Ian J. Reynolds's research while affiliated with Oregon Health and Science University and other places
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Publications (180)
Zinc is a highly abundant cation in the brain, essential for cellular functions, including transcription, enzymatic activity, and cell signaling. However, zinc can also trigger injurious cascades in neurons, contributing to the pathology of neurodegenerative diseases. Mitochondria, critical for meeting the high energy demands of the central nervous...
Zinc is a highly abundant cation in the brain, where it is essential for cellular function, including transcription, enzymatic activity, and cell signaling. However, zinc can also trigger injurious cascades in neurons, contributing to the pathology of neurodegenerative diseases. Mitochondria, critical for meeting the high energy demands of the cent...
This special issue of Frontiers in Neuroscience-Neurodegeneration celebrates the 50th anniversary of John Olney’s seminal work introducing the concept of excitotoxicity as a mechanism for neuronal cell death. Since that time, fundamental research on the pathophysiological activation of glutamate receptors has played a central role in our understand...
There is growing evidence that activation of the mGlu4 receptor leads to anxiolytic- and antipsychotic-like efficacy in rodent models, yet its relevance to depression-like reactivity remains unclear. Here, we present the pharmacological evaluation of ADX88178, a novel potent, selective and brain-penetrant positive allosteric modulator (PAM) of mGlu...
Adenosine A2A receptors are predominantly localized on striatopallidal gamma-aminobutyric acid (GABA) neurons, where they are colocalized with dopamine D2 receptors and are involved in the regulation of movement. Adenosine A2A receptor antagonists have been evaluated as a novel treatment for Parkinson's disease and have demonstrated efficacy in a b...
Affecting over 1.5 million people across the world, Parkinson's disease is a progressive neurological condition characterized, in part, by the loss of dopaminergic neurons in the substantia nigra pars compacta. It affects 1.5% of the global population over 65 years of age. As life expectancy is increasing, over the next few years the number of pati...
Positive allosteric modulators (PAMs) of metabotropic glutamate receptor 4 (mGluR4) have been proposed as a novel therapeutic approach for the treatment of Parkinson's disease. However, evaluation of this proposal has been limited by the availability of appropriate pharmacological tools to interrogate the target. In this study, we describe the prop...
Mutations in the leucine-rich repeat kinase 2 gene (LRRK2) are the most common genetic cause of Parkinson's disease (PD) and cause both autosomal dominant familial and sporadic PD. Currently, the physiological and pathogenic activities of LRRK2 are poorly understood. To decipher the biological functions of LRRK2, including the genes and pathways mo...
J. Neurochem. (2011) 118, 1016–1031.
The EphA4 receptor and its ephrin ligands are involved in astrocytic gliosis following CNS injury. Therefore, a strategy aimed at the blockade of EphA4 signaling could have broad therapeutic interest in brain disorders. We have identified novel small molecule inhibitors of EphA4 kinase in specific enzymatic and...
Accumulation of small soluble assemblies of amyloid-β (Aβ)(42) in the brain is thought to play a key role in the pathogenesis of Alzheimer's disease. As a result, there has been much interest in finding small molecules that inhibit the formation of synaptotoxic Aβ(42) oligomers that necessitates sensitive methods for detecting the initial steps in...
Apolipoprotein (apo) E4 is the major genetic risk factor for late-onset Alzheimer disease (AD). ApoE4 assumes a pathological conformation through an intramolecular interaction mediated by Arg-61 in the amino-terminal domain and Glu-255 in the carboxyl-terminal domain, referred to as apoE4 domain interaction. Because AD is associated with mitochondr...
ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.
Poly ADP-ribose polymerase (PARP) maintains genomic integrity by repairing DNA strand breaks, however over-activation of PARP following neural tissue injury is hypothesized to cause neuronal death. Therefore, PARP inhibitors have potential for limiting neural injury under certain conditions. A reliable method for assessing PARP activity in brain is...
ChemInform is a weekly Abstracting Service, delivering concise information at a glance that was extracted from about 100 leading journals. To access a ChemInform Abstract of an article which was published elsewhere, please select a “Full Text” option. The original article is trackable via the “References” option.
Alterations in mitochondrial function may have a central role in the pathogenesis of many neurodegenerative diseases. The study of mitochondrial dysfunction has typically focused on ATP generation, calcium homeostasis and the production of reactive oxygen species. However, there is a growing appreciation of the dynamic nature of mitochondria within...
Mitochondria are widely believed to be the source of reactive oxygen species (ROS) in a number of neurodegenerative disease states. However, conditions associated with neuronal injury are accompanied by other alterations in mitochondrial physiology, including profound changes in the mitochondrial membrane potential ΔΨm. In this study we have invest...
