Showing papers by "Mark P. Mattson published in 2000"

Apoptosis in neurodegenerative disorders

Abstract: Neuronal death underlies the symptoms of many human neurological disorders, including Alzheimer's, Parkinson's and Huntington's diseases, stroke, and amyotrophic lateral sclerosis. The identification of specific genetic and environmental factors responsible for these diseases has bolstered evidence for a shared pathway of neuronal death--apoptosis--involving oxidative stress, perturbed calcium homeostasis, mitochondrial dysfunction and activation of cysteine proteases called caspases. These death cascades are counteracted by survival signals, which suppress oxyradicals and stabilize calcium homeostasis and mitochondrial function. With the identification of mechanisms that either promote or prevent neuronal apoptosis come new approaches for preventing and treating neurodegenerative disorders.

Homocysteine Elicits a DNA Damage Response in Neurons That Promotes Apoptosis and Hypersensitivity to Excitotoxicity

Abstract: Elevated plasma levels of the sulfur-containing amino acid homocysteine increase the risk for atherosclerosis, stroke, and possibly Alzheimer's disease, but the underlying mechanisms are unknown. We now report that homocysteine induces apoptosis in rat hippocampal neurons. DNA strand breaks and associated activation of poly-ADP-ribose polymerase (PARP) and NAD depletion occur rapidly after exposure to homocysteine and precede mitochondrial dysfunction, oxidative stress, and caspase activation. The PARP inhibitor 3-aminobenzamide (3AB) protects neurons against homocysteine-induced NAD depletion, loss of mitochondrial transmembrane potential, and cell death, demonstrating a requirement for PARP activation and/or NAD depletion in homocysteine-induced apoptosis. Caspase inhibition accelerates the loss of mitochondrial potential and shifts the mode of cell death to necrosis; inhibition of PARP with 3AB attenuates this effect of caspase inhibition. Homocysteine markedly increases the vulnerability of hippocampal neurons to excitotoxic and oxidative injury in cell culture and in vivo, suggesting a mechanism by which homocysteine may contribute to the pathogenesis of neurodegenerative disorders.

Evidence for the involvement of TNF and NF-κB in hippocampal synaptic plasticity

Abstract: The cytokine tumor necrosis factor-alpha (TNF), well-known for its roles in cellular responses to tissue injury, has recently been shown to be produced in response to physiological activity in neuronal circuits. TNF stimulates receptors in neurons linked to the activation of the transcription factor NF-kappaB, and recent findings suggest that this signaling pathway can modulate neuronal excitability and vulnerability of neurons to excitotoxicity. Because data indicate that TNF is produced, and NF-kappaB activated, under conditions associated with learning and memory, we performed experiments in the hippocampal slice preparation aimed at elucidating roles for TNF and NF-kappaB in modulating synaptic plasticity. Whereas stimulation of Schaffer collateral axons at a frequency of 1 Hz induced long-term depression (LTD) of synaptic transmission in region CA1 of wild-type mice, LTD did not occur in slices from TNF receptor knockout mice. Stimulation at 100 Hz induced long-term potentiation (LTP) in slices from both wild-type mice and mice lacking TNF receptors. Basal transmission was unaltered in mice lacking TNF receptors. Pretreatment of slices from wild-type mice with kappaB decoy DNA prevented induction of LTD and significantly reduced the magnitude of LTP. Collectively, these data suggest important roles for TNF and signaling pathways that modulate NF-kappaB activity in regulation of hippocampal synaptic plasticity.

Dietary restriction increases the number of newly generated neural cells, and induces BDNF expression, in the dentate gyrus of rats.

