calpain and ifenprodil

The altered function and/or structure of tau protein is postulated to cause cell death in tauopathies and Alzheimer's disease. However, the mechanisms by which tau induces neuronal death remain unclear. Here we show that overexpression of human tau and of some of its N-terminal fragments in primary neuronal cultures leads to an N-methyl-D-aspartate receptor (NMDAR)-mediated and caspase-independent cell death. Death signaling likely originates from stimulation of extrasynaptic NR2B-subunit-containing NMDARs because it is accompanied by dephosphorylation of cAMP-response-element-binding protein (CREB) and it is inhibited by ifenprodil. Interestingly, activation of NMDAR leads to a crucial, sustained, and delayed phosphorylation of extracellular-regulated kinases 1 and 2, whose inhibition largely prevents tau-induced neuronal death. Moreover, NMDAR involvement causes the fatal activation of calpain, which, in turn, degrades tau protein into a 17-kDa peptide and possibly other highly toxic N-terminal peptides. Some of these peptides are hypothesized, on the basis of our in vitro experiments, to initiate a negative loop, ultimately leading to cell death. Thus, inhibition of calpain largely prevents tau degradation and cell death. Our findings unravel a cellular mechanism linking tau toxicity to NMDAR activation and might be relevant to Alzheimer's disease and tauopathies where NMDAR-mediated toxicity is postulated to play a pivotal role.

Topics: Alzheimer Disease; Animals; Calpain; Caspase Inhibitors; Caspases; Cells, Cultured; CREB-Binding Protein; Cysteine Proteinase Inhibitors; Enzyme Activation; Humans; Mice; Mice, Mutant Strains; Mitogen-Activated Protein Kinase 1; Mitogen-Activated Protein Kinase 3; Neurons; Phosphorylation; Piperidines; Protein Kinase Inhibitors; Rats; Rats, Wistar; Receptors, N-Methyl-D-Aspartate; tau Proteins; Transcriptional Activation; Transfection

Excitotoxicity is generally studied in dissociated neurons, cultured hippocampal slices, or intact animals. However, the requirements of dissociated neurons or cultured slices to use prenatal or juvenile rats seriously limit the advantages of these systems, whereas the complexity of intact animals prevents detailed molecular investigations. In the present experiments, we studied developmental changes in NMDA neurotoxicity in acute hippocampal slices with lactate dehydrogenase (LDH) release in medium, propidium iodide (PI) uptake, and Nissl staining as markers of cell damage. Calpain-mediated spectrin degradation was used to test calpain involvement in NMDA neurotoxicity. NMDA treatment produced increased LDH release, PI uptake, and spectrin degradation in slices from juvenile rats but not adult rats. NMDA-induced changes in slices from young rats were blocked completely by the NMDA receptor antagonist (+)-5-methyl-10,11-dihydro-5H-dibenzo [a,d] cyclohepten-5,10-imine maleate (MK-801) and by the antagonists of NR2B receptor ifenprodil and R-(R, S)-alpha-(4-hydroxyphenyl)-beta-methyl-4-(phenylmethyl)-1-piperidine propranol and were partly blocked by calpain inhibitor III but were not affected by the NR2A-specific antagonist [(R)-[(S)-1-(4-bromo-phenyl)-ethylamino]-(2,3-dioxo-1,2,3,4-tetrahydroquinoxalin-5-yl)-methyl]-phosphonic acid. NMDA-induced changes in Nissl staining were also different in slices from young and adult rats and blocked by NR2B but not NR2A antagonists. In contrast to NMDA treatment, oxygen/glucose deprivation (OGD) induced neurotoxicity in slices from both young and adult rats, although OGD-induced toxicity was attenuated by MK-801 only in slices from young rats. Our results are consistent with the idea that NMDA-mediated toxicity is caused by activation of NR2B- but not NR2A-containing NMDA receptors leading to calpain activation and that developmental changes in NMDA toxicity reflect developmental changes in NMDA receptor subunit composition.

Topics: Age Factors; Animals; Biomarkers; Calpain; Dipeptides; Disks Large Homolog 4 Protein; Dizocilpine Maleate; Enzyme Activation; Excitatory Amino Acid Antagonists; Fluorescent Dyes; Gene Expression Regulation, Developmental; Glucose; Hippocampus; In Vitro Techniques; Intracellular Signaling Peptides and Proteins; L-Lactate Dehydrogenase; Membrane Proteins; N-Methylaspartate; Neurons; Oxygen; Phenols; Piperidines; Propidium; Protein Subunits; Quinoxalines; Rats; Rats, Sprague-Dawley; Receptors, N-Methyl-D-Aspartate; Spectrin; Valine