The discovery of a novel series of potent and selective T-type calcium channel antagonists is reported. Initial optimization of high-throughput screening leads afforded a 1,4-substituted piperidine amide 6 with good potency and limited selectivity over hERG and L-type channels and other off-target activities. Further SAR on reducing the basicity of...
Liberation of zinc from intracellular stores contributes to oxidant-induced neuronal injury. However, little is known regarding how endogenous oxidant systems regulate intracellular free zinc ([Zn(2+)](i)). Here we simultaneously imaged [Ca(2+)](i) and [Zn(2+)](i) to study acute [Zn(2+)](i) changes in cultured rat forebrain neurons after glutamate...
Excessive accumulation of intracellular free zinc is lethal to neurons. The precise mechanism by which zinc kills neurons remains unclear, however, a number of reports suggest that zinc may impair neuronal metabolism. The present study examined how zinc affects the activity of isolated rat brain mitochondria. Using spectrofluorometry and polarograp...
Recent studies indicate that ATP and UTP act at G protein-coupled (P2Y) nucleotide receptors to excite nociceptive sensory neurons; nucleotides also potentiate signaling through the pro-nociceptive capsaicin receptor, TRPV1. We demonstrate here that P2Y(2) is the principal UTP receptor in somatosensory neurons: P2Y(2) is highly expressed in dorsal...
IntroductionMitochondrial mechanisms of peripheral neuropathyMitochondria and retinal drug toxicityMitochondria and ototoxicityMitochondrial mechanisms of CNS injuryConclusions
References
NADH dehydrogenase subunit 2, encoded by the mtDNA, has been associated with resistance to autoimmune type I diabetes (T1D)
in a case control study. Recently, we confirmed a role for the mouse ortholog of the protective allele (mt-Nd2a) in resistance to T1D using genetic analysis of outcrosses between T1D-resistant ALR and T1D-susceptible NOD mice....
There is an intimate relationship between ion transport and energy metabolism in the brain. All ion transport is driven directly
or indirectly by ATP, and the support of ion homeostasis represents the largest demand on energy production in the brain.
Failure of ion homeostasis because of the interruption of energy generation has devastating consequ...
There has been a steady accumulation of data that point to a role for mitochondria in the pathogenesis of Parkinson’s disease (PD). In broad terms, it is evident that toxins that impair mitochondrial function produce lesions resembling PD in animals and humans, and that mitochondria may be a major source of reactive oxygen species (ROS) that could...
Mitochondria are the primary generators of ATP and are important regulators of intracellular calcium homeostasis. These organelles are dynamically transported along lengthy neuronal processes, presumably for appropriate distribution to cellular regions of high metabolic demand and elevated intracellular calcium, such as synapses. The removal of dam...
Mitochondria have many roles critical to the function of neurons including the generation of ATP and regulation of intracellular Ca2+. Mitochondrial movement is highly dynamic in neurons and is thought to direct mitochondria to specific cellular regions of increased need and to transport damaged or old mitochondria to autophagosomes. Morphology als...
Functional synapses require mitochondria to supply ATP and regulate local [Ca2+]i for neurotransmission. Mitochondria are thought to be transported to specific cellular regions of increased need such as synapses. However, little is known about how this occurs, including the spatiotemporal distribution of mitochondria relative to presynaptic and pos...
Nitric oxide (NO) has a number of physiological and pathophysiological effects in the nervous system. One target of NO is the mitochondrion, where it inhibits respiration and ATP synthesis, which may contribute to NO-mediated neuronal injury. Our recent studies suggested that impaired mitochondrial function impairs mitochondrial trafficking, which...
Huntington's disease (HD) is a neurodegenerative disorder caused by a polyglutamine repeat in the huntingtin gene (Htt). Mitochondrial defects and protein aggregates are characteristic of affected neurons. Recent studies suggest that these aggregates impair cellular transport mechanisms by interacting with cytoskeletal components and molecular moto...
We have investigated the properties of the dopaminergic neurotoxins 6-hydroxydopamine, 1-methyl-4-phenylpyridinium and rotenone using an organotypic culture that included slices of substantia nigra, striatum and cortex maintained for about 20 days in vitro. At this age, the organotypic culture contains dopaminergic neurons, visualized using tyrosin...