Abstract: The adult brain contains neural stem cells that are capable of proliferating, differentiating into neurons or glia, and then either surviving or dying. This process of neural-cell production (neurogenesis) in the dentate gyrus of the hippocampus is responsive to brain injury, and both mental and physical activity. We now report that neurogenesis in the dentate gyrus can also be modified by diet. Previous studies have shown that dietary restriction (DR) can suppress age-related deficits in learning and memory, and can increase resistance of neurons to degeneration in experimental models of neurodegenerative disorders. We found that maintenance of adult rats on a DR regimen results in a significant increase in the numbers of newly produced neural cells in the dentate gyrus of the hippocampus, as determined by stereologic analysis of cells labeled with the DNA precursor analog bromodeoxyuridine. The increase in neurogenesis in rats maintained on DR appears to result from decreased death of newly produced cells, rather than from increased cell proliferation. We further show that the expression of brain-derived neurotrophic factor, a trophic factor recently associated with neurogenesis, is increased in hippocampal cells of rats maintained on DR. Our data are the first evidence that diet can affect the process of neurogenesis, as well as the first evidence that diet can affect neurotrophic factor production. These findings provide insight into the mechanisms whereby diet impacts on brain plasticity, aging and neurodegenerative disorders.

Antiinflammatory effects of estrogen on microglial activation.

Abstract: In the present study the effects of 17beta-estradiol on microglial activation are described. Estrogen replacement therapy has been associated with decreased severity of age-related neurodegenerative diseases such as Alzheimer's disease, and estrogens have potent immunosuppressive properties outside of the brain. To determine the role that microglial cells might play in estrogen-mediated neuroprotection, primary rat microglia and N9 microglial cell lines were treated with increasing doses of 17beta-estradiol before or during immunostimulation by lipopolysaccharide, phorbol ester, or interferon-gamma. Pretreatment with 17beta-estradiol, but not 17alpha-estradiol or progesterone, dose dependently attenuated microglial superoxide release and phagocytic activity. Additionally, 17beta-estradiol attenuated increases in inducible nitric oxide synthase protein expression, but did not alter nuclear factor-KB activation. The antiinflammatory effects of 17beta-estradiol were blocked by the antiestrogen ICI 182,780. Additionally, 17beta-estradiol induced rapid phosphorylation of the p42/p44 mitogen-activated protein kinase (MAP kinase), and the MAP kinase inhibitor PD 98059 blocked the antiinflammatory effects of 17beta-estradiol. Overall, these results suggest that estrogen receptor-dependent activation of MAP kinase is involved in estrogen-mediated antiinflammatory pathways in microglial cells. These results describe a novel mechanism by which estrogen may attenuate the progression of neurodegenerative disease and suggest new pathways for therapeutic intervention in clinical settings.

Capacitative Calcium Entry Deficits and Elevated Luminal Calcium Content in Mutant Presenilin-1 Knockin Mice

Abstract: Dysregulation of calcium signaling has been causally implicated in brain aging and Alzheimer's disease. Mutations in the presenilin genes (PS1, PS2), the leading cause of autosomal dominant familial Alzheimer's disease (FAD), cause highly specific alterations in intracellular calcium signaling pathways that may contribute to the neurodegenerative and pathological lesions of the disease. To elucidate the cellular mechanisms underlying these disturbances, we studied calcium signaling in fibroblasts isolated from mutant PS1 knockin mice. Mutant PS1 knockin cells exhibited a marked potentiation in the amplitude of calcium transients evoked by agonist stimulation. These cells also showed significant impairments in capacitative calcium entry (CCE, also known as store-operated calcium entry), an important cellular signaling pathway wherein depletion of intracellular calcium stores triggers influx of extracellular calcium into the cytosol. Notably, deficits in CCE were evident after agonist stimulation, but not if intracellular calcium stores were completely depleted with thapsigargin. Treatment with ionomycin and thapsigargin revealed that calcium levels within the ER were significantly increased in mutant PS1 knockin cells. Collectively, our findings suggest that the overfilling of calcium stores represents the fundamental cellular defect underlying the alterations in calcium signaling conferred by presenilin mutations.

