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Relationship: 361


A descriptive phrase which clearly defines the two KEs being considered and the sequential relationship between them (i.e., which is upstream, and which is downstream). More help

Overactivation, NMDARs leads to Increased, Intracellular Calcium overload

Upstream event
The causing Key Event (KE) in a Key Event Relationship (KER). More help
Downstream event
The responding Key Event (KE) in a Key Event Relationship (KER). More help

Key Event Relationship Overview

The utility of AOPs for regulatory application is defined, to a large extent, by the confidence and precision with which they facilitate extrapolation of data measured at low levels of biological organisation to predicted outcomes at higher levels of organisation and the extent to which they can link biological effect measurements to their specific causes.Within the AOP framework, the predictive relationships that facilitate extrapolation are represented by the KERs. Consequently, the overall WoE for an AOP is a reflection in part, of the level of confidence in the underlying series of KERs it encompasses. Therefore, describing the KERs in an AOP involves assembling and organising the types of information and evidence that defines the scientific basis for inferring the probable change in, or state of, a downstream KE from the known or measured state of an upstream KE. More help

AOPs Referencing Relationship

AOP Name Adjacency Weight of Evidence Quantitative Understanding Point of Contact Author Status OECD Status
Binding of agonists to ionotropic glutamate receptors in adult brain causes excitotoxicity that mediates neuronal cell death, contributing to learning and memory impairment. adjacent Moderate Allie Always (send email) Open for citation & comment WPHA/WNT Endorsed
Acetylcholinesterase Inhibition Leading to Neurodegeneration adjacent High Moderate Allie Always (send email) Under development: Not open for comment. Do not cite
Calcium overload in dopaminergic neurons of the substantia nigra leading to parkinsonian motor deficits adjacent Not Specified Not Specified Cataia Ives (send email) Under development: Not open for comment. Do not cite
Binding of chemicals to ionotropic glutamate receptors leads to impairment of learning and memory via loss of drebrin from dendritic spines of neurons adjacent Evgeniia Kazymova (send email) Under development: Not open for comment. Do not cite Under Development

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) that help to define the biological applicability domain of the KER.In general, this will be dictated by the more restrictive of the two KEs being linked together by the KER.  More help
Term Scientific Term Evidence Link
zebrafish Danio rerio Low NCBI
rat Rattus norvegicus High NCBI
mouse Mus musculus High NCBI

Sex Applicability

An indication of the the relevant sex for this KER. More help
Sex Evidence
Unspecific High

Life Stage Applicability

An indication of the the relevant life stage(s) for this KER.  More help
Term Evidence
All life stages High

Key Event Relationship Description

Provides a concise overview of the information given below as well as addressing details that aren’t inherent in the description of the KEs themselves. More help

The NMDA receptor is distinct from the other glutamate receptors in two ways: first, it is both ligand-gated and voltage-dependent; second, it requires co-activation by two ligands: glutamate and either glycine or D-serine. Following membrane depolarization, the co-agonists, L-glutamate and glycine must bind to their respective sites on the receptor to open the channel. On activation, the NMDA receptor allows the influx of extracellular calcium ions into the postsynaptic neuron and neurotransmission occurs (reviewed in Higley and Sabatini, 2012). Calcium flux through NMDA receptors is also thought to be critical in synaptic plasticity, a cellular mechanism for learning and memory. Indeed, NMDA receptor–dependent synaptic potentiation (LTP) and depression (LTD) are two forms of activity-dependent long-term changes in synaptic efficacy that are believed to represent cellular correlates of learning and memory processes. The best characterized form of NMDA receptor-dependent LTP and LTD occurs between CA3 and CA1 pyramidal neurons of the hippocampus (Luscher and Malenka, 2012). It is now well established that modest activation of NMDARs leads to modest increases in postsynaptic calcium, triggering LTD, whereas much stronger activation of NMDARs leading to much larger increases in postsynaptic calcium are required to trigger LTP (Luscher and Malenka, 2012). The high-frequency stimulation causes a strong temporal summation of the excitatory postsynaptic potentials, and depolarization of the postsynaptic cell is sufficient to relieve the Mg2+ block of the NMDAR and allow a large amount of calcium to enter into the post-synaptic cells.