![Figure 1. [Zn 2 ] i inhibits mitochondrial movement at cytotoxic...](profile/Anthony-Honick/publication/7544130/figure/fig1/AS:424376312045569@1478190605266/Zn-2-i-inhibits-mitochondrial-movement-at-cytotoxic-concentrations-a-Neurons_Q320.jpg)
Mitochondria have been identified as targets of the neurotoxic actions of zinc, possibly through decreased mitochondrial energy production and increased reactive oxygen species accumulation. It has been hypothesized that impairment of mitochondrial trafficking may be a mechanism of neuronal injury. Here, we report that elevated intraneuronal zinc i...
Ca(2+)-induced mitochondrial depolarization was studied in single isolated rat brain and liver mitochondria. Digital imaging techniques and rhodamine 123 were used for mitochondrial membrane potential measurements. Low Ca(2+) concentrations (about 30--100 nM) initiated oscillations of the membrane potential followed by complete depolarization in br...
Current evidence suggests that zinc kills neurons by disrupting energy production, specifically by inhibiting mitochondrial function. However it is unclear if the inhibitory effect requires zinc accumulation, and if so, precisely how zinc enters mitochondria. Here, using fluorescence microscopy to visualize individual rat brain mitochondria, we det...
Mitochondrial complex I dysfunction has been implicated in a number of brain pathologies, putatively owing to an increased rate of reactive oxygen species (ROS) release. However, the mechanisms regulating the ROS burden are poorly understood. In this study we investigated the effect of Ca2+ loads on ROS release from rat brain mitochondria with comp...
Functional deficits following traumatic brain injury (TBI) are associated with alterations in markers of dopaminergic neurotransmission. To assess the effects of TBI on the expression and functional integrity of dopamine transporters, we measured transporter protein levels and investigated synaptosomal dopamine uptake in the rat striatum. Two or fo...
Elevation of intracellular free zinc ([Zn2+]i) probably contributes to cell death in injury paradigms involving calcium deregulation and oxidative stress such as glutamate excitotoxicity. However, it is difficult to monitor both ions simultaneously in live cells. Here we present a new method using fluorescence microscopy and the ion sensitive indic...
Emerging evidence suggests that Zn2+ may impair neuronal metabolism. We examined how Zn2+ affects the activity of isolated brain mitochondria fueled with glutamate + malate, succinate or glycerol 3-phosphate. Submicromolar levels of Zn2+ dissipated membrane potential and inhibited oxygen utilization in all three substrate conditions. Zn(2+)-induced...
In this study we investigated fluctuations in mitochondrial membrane potential (DeltaPsim) in single isolated brain mitochondria using fluorescence imaging. Mitochondria were attached to coverslips and perfused with K+-based buffer containing 20 microM EDTA, supplemented with malate and glutamate, and rhodamine 123 for DeltaPsim determination. Delt...
We report here that reduced pyridine nucleotides and reduced glutathione result in an oxidation of Amplex Red by dioxygen that is dependent on the presence of horseradish peroxidase (HRP). Concentrations of NADH and glutathione typically found in biological systems result in the oxidation of Amplex Red at a rate comparable to that produced, for exa...
In order to satisfy the metabolic and ion homeostasis demands of neurons, mitochondria must be transported to appropriate locations within cells. Although it is well established that much of this trafficking occurs on microtubules and, to a lesser extent, actin, the mechanisms by which the trafficking of mitochondria is controlled are poorly unders...
Mitochondria are the proximate target of a number of different neurotoxins. Typically, impairing of the key bioenergetic function of mitochondria by toxins is considered as the main mechanism of action. However, the effective maintenance of energy generation in neurons depends on the biogenesis, trafficking, and degradation of mitochondria in addit...
HIV infection in humans and simian immunodeficiency virus (SIV) infection in macaques result in encephalitis in approximately one-quarter of infected individuals and is characterized by infiltration of the brain with infected and activated macrophages. 1-(2-chlorphenyl)-N-methyl-N-(1-methylpropyl)-3-isoquinoline-carboxamide (PK11195) is a ligand sp...
How m-calpain is activated in cells has challenged investigators because in vitro activation requires near-millimolar calcium.
Previously, we demonstrated that m-calpain activation by growth factors requires extracellular signal-regulated kinase (ERK);
this enables tail deadhesion and allows productive motility. We now show that ERK directly phosph...
Detection of neurotoxic metals in the intracellular milieu has made an important contribution to the understanding of the mechanism of metal-induced neuronal injury. Fluorescent, metal-sensitive dyes have proven to be valuable in the measurement of a variety of neurotoxic cations in neurons, and these dyes have provided a number of insights into th...
Excessive accumulation of the heavy metal zinc is cytotoxic. As a consequence, cellular vulnerability to zinc-induced injury may be regulated by the abundance of proteins that maintain intracellular free zinc concentrations ([Zn2+]i). In this study, we overexpressed the zinc-binding protein metallothionein-II (MT) in astrocytes to assess its impact...
