1,720,979 research outputs found
Dysregulation of System Xc- and Its Impact on Cocaine Seeking
Full text linkAbnormal glutamate signaling in the brain, particularly in the nucleus accumbens core, contributes to compulsive cocaine-seeking behavior. In the nucleus accumbens, two distinct pools of glutamate exist. The synaptic pool stimulates excitatory postsynaptic receptors, leading to cocaine seeking. Cystine-glutamate exchange by system xc- contributes to the extrasynaptic pool and regulates neurotransmission by activating extrasynaptic receptors including inhibitory Group II metabotropic glutamate receptors, positioning this as an important mechanism for regulating nucleus accumbens activity. We and others have found decreased levels of cystine-glutamate exchange in animals withdrawn from chronic cocaine administration. In chapter 1, we describe studies directly implicating diminished system xc- activity in abnormal glutamate signaling necessary for promoting drug seeking. Given that altered system xc- activity is essential to drug seeking, understanding the cellular basis of reduced cystine-glutamate exchange would advance our knowledge of the neural basis of addiction. Unfortunately, little is known about how the system is regulated. To address this, we sought to define conditions that permit the study of system xc- (chapter 2). In chapter 3, we examined potential regulation of xc- by dopamine, nonspecific dopamine agonists, D1-liked dopamine receptor agonists, and D2-like dopamine receptor agonists. While we found D1-like receptor regulation of cystine-glutamate exchange in mixed cortical culture, we did not observe this effect in tissue punches; thus, it is unlikely that abnormal regulation of system xc- by dopamine underlies reduced cystine-glutamate exchange in cocaine-withdrawn animals. In chapter 4, we examined pituitary adenylyl cyclase activating polypeptide (PACAP) because it has been recently shown to regulate extracellular glutamate homeostasis. We found acute and tonic regulation of cystine-glutamate exchange by PACAP-induced stimulation of Ca2+-dependent protein kinase (PKC) activity. More importantly, we also determined that PACAP regulation of system xc- is diminished in animals withdrawn from cocaine self-administration. These findings suggest dysregulated PACAP neurotransmission in cocaine-withdrawn animals contributes to the neural basis of addiction. By revealing novel mechanisms contributing to addiction, scientists can identify novel targets for therapeutic treatment
Regulation of System XC- and its Contribution to Cell Death
Full text linkThe main focus of the studies in this thesis involves examining the role of cystine/glutamate exchange (system xC-) in neuronal death in primary cortical cell culture, with an emphasis on how glial function affects neuronal cell death. System xC- is a sodium-independent transporter that mediates cystine uptake and glutamate release. It accounts for most of the cystine uptake in astrocytes in mature cultures, providing the rate limiting substrate for synthesis of the main endogenous antioxidant glutathione. The glutamate released by system xC- may lead to excessive extracellular glutamate and cause excitotoxicity.
β-N-methylamino-L-alanine (BMAA) is a non-protein amino acid that may be involved in neurodegenerative diseases. We found that BMAA induced oxidative stress by competing with cystine at system xC- leading to depletion of glutathione. BMAA also drives system xC- mediated glutamate release, which may contribute to its induction of excitotoxicity.
Fibroblast growth factor-2 (FGF-2) is involved in multiple processes in the central nervous system, including plasticity, neurogenesis, differentiation, and neuronal survival. Also, alterations in FGF-2 and its signaling have been implicated in neurodegenerative diseases and psychiatric disorders. We found that FGF-2 greatly increased cystine uptake through system xC- in astrocyte-enriched primary cultures, but not in neuronal or microglial cultures. Our data showed that FGF-2 increased cystine uptake by upregulating system xC- by acting on FGFR1, and signaling through the PI3K/Akt and MEK/ERK pathways.
FGF-2 treatment for 48 hours caused significant neuronal death only in mixed neuronal and glial cultures, but not in neuronal-enriched or astrocyte-enriched cultures. Blocking system xC-, or AMPA/kainate receptors, eliminated the neuronal death induced by FGF-2 treatment. Therefore, it is likely that 48 hour FGF-2 treatment induces AMPA receptor mediated toxicity through increased glutamate release from astrocytes due to increased system xC- function. However, we cannot exclude the possibility that FGF-2 treatment sensitizes the neurons to normal system xC- mediated glutamate release.
