Modulation of N-methyl-d-aspartate receptors by hydroxyl radicals in rat cortical neurons in vitro

doi:10.1016/0304-3940(95)11442-Y

Neuroscience Letters

Volume 189, Issue 1, 7 April 1995, Pages 57-59

https://doi.org/10.1016/0304-3940(95)11442-Y Get rights and content

Abstract

Oxygen-derived reactive species generated by xanthine/xanthine oxidase (X/XO) can modulate the N-methyl-d-aspartate (NMDA) receptor via its redox-sensitive site. Here it is shown that hydroxyl radicals are the agents responsible for NMDA receptor oxidation by X/XO. Spectrophotometric assays revealed that the amounts of superoxide anion and H₂O₂ produced by X/XO were not decreased by the hydroxyl radical-specific scavenger mannitol. This sugar, however, could prevent most of the oxidizing actions of NMDA receptors by X/XO, but not by the thiol oxidizing agent 5,5′-dithio-bis-nitrobenzoic acid. Finally, a non-enzymatic source of hydroxyl radicals was also effective in oxidizing the receptor's redox site. Hydroxyl radicals may thus represent the final common pathway for the modulation of NMDA receptor function by oxygen-derived free radical generating systems.

References (16)

E. Aizenman et al.
Oxygen free radicals regulate NMDA receptor function via a redox modulatory site
Neuron
(1990)
E. Aizenman et al.
Selective modulation of NMDA responses by reduction and oxidation
Neuron
(1989)
J.M. Braughler et al.
Central nervous system trauma and stroke. I. Biochemical considerations for oxygen radical formation and lipid peroxidation
Free Radical Biol. Med.
(1989)
B.E. Britigan et al.
Hydroxyl radical is not a product of the reaction of xanthine oxidase and xanthine
J. Biol. Chem.
(1990)
M. Culcasi et al.
Glutamate receptors induce a burst of superoxide via activation of nitric oxide synthase in arginine-depleted neurons
J. Biol. Chem.
(1994)
J.W. Heinecke et al.
Dityrosine, a specific marker of oxidation, is synthesized by the myeloperoxidase-hydrogen peroxide system of human neutrophils and macrophages
J. Biol. Chem.
(1993)
G. Köhr et al.
NMDA receptor channels: subunit-specific potentiation by reducing agents
Neuron
(1994)
M. Pernollet et al.
OH treatment of tetanus toxin reduces its susceptibility to limited proteolysis with more efficient presentation to specific T cells
Mol. Immunol.
(1993)

There are more references available in the full text version of this article.

Cited by (48)