The catalytic subunit of telomerase reverse transcriptase (TERT) protects dividing cells from replicative senescence in vitro. Here, we show that expression of TERT mRNA is induced in the ipsilateral cortical neurons after occlusion of the middle cerebral artery in adult mice. Transgenic mice that overexpress TERT showed significant resistance to i...
In this study we measured DeltaPsim in single isolated brain mitochondria using rhodamine 123. Mitochondria were attached to coverslips and superfused with K(+)-based HEPES-buffer medium supplemented with malate and glutamate. In approximately 70% of energized mitochondria we observed large amplitude spontaneous fluctuations in DeltaPsim with a tim...
In this study we investigated whether the link between mitochondrial dysfunction and deregulation of Ca(2+) homeostasis preceding excitotoxic cell death is mediated by cellular deenergization. Glycolytic and/or mitochondrial ATP synthesis was inhibited with 2-deoxy-D-glucose (2DG) and oligomycin, respectively. Changes in cytoplasmic Ca(2+) concentr...
In this study we investigated whether the link between mitochondrial dysfunction and deregulation of Ca2+ homeostasis preceding excitotoxic cell death is mediated by cellular deenergization. Glycolytic and/or mitochondrial ATP synthesis was inhibited with 2-deoxy-d-glucose (2DG) and oligomycin, respectively. Changes in cytoplasmic Ca2+ concentratio...
Mitochondria are essential to maintain neuronal viability. In addition to the generation of ATP and maintenance of calcium homeostasis, the effective delivery of mitochondria to the appropriate location within neurons is also likely to influence their function. In this study we examined mitochondrial movement and morphology in primary cultures of r...
Mitochondria are a node of integration for intracellular signaling pathways and their morphology changes seem to be tightly associated with their function. New data show that morphology is one of the parameters involved in mitochondria's choice between promoting cell death and protecting cells against general metabolic jeopardy.
An increasing body of evidence suggests that high intracellular free zinc promotes neuronal death by inhibiting cellular energy production. A number of targets have been postulated, including complexes of the mitochondrial electron transport chain, components of the tricarboxylic acid cycle, and enzymes of glycolysis. Consequences of cellular zinc...
Mitochondria may be both the source and the target of oxidative stress in neurodegenerative disease. In models of excitotoxicity, neuronal injury is triggered by the influx of calcium into neurons and then into mitochondria. Our studies suggest that an important consequence of this calcium movement is the generation of ROS by mitochondria. Studies...
![Fig. 4. [Zn 2 ] i-sensitive dyes exhibit similar response profiles,...](profile/Latha-Malaiyandi/publication/11206809/figure/fig2/AS:646705041113088@1531197905772/Zn-2-i-sensitive-dyes-exhibit-similar-response-profiles-despite-varying-affinities_Q320.jpg)
![Fig. 5. [Zn 2 ] i summary of [Zn 2 ] i-induced changes in Zn...](profile/Latha-Malaiyandi/publication/11206809/figure/fig3/AS:646705041137665@1531197905796/Zn-2-i-summary-of-Zn-2-i-induced-changes-in-Zn-2-sensitive-dyes-For-each-dye-in_Q320.jpg)
The emergence of zinc as a potent neurotoxin has prompted the development of techniques suitable for the measurement of intracellular free zinc ([Zn(2+)](i)) in cultured cells. Accordingly, a new family of Zn(2+)-sensitive fluorophores has become available. Using ionophore-induced elevations of [Zn(2+)](i) in cultured neurons, we measured [Zn(2+)](...
![Figure 4. Hydrogen peroxide modulation of [Fe 2 ] i. A, Neurons were...](profile/Geraldine-Kress/publication/11257024/figure/fig6/AS:668409213747210@1536372583277/Hydrogen-peroxide-modulation-of-Fe-2-i-A-Neurons-were-perfused-with-TBSS-and-exposed_Q320.jpg)
Iron is an essential element for cells but may also be an important cytotoxin. However, very little is known about iron transport, redox status, or toxicity specifically inside cells. In this study, we exploited the sensitivity of fura-2 to quenching by ferrous iron (Fe(2+)) to detect intracellular free iron ([Fe(2+)](i)) in neurons, astrocytes, an...
We hypothesized that: (a) S-nitrosylation of metallothionein (MT) is a component of pulmonary endothelial cell nitric oxide (NO) signaling that is associated with an increase in labile zinc; and (b) NO mediated increases in labile zinc in turn reduce the sensitivity of pulmonary endothelium to LPS-induced apoptosis. We used microspectrofluorometric...