Apoptotic and antiapoptotic mechanisms in stroke

Abstract: Apoptosis is a form of programmed cell death that occurs in neurons during development of the nervous system and may also be a prominent form of neuronal death in chronic neurodegenerative disorders such as Alzheimer's and Parkinson's diseases. Recent findings also implicate apoptosis in neuronal degeneration after ischemic brain injury in animal models of stroke. Activation of both apoptotic and antiapoptotic signaling cascades occurs in neurons in animal and cell culture models of stroke. Apoptotic cascades involve: increased levels of intracellular oxyradicals and calcium; induction of expression of proteins such as Par-4 (prostate apoptosis response-4), which act by promoting mitochondrial dysfunction and suppressing antiapoptotic mechanisms; mitochondrial membrane depolarization, calcium uptake, and release of factors (e.g., cytochrome c) that ultimately induce nuclear DNA condensation and fragmentation; activation of cysteine proteases of the caspase family; activation of transcription factors such as AP-1 that may induce expression of "killer genes." Antiapoptotic signaling pathways are activated by neurotrophic factors, certain cytokines, and increases in oxidative and metabolic stress. Such protective pathways include: activation of the transcription factors (e.g., nuclear factor-kappa B, NF-kappa B) that induce expression of stress proteins, antioxidant enzymes, and calcium-regulating proteins; phosphorylation-mediated modulation of ion channels and membrane transporters; cytoskeletal alterations that modulate calcium homeostasis; and modulation of proteins that stabilize mitochondrial function (e.g., Bcl-2). Intervention studies in experimental stroke models have identified a battery of approaches of potential benefit in reducing neuronal death in stroke patients, including administration of antioxidants, calcium-stabilizing agents, caspase inhibitors, and agents that activate NF-kappa B. Interestingly, recent studies suggest novel dietary approaches (e.g., food restriction and supplementation with antioxidants) that may reduce brain damage following stroke.

Pivotal role for acidic sphingomyelinase in cerebral ischemia-induced ceramide and cytokine production, and neuronal apoptosis.

Abstract: Stroke is a major cause of long-term disability, the severity of which is directly related to the numbers of neurons that succumb to the ischemic insult. The signaling cascades activated by cerebral ischemia that may either promote or protect against neuronal death are not well-understood. One injury-responsive signaling pathway that has recently been characterized in studies of non-neural cells involves cleavage of membrane sphingomyelin by acidic and/or neutral sphingomyelinase (ASMase) resulting in generation of the second messenger ceramide. We now report that transient focal cerebral ischemia induces large increases in ASMase activity, ceramide levels, and production of inflammatory cytokines in wild-type mice, but not in mice lacking ASMase. The extent of brain tissue damage is decreased and behavioral outcome improved in mice lacking ASMase. Neurons lacking ASMase exhibit decreased vulnerability to excitotoxicity and hypoxia, which is associated with decreased levels of intracellular calcium and oxyradicals. Treatment of mice with a drug that inhibits ASMase activity and ceramide production reduces ischemic neuronal injury and improves behavioral outcome, suggesting that drugs that inhibit this signaling pathway may prove beneficial in stroke patients.

Neurotoxicity and dysfunction of dopaminergic systems associated with AIDS dementia.

Abstract: Infection with the human immunodefiency virus (HIV) selectively targets the basal ganglia resulting in loss of dopaminergic neurons Although frequently asymptomatic, some patients may develop sign

The catalytic subunit of telomerase is expressed in developing brain neurons and serves a cell survival-promoting function.

Abstract: Telomerase, a specialized reverse transcriptase (RT) linked to cell immortalization and cancer, has been thought not to be expressed in postmitotic cells. We now report that telomerase activity and its essential catalytic subunit, telomerase reverse transcriptase (TERT), are expressed in neurons in the brains of rodents during embryonic and early postnatal development, and are subsequently downregulated. Suppression of TERT expression in cultured embryonic hippocampal neurons increases their vulnerability to apoptosis and excitotoxicity. Overexpression of TERT in PC12 cells suppresses apoptosis induced by trophic factor withdrawal. TERT exerts its anti-apoptotic action at an early stage of the cell death process prior to mitochondrial dysfunction and caspase activation. TERT may serve a neuron survival-promoting function in the developing brain, and downregulation of TERT in the adult brain may contribute to increased neuronal vulnerability in various age-related neurodegenerative disorders.

