazole-4-propionate; Ageing, cell death; alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid - pharmacology; Animals; Apoptosis; Apoptosis - drug effects; Apoptosis - physiology; Apoptosis Regulatory Proteins - drug effects; Apoptosis Regulatory Proteins - metabolism; Bax; Biochemistry; Biological and medical sciences; Brain damage; calpain; Calpain - metabolism; caspase 3; Caspases - drug effects; Caspases - metabolism; Cell culture; Cell physiology; Cells, Cultured; Cerebral Cortex - drug effects; Cerebral Cortex - metabolism; Cytochromes c - metabolism; Enzyme Activation - drug effects; Enzyme Activation - physiology; Enzyme Inhibitors - pharmacology; Excitatory Amino Acid Agonists - pharmacology; Fundamental and applied biological sciences. Psychology; Isolated neuron and nerve. Neuroglia; Membrane Potential, Mitochondrial - drug effects; Membrane Potential, Mitochondrial - physiology; Mice; Mitochondria - drug effects; Mitochondria - metabolism; mitochondrial membrane potential; mitochondrial pro-apoptotic proteins; Molecular and cellular biology; Neurology; Neurons - drug effects; Neurons - metabolism; Neurotoxins - pharmacology; Receptor Cross-Talk - drug effects; Receptor Cross-Talk - physiology; Receptors, AMPA - drug effects; Receptors, AMPA - metabolism; Signal Transduction - drug effects; Signal Transduction - physiology; Staurosporine - pharmacology; Vertebrates: nervous system and sense organs
• ISSN: 0022-3042; 1471-4159
• DOI: 10.1111/j.1471-4159.2007.04937.x

Excitotoxicity mediated via the (S)-α-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) subtype of receptor for l-glutamate contributes to various neuropathologies involving acute brain injury and chronic degenerative disorders. In this study, AMPA-induced neuronal injury and staurosporine (STS)-mediated apoptosis were compared in primary neuronal cultures of murine cerebral cortex by analyzing indices up- and downstream of mitochondrial activation. AMPA-mediated apoptosis involved induction of Bax, loss of mitochondrial transmembrane potential (ΔΨm), early release of cytochrome c (cyt c), and more delayed release of second mitochondrial activator of caspases (SMAC), Omi, and apoptosis-inducing factor (AIF) with early calpain and minor late activation of caspase 3. STS-induced apoptosis was characterized by a number of differences, a more rapid time course, non-involvement of ΔΨm, and relatively early recruitment of SMAC and caspase 3. The AMPA-induced rise in intracellular calcium appeared insufficient to evoke ΔΨm as release of cyt c preceded mitochondrial depolarization, which was followed by the cytosolic translocation of SMAC, Omi, and AIF. Bax translocation preceded cyt c release for both stimuli inferring its involvement in apoptotic induction. Inclusion of the broad spectrum caspase inhibitor zVAD-fmk reduced the AMPA-induced release of cyt c, SMAC, and AIF, while only affecting the redistribution of Omi and AIF in the STS-treated neurons. Only AIF release was affected by a calpain inhibitor (calpastatin) which exerted relatively minor effects on the progression of cellular injury. AMPA-mediated release of apoptogenic proteins was more hierarchical relative to STS with its calpain activation and caspase-dependent AIF redistribution arguing for a model with cross-talk between caspase-dependent/independent apoptosis.