In vivo activation and nuclear translocation of phosphorylated glycogen synthase kinase-3beta in neuronal apoptosis: links to tau phosphorylation.

The roles of glycogen synthase kinase-3beta (GSK-3beta) and tau phosphorylation were examined in seven-day-old rats injected with the NMDA receptor antagonist (MK801) that is known to induce neuronal apoptosis. Immunoblot and immunohistochemical analysis of brain samples demonstrated a site-specific increase in tau phosphorylation associated with the relocalization of the protein to the nuclear/perinuclear region of apoptotic neurons. In addition, a tau 32-kDa fragment was detected, suggesting that tau was a target of intracellular proteolysis in MK801-treated brains.

Cortical cell elution by sedimentation field-flow fractionation.

As a cell sorter, Sedimentation field-flow fractionation (SdFFF) can be defined as an effective tool for cell separation and purification, respecting integrity and viability as well as providing enhanced recovery and purified sterile fraction collection. The complex cell suspension containing both neurons and glial cells of all types, obtained from cerebral cortices of 17-day-old rat fetuses, is routinely used as a model of primary neuronal culture. Using SdFFF, this complex cell mixture was eluted in sterile fractions which were collected and cultured.

Markers of apoptosis and models of programmed cell death in Alzheimer's disease.

Alzheimer's disease (AD) is neuropathologically marked by the presence of senile plaques composed of beta-amyloid peptide and by neurofibrillary tangles formed by abnormally phosphorylated tau protein. Many authors have also reported a neuronal loss in affected regions of the brain in AD patients. This neuronal degeneration could be linked to the triggering of intracellular pathways leading to apoptosis.

N-methyl-D-aspartate receptor blockade enhances neuronal apoptosis induced by serum deprivation.

Neuronal apoptosis a hallmark of brain development could also be involved in neurodegenerative diseases. Glutamate toxicity is widely proposed as an important factor in the pathogenesis of neurological disorders. We show here that, in rat primary cortical cultures, the blockade of N-methyl-D-aspartate (NMDA) glutamate receptors exacerbated neuronal apoptosis induced by serum deprivation.

Toxic neuronal apoptosis and modifications of tau and APP gene and protein expressions.

The causes and the mechanisms of neuronal death in Alzheimer's disease are not elucidated, although some new insights have been proposed over the past years, including free-radical toxicity, beta-amyloid toxicity, excitotoxicity, and disturbed cellular calcium metabolism. Some authors have also pointed out that apoptosis could play a role in neuronal degeneration, but it is still largely debated. Here, we review some recent data linking the induction of experimental neuronal apoptosis in vitro and the molecular pathology of the tau protein and amyloid precursor protein (APP).

Damage and repair of nerve cell DNA in toxic stress.

It is generally agreed that ALS/PDC is triggered by a disappearing environmental factor peculiar to the lifestyle of people of the western Pacific (i.e., Guam, Irian Jaya, Indonesia, and the Kii Peninsula of Japan).

The Guam cycad toxin methylazoxymethanol damages neuronal DNA and modulates tau mRNA expression and excitotoxicity.

As in Alzheimer's disease, brains of Guam Chamorros with amyotrophic lateral sclerosis (ALS) and Parkinsonism-dementia complex (PDC) contain intraneuronal-paired helical filaments composed of accumulated phosphorylated tau protein. Tau mRNA expression in rat neuronal cultures-normally modulated by glutamate-increases after treatment with the aglycone of cycasin, a cycad-derived toxin whose concentration in Chamorro food varies with disease incidence. Elevated Tau gene expression in vitro is coincident with increased cycasin-related DNA adducts and reduced DNA repair.

Effects of depolarizing stimuli on calcium homeostasis in cultured rat motoneurones.

Intracellular calcium concentrations in individual rat motoneurones in enriched primary cultures were measured by Indo-1 fluorimetry. Motoneurones in the cultures were characterized morphometrically and by cholineacetyltransferase immunocytochemistry. Depolarization of the cells with glutamic acid or veratridine increased intracellular calcium levels, which returned to baseline only slowly after removal of the depolarizing agent.