We hypothesized that metallothionein (MT), a cysteine-rich protein with a strong affinity for Zn(2+), plays a role in nitric oxide (NO) signaling events via sequestration or release of Zn(2+) by the unique thiolate clusters of the protein. Exposing mouse lung fibroblasts (MLF) to the NO donor S-nitrosocysteine resulted in 20-30% increases in fluore...
Using the H(+)-sensitive fluorophore 2',7'-bis-(2-carboxyethyl)-5-(and-6)-carboxyfluorescein (BCECF) and microfluorimetry, we investigated how elevated intracellular free zinc ([Zn(2+)](i)) altered intracellular proton concentration (pH(i)) in dissociated cultures of rat forebrain neurons. Neurons exposed to extracellular zinc (3 microM) in the pre...
We hypothesized that metallothionein (MT), a cysteine-rich protein with a strong affinity for Zn2+, plays a role in nitric oxide (NO) signaling events via sequestration or release of Zn2+ by the unique thiolate clusters of the protein. Exposing mouse lung fibroblasts (MLF) to the NO donor S-nitrosocysteine resulted in 20-30% increases in fluorescen...
Glutamate can injure neurons in a way that is likely relevant to a number of acute neurodegenerative states, including stroke
and head trauma. It is also likely that glutamate contributes to neurons degeneration in chronic diseases as well. Many studies
have established a relationship between the bioenergetic state of neurons and their vulnerabilit...
![Fig. 4. Vanilloid compounds elicit increased [Ca 2 ϩ ] i and NO release...](profile/William-Groat/publication/11743363/figure/fig4/AS:281996745035781@1444244671481/Vanilloid-compounds-elicit-increased-Ca-2-i-and-NO-release-from-isolated-rat-R-and_Q320.jpg)
P ainful sensations induced by capsaicin, the pungent substance in hot peppers, are caused by stimulation of vanilloid receptor 1 (VR1), an ion channel protein expressed by nocicep-tive primary afferent neurons. VR1 also participates in the detection of at least two additional noxious stimuli, acid (pH 6) and heat (43°C). The urinary bladder is ric...
Mitochondria are widely believed to be the source of reactive oxygen species (ROS) in a number of neurodegenerative disease states. However, conditions associated with neuronal injury are accompanied by other alterations in mitochondrial physiology, including profound changes in the mitochondrial membrane potential DeltaPsi(m). In this study we hav...
Using the mitochondrial membrane potential (DeltaPsi(m))-sensitive fluorescent dyes 5,5',6,6'-tetrachloro-1,1',3,3'-tetraethylbenzimidazolocarbocyanine iodide (JC-1) and tetramethylrhodamine methyl ester (TMRM), we have observed spontaneous changes in the DeltaPsi(m) of cultured forebrain neurons. These fluctuations in DeltaPsi(m) appear to represe...
Ca(2+), Na(+), K(+), and Mg(2+) have a central role in neuronal excitability. The concentration of these cations in the cytoplasm of neurons (generically termed [ion(+)]i) provides a marker of the excitation state of the neurons, and may also illuminate the activity of specific signaling mechanisms that involve Ca(2+)- or Mg(2+)-activated enzymes....
The N-methyl-D-aspartate subtype of glutamate receptor is unusual in that it requires two endogenous agonists for activation. Thus, in addition to glutamate, the amino acid glycine (or possibly D-serine) is an essential co-agonist. This unit presents a radioligand binding protocol that detects ligand activity at the NMDA receptor-associated glycine...
1. Recent observations showed that a mitochondrial Ca2+ increase is necessary for an NMDA receptor stimulus to be toxic to cortical neurones. In an attempt to determine the magnitude of the Ca2+ fluxes involved in this phenomenon, we used carbonylcyanide-p-(trifluoromethoxy)phenylhydrazone (FCCP), a mitochondrial proton gradient uncoupler, to relea...
MitoTracker dyes are fluorescent mitochondrial markers that covalently bind free sulfhydryls. The impact of alterations in mitochondrial membrane potential (Delta Psi(m)) and oxidant stress on MitoTracker staining in mitochondria in cultured neurons and astrocytes has been investigated. p-(Trifluoromethoxy) phenyl-hydrazone (FCCP) significantly dec...
Interest in mitochondria has experienced a “re-awakening” in recent years as new discoveries of latent mitochondrial properties, expanding beyond their traditional function in cellular bioenergetics, have fostered a closer examination of the mitochondria’s role in health and disease. This growing interest is especially noted in the neurosciences wh...