Presenilin-1 Mutation Increases Neuronal Vulnerability to Focal Ischemia In Vivo and to Hypoxia and Glucose Deprivation in Cell Culture: Involvement of Perturbed Calcium Homeostasis

Abstract: Many cases of early-onset inherited Alzheimer9s disease (AD) are caused by mutations in the presenilin-1 (PS1) gene. Studies of cultured neural cells suggest that PS1 mutations result in perturbed cellular calcium homeostasis and may thereby render neurons vulnerable to apoptosis. In light of evidence that metabolic impairment plays a role in AD, that cerebral ischemia may be a risk factor for AD, and that individuals with AD have increased morbidity and mortality after stroke, we examined the impact of a PS1 mutation on neuronal vulnerability to ischemic injury. We report that the extent of brain injury after focal cerebral ischemia reperfusion is increased, and behavioral outcome is worsened, in PS1 mutant knock-in mice compared to wild-type mice. Cultured cortical neurons from PS1 mutant mice exhibit increased vulnerability to glucose deprivation and chemical hypoxia compared to their wild-type counterparts. Calcium imaging studies demonstrated enhanced elevation of intracellular calcium levels after glucose deprivation and chemical hypoxia in neurons from PS1 mutant mice. Agents that block calcium release from IP 3 - and ryanodine-sensitive stores (xestospongin and dantrolene, respectively) protected against the endangering action of the PS1 mutation. Our data suggest that presenilin mutations may promote neuronal degeneration in AD by increasing the sensitivity of neurons to age-related ischemia-like conditions. The data further suggest that drugs that stabilize endoplasmic reticulum calcium homeostasis may prove effective in suppressing the neurodegenerative process in AD patients.

Caspase-mediated degradation of AMPA receptor subunits: a mechanism for preventing excitotoxic necrosis and ensuring apoptosis.

Abstract: Activation of ionotropic glutamate receptors of the AMPA and NMDA subtypes likely contributes to neuronal injury and death in various neurodegenerative disorders. Excitotoxicity can manifest as either apoptosis or necrosis, but the mechanisms that determine the mode of cell death are not known. We now report that levels of AMPA receptor subunits GluR-1 and GluR-4 are rapidly decreased in cultured rat hippocampal neurons undergoing apoptosis in response to withdrawal of trophic support (WTS), whereas levels of NMDA receptor subunits NR1, NR2A, and NR2B are unchanged. Exposure of isolated synaptosomal membranes to “apoptotic” cytosolic extracts resulted in rapid degradation of AMPA receptor subunits. Treatment of cells and synaptosomal membranes with the caspase inhibitors prevented degradation of AMPA receptor subunits, demonstrating a requirement for caspases in the process. Calcium responses to AMPA receptor activation were reduced after withdrawal of trophic support and enhanced after treatment with caspase inhibitors. Vulnerability of neurons to excitotoxic necrosis was decreased after withdrawal of trophic support and potentiated by treatment with caspase inhibitors. Our data indicate that caspase-mediated degradation of AMPA receptor subunits occurs during early periods of cell stress and may serve to ensure apoptosis by preventing excitotoxic necrosis.

Neurotrophic factors protect cortical synaptic terminals against amyloid and oxidative stress-induced impairment of glucose transport, glutamate transport and mitochondrial function.

