Mitochondrial apoptotic cell death and moderate superoxide generation upon selective activation of non-desensitizing AMPA receptors in hippocampal cultures

Authors

  • A. Cristina Rego,

    1. Institute of Biochemistry, Faculty of Medicine and Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal
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  • Nuno M. Monteiro,

    1. Institute of Biochemistry, Faculty of Medicine and Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal
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  • Ana P. Silva,

    1. Institute of Biochemistry, Faculty of Medicine and Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal
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  • Joana Gil,

    1. Institute of Biochemistry, Faculty of Medicine and Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal
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  • João O. Malva,

    1. Institute of Biochemistry, Faculty of Medicine and Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal
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  • Catarina R. Oliveira

    1. Institute of Biochemistry, Faculty of Medicine and Center for Neuroscience and Cell Biology, University of Coimbra, Coimbra, Portugal
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Address correspondence and reprint requests to Ana Cristina Rego, PhD, Institute of Biochemistry, Faculty of Medicine, and Center for Neuroscience and Cell Biology, University of Coimbra, 3004–504 Coimbra, Portugal. E-mail: acrego@cnc.cj.uc.pt

Abstract

In the present work we investigated the effect of selective stimulation of non-desensitizing α-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA) receptors in the intracellular processes leading to hippocampal neuronal death and production of reactive oxygen species (ROS). Activation of AMPA receptors in the presence of cyclothiazide (CYZ), a blocker of AMPA receptor desensitization, resulted in the death of approximately 25% of neurones, which was prevented by 2,3-dihydroxy-6-nitro-7-sulphamoyl-benzo(f)quinoxaline (NBQX), an AMPA-preferring receptor antagonist. (+)-5-Methyl-10,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine hydrogen maleate (MK-801) protected the neurones from necrotic death induced by AMPA or NMDA receptor activation. Neurodegeneration caused by selective activation of non-desensitizing AMPA receptors, in the presence of AMPA, CYZ and MK-801, significantly decreased the number of Co2+-positive neurones, used as a cytochemical marker of Ca2+-permeable AMPA receptors, but maintained intracellular ATP/ADP. The AMPA-mediated apoptotic cell death involved mitochondrial cytochrome c release and the activation of caspases-1 and -3, which was prevented by NBQX. Interestingly, although selective activation of AMPA receptors was not associated with production of intracellular peroxides, a moderate increase in superoxide production was observed upon exposure to antimycin A (AA). Furthermore, increased activity of Mn- superoxide dismutase (SOD) was observed on selective activation of non-desensitizing AMPA receptors. Taken together, these data make important contributions to the elucidation of the downstream pathways activated in AMPA receptor-mediated excitotoxicity in cultured rat hippocampal neurones.

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