The membrane-permeant oxidizing agent 2,2'-dithiodipyridine (DTDP) can induce Zn(2+) release from metalloproteins in cell-free systems. Here, we report that brief exposure to DTDP triggers apoptotic cell death in cultured neurons, detected by the presence of both DNA laddering and asymmetric chromatin formation. Neuronal death was blocked by increa...
The membrane-permeant oxidizing agent 2,2′-dithiodipyridine (DTDP) can induce Zn2+ release from metalloproteins in cell-free systems. Here, we report that brief exposure to DTDP triggers apoptotic cell death in cultured neurons, detected by the presence of both DNA laddering and asymmetric chromatin formation. Neuronal death was blocked by increase...
Mitochondria buffer large changes in [Ca(2+)](i)following an excitotoxic glutamate stimulus. Mitochondrial sequestration of [Ca(2+)](i)can beneficially stimulate oxidative metabolism and ATP production. However, Ca(2+)overload may have deleterious effects on mitochondrial function and cell survival, particularly Ca(2+)-dependent production of react...
Increased intracellular free Zn(2+) ([Zn(2+)](i)) is toxic to neurons. Glia are more resistant to Zn(2+)-mediated toxicity; however, it is not known if this is because glia are less permeable to Zn(2+) or if glia possess intrinsic mechanisms that serve to buffer or extrude excess [Zn(2+)](i). We used the Zn(2+)-selective ionophore pyrithione to dir...
In central neurons, glutamate receptor activation causes massive calcium influx and induces a mitochondrial depolarization, which is partially blocked by cyclosporin A, suggesting a possible activation of the mitochondrial permeability transition pore (PTP) as a mechanism. It has been recently reported that tamoxifen (an antiestrogen chemotherapeut...
Oxidative stress can trigger neuronal cell death and has been implicated in several chronic neurological diseases and in acute neurological injury. Oxidative toxicity can be induced by glutamate treatment in cells that lack ionotrophic glutamate receptors, such as the immortalized HT22 hippocampal cell line and immature primary cortical neurons. Pr...
Glutamate can stimulate increases in intracellular magnesium concentration ([Mg2+]i) and induce neurotoxicity, both independent of Ca2+ changes. Although Mg2+ is essential within the cell, very little is known about how it is regulated, especially in neurons. Therefore we used the fluorescent indicator, magindo-1 and confocal microscopy to examine...
We examined several factors related to the increase in susceptibility to excitotoxicity that occurs in embryonic forebrain neurons over time in culture. Neuronal cultures were resistant to a 5-min exposure to 100 microM glutamate/10 microM glycine at 5 days in vitro (DIV), but became vulnerable to the same stimulus by 14 DIV. We used the fluorescen...
This project is designed to investigate intracellular signaling mechanisms associated with neuronal cell injury. In the acute form, this injury accounts for neural injury following stroke and head trauma, while in the chronic phenotype, it may account for degenerative diseases such as Parkinson's disease. Our preliminary studies have suggested that...
A series of novel bisbenzamidines and bisbenzimidazolines with different linkers connecting the aromatic groups was tested in vitro for NMDA receptor antagonist activity. IC50 values for these compounds ranged from 1.2 to >200 microM. The bisbenzamidine with a homopiperazine ring as the central linker was found to be the most potent NMDA receptor a...
The assembly of heterogeneous populations of native N-methyl-D-aspartate receptors results in receptors with multiple pharmacological properties dependent on subunit combinations. Using stably transfected ML(tk-) mouse fibroblasts expressing N-methyl-D-aspartate R1a and either R2A or R2B, we evaluated polyamine effects on [125I]dizocilpine (MK-801)...
We measured glutamate-stimulated increases in intracellular free Ca2+ concentrations ([Ca2+]i) in cultured rat forebrain neurons loaded with both a high- and a low-affinity Ca2+ indicator. In these dual-dye studies, the high-affinity indicators Fluo-3 and Fura-2 gave both qualitatively and quantitatively different results than the low-affinity indi...
Acute neuronal injury caused by activation of glutamate receptors in neurons, or excitotoxicity, can be triggered by the activation of N-methyl-D-aspartate receptors and the entry of large amounts of Ca2+. Recent studies have suggested that mitochondria have a critical role in the excitotoxicity injury mechanism. Mitochondria accumulate large amoun...
The quantification of drug interaction with NMDA receptors has been greatly facilitated by the use of ligand binding assays.
The first assays for this receptor measured NMDA-sensitive [3H]glutamate binding (see
1 for review). However, [3H]glutamate has relatively low affinity for the receptor, and is also a substrate for transport processes in brai...