Abstract: Previous studies have shown that several different neurotrophic factors can prevent death of cortical and hippocampal neurons induced by excitotoxic and oxidative insults in cell culture and in vivo. Because neuronal degeneration may be initiated by alterations occurring in synaptic compartments in disorders ranging from Alzheimer's disease to stroke, we tested the hypothesis that neurotrophic factors can exert direct protective actions at the level of the synapse. We now report that a nine amino acid bioactive fragment of activity-dependent neurotrophic factor (ADNF-9) enhances basal glucose and glutamate transport, and attenuates oxidative impairment of glucose and glutamate transport induced by amyloid beta-peptide and Fe(2+), in neocortical synaptosomes. Preservation of transporter function required only short-term (1-2 h) pretreatments. Basic fibroblast growth factor (bFGF) was also effective in suppressing oxidative impairment of synaptic transporter functions, while nerve growth factor (NGF) was less effective. Additional analyses showed that ADNF-9, bFGF and NGF suppress oxidative stress and mitochondrial dysfunction induced by amyloid beta-peptide and Fe(2+) in synaptosomes. Our data suggest that ADNF-9 can act locally in synaptic compartments to suppress oxidative stress and preserve function of glucose and glutamate transporters. Such synapto-protective actions suggest roles for activity-dependent trophic signaling in preventing degeneration of neuronal circuits, and indicate possible therapeutic applications of agents that stimulate local synaptic (transcription-independent) neurotrophic factor signaling pathways.

Apoptotic and anti-apoptotic synaptic signaling mechanisms.

Abstract: Although several prominent morphological features of apoptosis are evident in the cell body (e.g., cell shrinkage, membrane blebbing, and nuclear DNA condensation and fragmentation) the biochemical and molecular cascades that constitute the cell death machinery can be engaged in synaptic terminals and neurites. Initiating events such as oxyradical production and calcium influx, and effector processes such as Par-4 production, mitochondrial alterations and caspase activation, can be induced in synapses and neurites. Several prominent signal transduction pathways in synaptic terminals play important roles in either promoting or preventing neuronal death in physiological and pathological conditions. For example, activation of glutamate receptors in postsynaptic spines can induce neuronal apoptosis, whereas local activation of neurotrophic factor receptors in presynaptic terminals can prevent neuronal death. Factors capable of inducing nuclear chromatin condensation and fragmentation can be produced locally in synaptic terminals and neurites, and may propogate to the cell body. Recent findings suggest that, beyond their roles in inducing or preventing cell death, apoptotic and anti-apoptotic cascades play roles in synaptic plasticity (structural remodelling and long-term functional changes). For example, caspase activation results in proteolysis of glutamate receptor (AMPA) subunits, which results in altered neuronal responsivity to glutamate. Activation of neurotrophic factor receptors in synaptic terminals can result in local changes in energy metabolism and calcium homeostasis, and can induce long-term changes in synaptic transmission. The emerging data therefore suggest that synapses can be considered as autonomous compartments in which both pro- and anti-apoptotic signaling pathways are activated resulting in structural and functional changes in neuronal circuits. A better understanding of such synaptic signaling mechanisms may reveal novel approaches for preventing and treating an array of neurodegenerative conditions that are initiated by perturbed synaptic homeostasis.

Pro-apoptotic action of PAR-4 involves inhibition of NF-kappaB activity and suppression of BCL-2 expression.

Abstract: Par-4(1) (prostate apoptosis response 4) is known to function at an early stage in apoptosis in several different cell types, including neurons. On the other hand, activation of the transcription factor NF-kappaB can prevent apoptosis in various cancer cells and neurons. We now report that overexpression of full-length Par-4 in cultured PC12 cells results in a suppression of basal NF-kappaB DNA-binding activity and NF-kappaB activation following trophic factor withdrawal (TFW). The decreased NF-kappaB activity is correlated with enhanced apoptosis. Conversely, NF-kappaB activity is increased and vulnerability to apoptosis reduced in cells overexpressing a dominant-negative form of Par-4. Par-4 overexpression or functional blockade had no effect on AP-1 DNA-binding activity. Expression of the antiapoptotic protein Bcl-2 was dramatically reduced in PC12 cells overexpressing Par-4. Our data suggest that suppression of NF-kappaB activation plays a major role in the proapoptotic function of Par-4.