Glutaminergic tonic action potentiate MPP <sup>+</sup> -induced hydroxyl radical production in rat striatum
2019, Neuroscience Letters
Citation Excerpt :
First is the NMDA receptor mediated oxidation of MPP+ which has been described as a source of OH [1,9,16]. Oxygen derived free radical can modulates NMDA receptor [18]. It is known that the OH generation was virtually abolished by NMDA antagonist [19].
The effect of glycine on 1-methyl-4-phenylpyridinium ion (MPP⁺)-induced hydroxyl radical (OH) formation in the extracellular fluid of rat striatum were investigated. Rats were anesthetized and sodium salicylate in Ringer's solution (0.5 nmol/μl/min) was infused through a microdialysis probe to detect the generation of OH as reflected by the non-enzymatic formation of 2,3-dihydroxybenzoic acid (2,3-DHBA) in rat striatum. MPP⁺ (5 mmol/L) produced an increase in OH formation. When glycine (1 mmol/L) was infused into the rat striatum through a microdialysis probe after MPP⁺ treatment, the marked in the level of 2,3-DHBA was observed in the brain dialysate. However, in the presence of MK-801 (100 μmol/L), a non competitive antagonist of N-methyl-D-aspartate (NMDA), glycine failed to increase the 2,3-DHBA formation by MPP⁺. When corresponding experiments were performed with nitro-L arginine (L-NNA) (1 mmol/L), a nitric oxide synthase (NOS) inhibitor, same result was obtained. These results suggest that MPP⁺-induced OH generation may modulated by glycine via NMDA receptor in rat striatum. This increase might be explained because of the presence of a glutaminergic tonic action
Cellular calcium signaling in the aging brain
2019, Journal of Chemical Neuroanatomy
Citation Excerpt :
Oxidation and reduction of sulfhydryl moieties alter receptor function. An oxidative environment can decrease NMDAR function in neuronal cell cultures (Aizenman et al., 1989; Aizenman, 1995; Sucher and Lipton, 1991) owing to the formation of disulfide bonds on the sulfhydryl group-containing amino acid residues in the receptor (Aizenman et al., 1990; Choi et al., 2001; Sullivan et al., 1994). On the other hand, NMDAR can also mediate excitotoxic cell death.
Aging in the biological system is an irreversible process. In the initial stages of lifespan aging improves survival skills of an organism while in the later stages aging reduce the survival skills. Aging is associated with changes in several cellular and molecular functions among which calcium signaling is a prominent one. Calcium signaling is essential for many vital functions of the brain and even minor impairments in calcium signaling can lead to deleterious consequences including neuronal death. Calcium signaling proteins are pursued as promising drug targets for many aging related diseases. This review attempts to summarize changes in calcium signaling in the brain as a result of aging.
Age-related NMDA signaling alterations in SOD2 deficient mice
2018, Biochimica et Biophysica Acta - Molecular Basis of Disease
Oxidative stress affects the survival and function of neurons. Hence, they have a complex and highly regulated machinery to handle oxidative changes. The dysregulation of this antioxidant machinery is associated with a wide range of neurodegenerative conditions. Therefore, we evaluated signaling alterations, synaptic properties and behavioral performance in 2 and 6-month-old heterozygous manganese superoxide dismutase knockout mice (SOD2^+/− mice). We found that their low antioxidant capacity generated direct oxidative damage in proteins, lipids, and DNA. However, only 6-month-old heterozygous knockout mice presented behavioral impairments. On the other hand, synaptic plasticity, synaptic strength and NMDA receptor (NMDAR) dependent postsynaptic potentials were decreased in an age-dependent manner. We also analyzed the phosphorylation state of the NMDAR subunit GluN2B. We found that while the levels of GluN2B phosphorylated on tyrosine 1472 (synaptic form) remain unchanged, we detected increased levels of GluN2B phosphorylated on tyrosine 1336 (extrasynaptic form), establishing alterations in the synaptic/extrasynaptic ratio of GluN2B. Additionally, we found increased levels of two phosphatases associated with dephosphorylation of p-1472: striatal-enriched protein tyrosine phosphatase (STEP) and phosphatase and tensin homolog deleted on chromosome Ten (PTEN). Moreover, we found decreased levels of p-CREB, a master transcription factor activated by synaptic stimulation. In summary, we describe mechanisms by which glutamatergic synapses are altered under oxidative stress conditions. Our results uncovered new putative therapeutic targets for conditions where NMDAR downstream signaling is altered. This work also contributes to our understanding of processes such as synapse formation, learning, and memory in neuropathological conditions.
Central role for NMDA receptors in redox mediated impairment of synaptic function during aging and Alzheimer's disease
2017, Behavioural Brain Research
Citation Excerpt :
The introduction of oxidizing agents, including oxidized glutathione and xanthine/xanthine oxidase, reduces the NMDA receptor synaptic response in hippocampal slices from young relative to older animals, while bath application of the reducing agent dithiothreitol (DTT) enhances NMDA receptor synaptic responses specifically in older cognitively impaired animals [71–73,76]. The NMDA receptor contains three pairs of extracellular cysteine residues, and with increased oxidative stress, the cysteine residues may interact through the redox-mediated formation of disulfide bonds (Fig. 1), altering receptor confirmation to decrease receptor function [78–82]. While this extracellular modification presents an attractive model of NMDA receptor impairment, our work indicates a primarily intracellular mechanism.