Both glutamate and reactive oxygen species have been implicated in excitotoxic neuronal injury, and mitochondria may play a key role in the mediation of this process. In this study, we examined whether glutamate-receptor stimulation and oxidative stress interact to affect the mitochondrial membrane potential (delta psi). We measured delta psi in ra...
High concentrations of Zn2+ are found in presynaptic terminals of excitatory neurons in the CNS. Zn2+ can be released during synaptic activity and modulate postsynaptic receptors, but little is known about the possibility that Zn2+ may enter postsynaptic cells and produce dynamic changes in the intracellular Zn2+ concentration ([Zn2+]i). We used fu...
Aromatic analogs of arcaine were shown to have inhibitory effects on the binding of the channel blocking drug [3H]MK-801 to the NMDA receptor complex. The most potent compound of the series was an N,N'-bis(propyl)guanidinium which inhibited [3H]MK-801 binding with an IC50 of 0.58 microM and an IC50 of 12.17 microM upon addition of 100 microM spermi...
Prior studies identified several forms of toxicity that could be associated with the activation of different subtypes of ionic glutamate receptors. The most rapidly induced form of injury occurred as the result of N-methyl D-aspartate (NMDA) receptor activation. NMDA mediated injury could be induced with agonist exposure lasting as little as five m...
We have investigated the role of mitochondrial calcium buffering in excitotoxic cell death. Glutamate acts at NMDA receptors in cultured rat forebrain neurons to increase the intracellular free calcium concentration. Although concurrent inhibition of mitochondrial calcium uptake substantially enhanced this cytoplasmic calcium increase, it significa...
Increasing extracellular pH from 7.4 to 8.5 caused a dramatic increase in the time required to recover from a glutamate (3 microM, for 15 s)-induced increase in intracellular Ca2+ concentration ([Ca2+]i) in indo-1-loaded cultured cortical neurons. Recovery time in pH 7.4 HEPES-buffered saline solution (HBSS) was 126 +/- 30 s, whereas recovery time...
Several studies have suggested that polyamines modulate the interaction of glycine with the NMDA receptor. We have further investigated the effects of polyamines using the NMDA receptor glycine site antagonist [(E)-3-(2-phenyl-2-carboxyethenyl)-4,6-dichloro-1H-indole-2-carbox ylic acid] ([3H]MDL 105,519). [3H]MDL 105,519 binding assays were perform...
We have examined the mechanisms by which cultured central neurones from embryonic rat brain buffer intracellular Ca ²⁺ loads following kainate receptor activation using fluorescent indicators of [Ca ²⁺ ] i and [Na ⁺ ] i .
Stimulation of cultured forebrain neurones with 100 μ m kainate produced a rapid increase in [Ca ²⁺ ] i that displayed a variabl...
![Fig. 2. Effect of KB-R7943 on recovery from a glutamate-induced [Ca 2 ]...](profile/Elias-Aizenman/publication/13725922/figure/fig2/AS:349436346683397@1460323525185/Effect-of-KB-R7943-on-recovery-from-a-glutamate-induced-Ca-2-i-increase-as-a-measure_Q320.jpg)
![Fig. 3. KB-R7943 inhibits acute [Ca 2 ] i increases caused by NMDA and...](profile/Elias-Aizenman/publication/13725922/figure/fig3/AS:349436346683398@1460323525211/KB-R7943-inhibits-acute-Ca-2-i-increases-caused-by-NMDA-and-non-NMDA-receptor_Q320.jpg)
![Fig. 5. KB-R7943 potentiates KA-induced [Ca 2 ] i increases in neurons...](profile/Elias-Aizenman/publication/13725922/figure/fig6/AS:668674805489671@1536435905330/KB-R7943-potentiates-KA-induced-Ca-2-i-increases-in-neurons-depleted-of-intracellular_Q320.jpg)
![Fig. 6. Effect of KB-R7943 on increases in [Ca 2 ] i caused by acute...](profile/Elias-Aizenman/publication/13725922/figure/fig5/AS:349436346683401@1460323525285/Effect-of-KB-R7943-on-increases-in-Ca-2-i-caused-by-acute-15-sec-or-prolonged-5_Q320.jpg)
Activation of ionotropic glutamate receptors causes increases in intracellular Ca2+ concentration ([Ca2+]i) and intracellular Na+ concentration in neurons. It has been suggested that reversal of the plasma membrane Na+/Ca2+ exchanger (NCE) may account in part for the rise in [Ca2+]i. Recently, KB-R7943 (2-[2-[4-(4-nitrobenzyloxy)phenyl]ethyl]isothi...