Mild kainate toxicity produces selective motoneuron death with marked activation of CA(2+)-permeable AMPA/kainate receptors.

Motoneuron death could be produced by higher sensitivity to excitoxicity during the development and pathological conditions. We report here that in ventral spinal cord cultures mild kainate exposure (12.5 microM, 20 min or 100 microM, 2.

Characterization of human presenilin 1 transgenic rats: increased sensitivity to apoptosis in primary neuronal cultures.

Mutations in the gene for presenilin 1 are causative for the majority of cases of early onset familial Alzheimer's disease. Yet, the physiological function of presenilin 1 and the pathological mechanisms of the mutations leading to Alzheimer's disease are still unknown. To analyse potential pathological effects of presenilin 1 over-expression, we have generated transgenic rats which express high levels of human presenilin 1 protein in the brain.

Brefeldin A-induced apoptosis is expressed in rat neurons with dephosphorylated tau protein.

Brefeldin A (BFA) is a fungus metabolite (penicillum brefeldanum) that is known to produce the disintegration of the Golgi apparatus in exposed cells, and apoptosis in various cancer cells. This study reports that in rat primary cortical cell cultures BFA also produces apoptosis assessed by the TUNEL method and DAPI (4',6-diemidino-2-phenylindole) staining. The percentages of apoptotic neurons range from 26.

FK506 antagonizes apoptosis and c-jun protein expression in neuronal cultures.

FK506 is an immunosuppressive drug that binds to FK506 binding protein (FKBPs), a subgroup of cytosolic proteins called immunophillins. Previous works have revealed that FK506 protects neural cells from ischemia or excitotoxicity. Here we report that FK506 (10(-6) M) and not cyclosporine A (10(-6) M) blocks neuronal apoptosis induced by serum deprivation in rat neuronal cultures.

Neuronal apoptosis is associated with a decrease in tau mRNA expression.

Apoptosis is a programmed cell death that occurs during the development of the nervous system and in neurodegenerative disorders. Tau protein is a microtubule-associated protein which promotes microtubule polymerization and stabilization. Apoptosis was induced in primary neuronal cultures by glutamate exposure (200 microM, 15 min) or by serum deprivation, and tau mRNA levels were studied by quantitative in situ hybridization in apoptotic and non apoptotic neurons.

Glutamate toxicity enhances tau gene expression in neuronal cultures.

Tau protein is a microtubule-associated protein normally expressed in neurons. In Alzheimer's disease (AD) brains, phosphorylated tau accumulates in paired helical filaments which form neurofibrillary tangles in affected neurons; moreover, tau mRNA expression is increased in affected regions of AD brains. Glutamate, an excitatory neurotransmitter but also a potent neurotoxin under pathologic conditions, is known to produce neuronal degeneration and death accompanied by an increase in tau immunoreactivity in primary neuronal cultures.

Therapeutic advances in amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive and rapidly fatal neurodegenerative disease in which both upper and lower motoneurones are involved. The recent discovery of mutations affecting the superoxide dismutase (SOD) gene has given impetus to research on the role of oxidative stress in the pathogenesis of familial ALS, while further evidence for a role of excitotoxicity in the disease process has arisen. In this review, Erik Louvel, Jacques Hugon and Adam Doble discuss these findings and, in addition, describe how a number of large, well-controlled clinical trials have taken place to test potential therapies suggested by different aetiological hypotheses, including immunosuppressive therapies, neurotrophic factors, antioxidants and anti-excitotoxic drugs.

Cultured neurons expressing phosphorylated tau are more resistant to apoptosis induced by NMDA or serum deprivation.

Apoptosis is a programmed cell death that occurs during the development of the nervous system and in neurodegenerative disorders. Tau protein is a cytoskeletal component that promotes microtubule polymerization and stabilization. Apoptosis was induced in primary neuronal cultures by a prolonged exposure (16 h) to the NMDA (N-methyl-D-aspartate 20 microM) or by serum deprivation.