Increased human longevity has magnified the negative impact that aging can have on cognitive integrity of older individuals experiencing some decline in cognitive function. Approximately 30% of the elderly will have cognitive problems that influence their independence. Impaired executive function and memory performance are observed in normal aging and yet can be an early sign of a progressive cognitive impairment of Alzheimer’s disease (AD), the most common form of dementia. Brain regions that are vulnerable to aging exhibit the earliest pathology of AD. Senescent synaptic function is observed as a shift in Ca²⁺-dependent synaptic plasticity and similar mechanisms are thought to contribute to the early cognitive deficits associated with AD. In the case of aging, intracellular redox state mediates a shift in Ca²⁺ regulation including N-methyl-d-aspartate (NMDA) receptor hypofunction and increased Ca²⁺ release from intracellular stores to alter synaptic plasticity. AD can interact with these aging processes such that molecules linked to AD, β-amyloid (Aβ) and mutated presenilin 1 (PS1), can also degrade NMDA receptor function, promote Ca²⁺ release from intracellular stores, and may increase oxidative stress. Thus, age is one of the most important predictors of AD and brain aging likely contributes to the onset of AD. The focus of this review article is to provide an update on mechanisms that contribute to the senescent synapse and possible interactions with AD-related molecules, with special emphasis on regulation of NMDA receptors.
Photodamage of lipid bilayers by irradiation of a fluorescently labeled cell-penetrating peptide
2014, Biochimica et Biophysica Acta - General Subjects
Citation Excerpt :
However, a concern is that the hydroxyl radical, a ROS that can potentially be produced from superoxide, might interfere with the NBT assay (tiron can also potentially quench the hydroxyl radical [36]). To address this issue, mannitol, a quencher with specificity for the hydroxyl radical over other ROS, was added to the NBT assay [37,38]. As shown in Fig. 2D, addition of mannitol had a relatively small effect on the chemical reduction of NBT mediated by TMR-irradiation.
Fluorescently labeled cell-penetrating peptides can translocate into cells by endocytosis and upon light irradiation, lyse the endocytic vesicles. This photo-inducible endosomolytic activity of Fl–CPPs can be used to efficiently deliver macromolecules such as proteins and nucleic acids and other small organic molecules into the cytosol of live cells. The requirement of a light trigger to induce photolysis provides a more spatial and temporal control to the intracellular delivery process.
In this report, we examine the molecular level mechanisms by which cell-penetrating peptides such as TAT when labeled with small organic fluorophore molecules acquire a photo-induced lytic activity using a simplified model of lipid vesicles.
The peptide TAT labeled with 5(6)-carboxytetramethylrhodamine binds to negatively charged phospholipids, thereby bringing the fluorophore in close proximity to the membrane of liposomes. Upon light irradiation, the excited fluorophore produces reactive oxygen species at the lipid bilayer and oxidation of the membrane is achieved. In addition, the fluorescent peptide causes aggregation of photo-oxidized lipids, an activity that requires the presence of arginine residues in the peptide sequence.
These results suggest that the cell-penetrating peptide plays a dual role. On one hand, TAT targets a conjugated fluorophore to membranes. On the other hand, TAT participates directly in the destabilization of photosensitized membranes. Peptide and fluorophore therefore appear to act in synergy to destroy membranes efficiently.
Understanding the mechanism behind Fl–CPP mediated membrane photodamage will help to design optimally photo-endosomolytic compounds.
Pilocapine alters NMDA receptor expression and function in hippocampal neurons: NADPH oxidase and ERK1/2 mechanisms
2011, Neurobiology of Disease
Citation Excerpt :
Both pathways are required for NMDAR upregulation. The cysteine residues on NMDARs are targets for S-nitrosylation (Eu et al., 2000; Stamler et al., 2001) and disulfide bond formation (Choi et al., 2001; Lipton et al., 2002), implicating a critical role for reactive oxygen and nitrogen species in modulating NMDAR function (Aizenman et al., 1989, 1990, Aizenman, 1995). In order to investigate on how PILO-induced oxidative stress modulates NMDAR functional activity, we examined neuronal Ca2+ influx after acute application of low concentrations of glutamate.
The molecular basis for epileptogenesis remains poorly defined, but repeated or prolonged seizures can cause altered hippocampal N-methyl d-aspartate receptor (NMDAR) stoichiometry, loss of hippocampal neurons, and aberrant mossy fiber sprouting. Using the muscarinic receptor 1 (m1R) agonist, pilocarpine (PILO), in hippocampal cell cultures we explored the early sequence of molecular events that occur within 24 h of the initial insult and result in altered neuronal function during epileptogenesis. Our findings show that PILO-induced, m1R-mediated, inositol 1,4,5-trisphosphate (IP3) synthesis constitutes an early, crucial biochemical event required for NMDAR hyperactivation and subsequent NADPH oxidase (NOX) activation and NMDAR-independent ERK1/2 phoshorylation. Together, but not separately, NOX activation and ERK1/2 phosphorylation induce alterations in NMDAR stoichiometry through the upregulation of NR1 and NR2B subunits. Lastly, we demonstrated that PILO-mediated oxidative stress alters NMDAR function through the redox modulation of cysteine residues. The in vitro results related to thiol oxidation, NOX activation, ERK1/2 phosphorylation and NMDAR upregulation were confirmed in vivo, 24 h after treatment of adult rats with PILO. These results obtained in PILO-treated primary hippocampal neurons – and confirmed in vivo at the same time-point after PILO – provide a better understanding of the early cellular responses during epileptogenesis and identify potential therapeutic targets to prevent development of chronic epilepsy.

View all citing articles on Scopus

View full text

Modulation of N-methyl-d-aspartate receptors by hydroxyl radicals in rat cortical neurons in vitro

Abstract

Neuron

Neuron

Free Radical Biol. Med.

J. Biol. Chem.

J. Biol. Chem.

J. Biol. Chem.

Neuron

Mol. Immunol.