A series of gamma-amino alcohols were synthesized and screened for reuptake inhibition and noncompetitive NMDA antagonism. Compound (+/-)-3f simultaneously and potently inhibits reuptake of 5-HT, NE, and DA, representing a potential wide-spectrum reuptake inhibitor antidepressant. In addition, comparative rat and human studies uncovered a species-s...
Citations
... Zn 2+ and Ca 2+ fluxes are critical to mitochondrial function [42,43], and human platelets typically contain several mitochondria [39]. The possibility that platelet mitochondria contain detectable concentrations of free Zn 2+ was examined by imaging ND platelets that were simultaneously treated with FluoZn-3-AM and Mitotracker-DR. ...
![[object Object]](https://i1.rgstatic.net/ii/profile.image/276914599153675-1443032993460_Q64/Elias-Aizenman.jpg)
... The increased expression of iNOS suggests an increased level of NO, which could induce the release of Cd from MT, increasing the levels of free Cd and therefore, the direct effects of Cd toxicity. Additionally, MT plays an important role in determining neuronal fate by modulating the localization and the level of intracellular free Zinc (Aizenman et al. 2020). Thus, the increased Zn level observed in the CasCd group may also be related to the increased MT3 expression in the same group. ...
... This movement of electrons generates an alkaline matrix and an acidic intermembrane space. Protons then flow through complex V (ATP synthase), which utilizes the energy to synthesize ATP from ADP [20]. Since molecular oxygen is the terminal electron acceptor in the mitochondrial electron transport chain, assessments of mitochondrial function are often performed through the measurement of oxygen consumption in isolated mitochondria . ...
... The zinc-thiolate groups within MT are crucial targets for NO, directly influencing intracellular zinc homeostasis [58]. It has been reported that NO does not react with thiols alone to produce nitrosothiol [59]. Instead, in the presence of O 2 , NO is converted into N 2 O 3 and/or NO 2 , which can then nitrosylate MT, leading to zinc release by disrupting zinc-sulfur clusters [59,60]. ...
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... Acetylcholine was purchased from Sigma-Aldrich Incorporated (St. Louis, MO). cis-2-[[(3,5-Dichlorophenyl)amino]carbonyl]cyclohexanecarboxylic acid (VU0155041), N-(4-(N-(2-chlorophenyl)sulfamoyl)phenyl)picolinamide (4PAM2), and 5-methyl-N-(4methylpyrimidin-2-yl)-4-(1H-pyrazol-4-yl)thiazol-2-amine (ADX-88178) were synthesized in-house according to methods in (Niswender et al., 2008;Reynolds, 2008;Engers et al., 2010;Celanire et al., 2011). ...
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... The decrease in proliferation rate of LHON fibroblasts in DMEM-Gal does not appear to result from increases in cell death. Nuclear condensation and fragmentation during apoptosis is readily detected by Hoechst staining [57][58][59]. Very few apoptotic nuclei were observed in Hoechst-stained fields of either cell line, far too few to account for the changes in cell number observed daily. Likewise, the number of detached cells observed on successive days remained very low for all conditions. ...
... Due to its large lipid content, relatively high oxygen consumption, and low levels of antioxidant defenses, the brain is particularly susceptible to oxidative stress [31][32][33][34][35][36]. The mitochondrial electron transport chain (ETC) complexes I and III, as well as the TCA cycles of a-ketoglutarate dehydrogenase, are believed to be the sources of superoxide radicals [37,38]. ...
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... injection with L-DOPA + Bz to trigger the dyskinesia. The doses selected were those shown previously to be effective at inducing behavioral responses in rats (e.g., [13,16,26]). The pre-treatment time was selected following pharmacokinetic studies which showed t max for both drugs was 1 h (Supplementary Figure 1A, B), thereby ensuring maximal plasma concentrations of the test drug and L-DOPA coincided. ...
... Unilateral 6-OHDA injection produces deficits in balance, motor coordination and rotarod performance as reported by several studies (Lundblad et al., 2003a;Monville et al., 2006;Smith et al., 2014). In agreement with previous reports, our results show a decrease in motor function and coordination in animals after 6-OHDA lesioning. ...
... Organotypic cell culture models are often used to determine biochemical and physiological pathways at a cellular level. Importantly they retain the cytoarchitecture of adult tissue and therefore largely replicate in vivo environments [8,10,20]. Organotypic culture of the ventral mesencephalon (VM) and striatal tissue has been particularly useful for mechanistic and dopaminergic cell survival studies as interconnections between the striatum and VM are retained. Due to the controlled nature of the culture environment, determination of factors that result in dopaminergic cell death can be easily studied with use of various receptor agonists and antagonists. ...