Together the results indicate that 1) competitive substrates of system xC-, such as BMAA, that do not lead to glutathione production are particularly toxic; and 2) upregulation of system xC- on astrocytes may be toxic to surrounding neurons
Going Beyond Counting First Authors in Author Co-citation Analysis
Full text linkThe present study examines one of the fundamental aspects of author co-citation analysis (ACA) - the way co-citation
counts are defined. Co-citation counting provides the data on which all subsequent statistical analyses and mappings
are based, and we compare ACA results based on two different types of co-citation counting - the traditional type that
only counts the first one among a cited work's authors on the one hand and a non-traditional type that takes into
account the first 5 authors of a cited work on the other hand. Results indicate that the picture produced through this non-traditional author co-citation counting contains more coherent author groups and is therefore considerably clearer. However, this picture represents fewer specialties in the research field being studied than that produced through the traditional first-author co-citation counting when the same number of top-ranked authors is selected and analyzed. Reasons for these effects are discussed
Mechanisms of Neuronal Death Induced by Environmental Toxicants in Murine Cortical Culture
Full text linkThis study was directed at examining the neurotoxic mechanisms of several classes of environmental toxicants implicated in neurodegenerative disease. Primary cortical cultures were exposed to organophosphorus pesticides, heavy metals and the cyanobacterial toxin, beta-N-methylamino-L-alanine (BMAA). Several components relating to neuronal injury were assessed in each study and novel aspects are described.
The main action of organophosphorous insecticides is generally believed to be the inhibition of acetylcholinesterase. However, these compounds are now recognized to inhibit many other enzymes and cause neuronal death through a variety of mechanisms. I found that exposure to chlorpyrifos or diazinon caused concentration-dependent neurotoxicity that could not be attributed to acetylcholinesterase inhibition. Chlorpyrifos exposure increased extracellular glutamate and induced a diffuse nuclear staining characteristic of necrosis; the toxicity was sensitive to ionotropic glutamate receptor antagonists. Diazinon toxicity was blocked by caspase inhibitors. Additionally, diazinon induced punctuate chromatin staining characteristic of apoptosis. These results represent two distinct, novel mechanisms of organophosphorous neurotoxicity.
Heavy metals are ubiquitous in the environment and are of significant health concern worldwide. Exposure to lead, iron, mercurials (inorganic mercury, methylmercury, or thimerosal, i.e. ethylmercury) or other heavy metals is implicated as a risk factor for neurodegenerative disease. I found that the toxicity of these metals may be enhanced when interacting with chelators used to treat metal intoxication. As well, my studies describe a new role for mercury-induced oxidative stress as a cytoprotective signal to enhance glutathione levels. My data also suggests an obligate role for MRP1 in the detoxification of methylmercury.
Neurodegenerative diseases likely involve complex interactions between genetic predisposition and multiple environmental factors. My final study tested the interaction of the methylmercury and BMAA. Importantly, concentrations of BMAA that caused no toxicity by themselves potentiated methyl mercury toxicity. BMAA plus methylmercury, at concentrations that had no effect by themselves, depleted cellular glutathione. The combined toxicity was attenuated by glutathione monoethyl ester, and the free radical scavenger, trolox, but not by the NMDA receptor antagonist, MK-801. The results indicate a synergistic neurotoxic interaction targeting the cellular redox state. This finding may have implications for neurodegenerative disease caused by environmental toxicant exposure
Neurotoxic Evaluation of Root-End Filling Materials in Cortical Culture
No full textTo date, there have not been any published studies designed to determine the toxicity of MTA, amalgam, Super EBA, and Diaket on neurons in culture systems. The objective of this in vitro study was, therefore, to evaluate the neurotoxicity of these four commonly used root-end filling materials in murine cortical cell cultures
Regulation of the Cystine/Glutamate Antiporter and its Contribution to Neuronal Death
Full text linkThe aim of this thesis is to better understand the regulation of the cystine/glutamate antiporter (system xc-) and its role in regulating neuronal survival and death. Expressed primarily on astrocytes, system xc- takes up cystine and releases glutamate in a 1:1 ratio. Cystine uptake is the rate-limiting step in glutathione synthesis, the brain’s main antioxidant. Glutamate released into the extrasynaptic space can regulate neuronal function; however excessive glutamate release can cause excitotoxicity. The dual actions of system xc- make it of interest in many neurodegenerative diseases where oxidative stress and excitotoxicity are involved. We investigated the regulation of system xc- in SOD1-G93A transgenic mouse model of ALS. We observed an increase in cystine uptake and glutamate