Increased hypoxic tolerance by chemical inhibition of oxidative phosphorylation: "chemical preconditioning".

A short ischemic episode preceding sustained ischemia is known to increase tolerance against ischemic cell death. We report early-onset long-lasting neuroprotection against in vitro hypoxia by preceding selective chemical inhibition of oxidative phosphorylation: "chemical preconditioning." The amplitude of CA1 population spikes (psap) in hippocampal slices prepared from control animals (control slices) was 31 +/- 27% (mean +/- SD) upon 45-min recovery from 15-min in vitro hypoxia.

NMDA induces apoptosis and necrosis in neuronal cultures. Increased APP immunoreactivity is linked to apoptotic cells.

In rat neuronal cultures exposed for a prolonged period (16 h) to a low concentration of N-methyl-D-aspartate (NMDA; 20 microM) two different types of neuronal death were observed, necrosis and apoptosis. The immunocytochemical study of necrotic neurons revealed the absence of amyloid precursor protein (APP) labeling in the cytoplasmic region. In contrast, neuronal apoptosis was associated with an increase in cytoplasmic APP immunoreactivity.

Identical chromosome imbalance in two siblings born to a mother with a double reciprocal translocation.

We report the case of a woman who carried two reciprocal translocations. Her karyotype was 46,XX,t(3;12)(q12;q21)(4;17)(p14;p13). She had two children, a phenotypically normal daughter (karyotype (46,XX,t(3;12)(q12;q21)) and a son with partial 4p trisomy (karyotype 46,XY,t(3;12) (q12;q21),-17,+maternal der(17)).

Neuronal APP accumulates in toxic membrane blebbings.

The occurence of plasma membrane blebbings is an early cytotoxic event, associated with the reorganization of cytoskeletal proteins, the alteration of interactions between the plasma membrane and the underlying cytoskeleton. The blebbing formation remains poorly understood but the involvement of cytosolic Ca2+ and the production of free radicals may contribute to this cellular phenomenom. The amyloid precursor protein (APP), is a transmembrane protein that can be cleaved to produce the beta amyloid peptide (Abeta) which accumulates in brain senile plaques of Alzheimer's disease.

ALS therapy: targets for the future.

There are four main hypotheses about the cause of ALS: excitotoxicity linked to glutamate receptor overactivation; mutation of the superoxide dismutase gene; production of autoantibodies to calcium channels; neurofilament accumulation. The motoneuron degeneration characteristic of ALS could be caused by any one or a combination of these mechanisms. Future therapeutic approaches should be based on these mechanisms and given in combination so that different levels of the degenerative process are targeted.

Antioxidant drugs block in vitro the neurotoxicity of CSF from patients with amyotrophic lateral sclerosis.

Amyotrophic lateral sclerosis (ALS) is a progressive neurological disease characterized by upper and lower motoneurone degeneration. Excitotoxicity and oxidative stress have been proposed as possible aetiological factors. We measured the neuronal death induced in rat cortical cell cultures by CSF taken from seven ALS patient and seven control subjects with lumbar radiculopathies.

Modifications of neuronal phosphorylated tau immunoreactivity induced by NMDA toxicity.

Glutamate toxicity has been involved in the pathophysiology of a large variety of neurodegenerative disorders. Tau Protein is a micro-tubule-associated protein that promotes microtubule polymerization and stabilization. Phosphorylated tau protein accumulates in paired helical neurofilaments, the major constituent of neurofibrillary tangles observed in the brain of patients suffering from Alzheimer disease (AD).

[Role of glutamate and excitotoxicity in neurologic diseases].

Glutamate is one of the major excitatory neurotransmitter in the central nervous system. Glutamate acts on 4 different post synaptic receptors; NMDA (N-Methyl-D-aspartate) AMPA (alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid), Kainate and metabotropic receptors. The three former receptors are linked to membrane ion channels whereas metabotropic receptors are coupled with a G protein.