The prostate apoptosis response-4 protein participates in motor neuron degeneration in amyotrophic lateral sclerosis

Abstract: Prostate apoptosis response-4 (Par-4), a protein containing a leucine zipper domain within a death domain, is up-regulated in prostate cancer cells and hippocampal neurons induced to undergo apoptosis. Here, we report higher Par-4 levels in lumbar spinal cord samples from patients with amyotrophic lateral sclerosis (ALS) than in lumbar spinal cord samples from neurologically normal patients. We also compared the levels of Par-4 in lumbar spinal cord samples from wild-type and transgenic mice expressing the human Cu/Zn-superoxide dismutase gene with a familial ALS mutation. Relative to control samples, higher Par-4 levels were observed in lumbar spinal cord samples prepared from the transgenic mice at a time when they had hind-limb paralysis. Immunohistochemical analyses of human and mouse lumbar spinal cord sections revealed that Par-4 is localized to motor neurons in the ventral horn region. In culture studies, exposure of primary mouse spinal cord motor neurons or NSC-19 motor neuron cells to oxidative ...

Neuroprotective role for the p50 subunit of NF-kappaB in an experimental model of Huntington's disease.

Abstract: Prototypical NF-kappaB consists of a transcription factor dimer of p50 and p65, and an inhibitory subunit called I-kappaB. NF-kappaB is activated in neurons in response to excitotoxic, metabolic, and oxidative stress. Cell-culture data suggest that activation of NF-kappaB can prevent neuronal apoptosis, but its role in vivo is unclear and the specific kappaB subunits involved are unknown. In Huntington's disease (HD), striatal neurons degenerate, and a similar pattern of neuronal vulnerability occurs in rats and mice following exposure to the mitochondrial toxin 3-nitropropionic acid (3NP). We report that mice lacking the p50 subunit of NF-kappaB exhibit increased damage to striatal neurons following administration of 3NP. The neuronal death occurs by apoptosis as indicated by increased caspase activation and DNA fragmentation into oligonucleosomes. NF-kappaB activity is markedly increased in striatum 24-72 h following 3NP administration in wild-type mice, but not in mice lacking p50, indicating that p50 is necessary for the vast majority of 3NP-induced NF-kappaB DNA-binding activity in striatum. Cultured striatal neurons from p50-/- mice exhibited enhanced oxidative stress, perturbed calcium regulation, and increased cell death following exposure to 3NP, suggesting a direct adverse effect of p50 deficiency in striatal neurons.

ALS-Linked Cu/Zn-SOD Mutation Impairs Cerebral Synaptic Glucose and Glutamate Transport and Exacerbates Ischemic Brain Injury:

Abstract: Although degeneration of lower motor neurons is the most striking abnormality in amyotrophic lateral sclerosis (ALS), more subtle alterations may occur in the brain. Mutations in copper/zinc superoxide dismutase (Cu/Zn-SOD) are responsible for some cases of inherited ALS, and expression of mutant Cu/Zn-SOD in transgenic mice results in progressive motor neuron loss and a clinical phenotype similar to that of ALS patients. It is now reported that Cu/Zn-SOD mutant mice exhibit increased vulnerability to focal ischemic brain injury after transient occlusion of the middle cerebral artery. Levels of glucose and glutamate transport in cerebral cortex synaptic terminals were markedly decreased, and levels of membrane lipid peroxidation were increased in Cu/Zn-SOD mutant mice compared to nontransgenic mice. These findings demonstrate that mutant Cu/Zn-SOD may endanger brain neurons by a mechanism involving impairment of glucose and glutamate transporters. Moreover, our data demonstrate a direct adverse effect of the mutant enzyme on synaptic function.