release through system xc- in spinal cord slices of SOD1-G93A transgenic mice. We did not observe a change in the function of the main glutamate clearance transporter, excitatory amino acid transporter (EAAT). This study was the first to show that system xc- activity is dysregulated in an ALS model and suggests that the excitotoxicity in the SOD1-G93A transgenic mouse may be due to increased system xc- activity. Using primary mixed cortical cultures we assessed how different compounds that deplete intracellular glutathione (GSH), L-buthionine-sulfoximine (BSO) and diethyl maleate (DEM), affect system xc- function. Both compounds caused significant decreases in intracellular GSH levels; however, DEM caused an increase in cystine uptake through system xc-, while unexpectedly BSO caused a decrease in uptake. Also, DEM caused a decrease in intracellular cysteine, while BSO increased cysteine levels. The results suggest that negative feedback by intracellular cysteine is a more important regulator of system xc- than intracellular GSH. Transforming growth factor-β1 (TGF-β1) is a cytokine involved in regulating many cellular processes, including neuronal survival and death. We found that TGF-β1 increased cystine uptake through system xc- in astrocyte-enriched glial cultures via the MAPK/ERK pathway. TGF-β1 increased the export of GSH from astrocytes, which suggests a neuroprotective role; however, in mixed cortical cultures TGF-β1 enhanced rotenone-induced neurotoxicity through AMPA receptors. The data suggests that the increase in system xc- activity by TGF-β1 may have antioxidant defenses, but also exacerbates excitotoxicity
In Vitro Cytotoxicity of Orthodontic Cements and Archwires
No full textThe orthodontic literature has reported on the cytotoxicity of orthodontic materials in various culture systems. However, there have been no studies that have examined the cytotoxic effects of orthodontic archwires and cements on neuronal cells. Moreover, little is known concerning the mechanism of toxicity for these popular compounds. This investigation used murine cortical cell cultures to examine the in vitro neurotoxicity of two orthodontic adhesives and five commonly used orthodontic metallic archwire alloys. The materials examined included the composite resin, Transbond TM XT, the compomer cement. Band-Lok TM, 0.016 inch nickel-titanium. copper-nickel-titanium, titanium-molybdenum, Elgiloy, and stainless steel archwire alloys. Standard sized samples of each material were placed on tissue culture inserts suspended above the cell cultures. Neuronal death was determined using the lactate dehydrogenase release assay 24 hours after exposure to the archwires. The results indicated that Band-Lok™ was significantly toxic and remained so even after seven days of setting. Washing Band-Lok for seven days in Eagles \u27 MEM virtually eliminated neurotoxicity. Transbond™ XT was not significantly toxic. With respect to archwire alloys, the results indicated that nickel-titanium, copper-nickel-titanium and titanium-molybdenum alloys were not neurotoxic, while stainless steel and Elgiloy were significantly toxic. Washing the archwires for 7 days in Eagle\u27s MEM did not alter the toxicity. However, the free radical scavenger, trolox blocked the toxicity of both stainless steel and Elgiloy indicating that the death was free radical mediated. The caspase inhibitor, ZVAD blocked the toxicity of stainless steel, but not Elgiloy, suggesting that stainless steel induced apoptosis. Further evidence that stainless steel induced apoptosis was provided by propidium iodide staining which showed nuclear chromatin condensation and fragmentation into discrete spherical or irregular shapes characteristic of apoptosis. Given the composition of the toxic archwires, the fact that the death was free radical mediated, and the type of death occurring, the results are most consistent with the toxicity of Elgiloy being caused by the release of iron and the toxicity of stainless steel being mediated by the release of chromium. While these results may not have direct clinical applications to orthodontic practice, clinicians should consider the biological properties of the materials they use in patients when performing orthodontic treatment
Variations on the Author
Full text link“Variations on the Author” discusses two of Eduardo Coutinho’s recent films (Um Dia na Vida, from 2010, and Últimas Conversas, posthumously released in 2015) and their contribution to the general question of documentary authorship. The director’s filmography is characterized by a consistent yet self-effacing form of authorial self-inscription: Coutinho often features as an interviewer that rather than express opinions propels discourses; an interviewer that is good at listening. This mode of self-inscription characterizes him as an author who is not expressive but who is nonetheless markedly present on the screen. In Um Dia na Vida, however, Coutinho is completely absent form the image, while Últimas Conversas, on the contrary, includes a confessional prologue that moves the director from the margins to the center of his films. This article examines the ways in which these works stand out in the filmography of a director who offers new insights into the notion of cinematic authorship
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