Evidence Collection Strategy

Include a description of the approach for identification and assembly of the evidence base for the KER. For evidence identification, include, for example, a description of the sources and dates of information consulted including expert knowledge, databases searched and associated search terms/strings.  Include also a description of study screening criteria and methodology, study quality assessment considerations, the data extraction strategy and links to any repositories/databases of relevant references.Tabular summaries and links to relevant supporting documentation are encouraged, wherever possible. More help

As it pertains to AOP 281, evidence was collected in multiple ways: literature searches of external databases, review of related KEs and KERS in the AOPWiki, and consultation with experts.   Extensive literature searches were conducted in Scopus, Pubmed, and Google Scholar using keywords applicable to each KE, with an initial focus on zebrafish data to then focusing on rat data. Related KEs and KERs in the AOPWiki were also reviewed for relevant evidence and their sources.  The “snowball method” was used to find additional articles, i.e., relevant citations within an article were obtained if they provided additional evidence. EndNote reference managing software was used to store results from the literature searches and when possible, a pdf of the manuscript was attached to each record. Papers were reviewed and categorized by whether they contained data to support one or more parts of the AOP. An Excel spreadsheet was used to record reviewed papers and any information worth noting.

Evidence Supporting this KER

Addresses the scientific evidence supporting KERs in an AOP setting the stage for overall assessment of the AOP. More help
Biological Plausibility
Addresses the biological rationale for a connection between KEupstream and KEdownstream.  This field can also incorporate additional mechanistic details that help inform the relationship between KEs, this is useful when it is not practical/pragmatic to represent these details as separate KEs due to the difficulty or relative infrequency with which it is likely to be measured.   More help

There is structural and functional mechanistic understanding supporting this relationship between NMDAR overactivation and increased intracellular calcium.

The relationship between the upstream and downstream key event is plausible as the expression of the functional NMDA receptors is commonly carried out or assessed by Ca2+ imaging method. Calcium imaging techniques have been extensively utilized in the literature to investigate the potential interactions between NMDA-evoked Ca2+ influx and NMDA receptor activation. Approximately 15% of the current through NMDA receptors is mediated by Ca2+ under physiological conditions (Higley and Sabatini, 2012).

It has been shown that less than five and, occasionally, only a single NMDA receptor opens under physiological conditions, causing a total Ca2+ influx of about 6000 ions into a spine head reaching a concentration of ∼10 µm (Higley and Sabatini, 2012). However, the majority of the ions are rapidly eliminated by binding to Ca2+ proteins, reaching ∼1 µM of free Ca2+ concentration (Higley and Sabatini, 2012).

It has been shown that in rat primary forebrain cultures the intracellular Ca2+ increases after activation of the NMDA receptor through administration of NMDA but this increase in Ca2+ is blocked when the cells are cultured under Ca2+ free conditions, demonstrating that the NMDA-evoked increase in intracellular Ca2+ derives from extracellular and not intracellular sources (Liu et al., 2013).

Indirect mechanism of domoic acid (DA) induced overactivation of NMDARs that result in Ca2+ overload: depolarization of the pre-synaptic cell activates the release of endogenous Ca2+ which mobilizes vesicles containing GLU to the membrane surface. Glutamate (GLU) is then released into the synaptic cleft by exocytosis where it is able to interact with cell surface receptors. Exogenous DA can interact within the synaptic cleft with each of the three ionotropic receptor subtypes including the kainate, AMPA, and NMDA receptors on cell membranes. Activation of the kainate and AMPA receptors results in release of Ca2+ via coupled ion channels, into the post-synaptic cell. DA is also able to bind to NMDA receptors that are linked to both Ca2+ and NA/K+ ion channels and results in a cellular influx of both Na+ and Ca2+. Unlike GLU, DA induces prolonged receptor activation causing a constant influx of cations into the cell and the appropriate chemical cues for desensitization are blocked. The excess intracellular Ca2+ causes disruption of cellular function, cell swelling and ultimately cell death (Lefebvre and Robertson,2010).

Glufosinate (GLF) is the methylphosphinate analog of glutamate that directly can activate NMDARs (Lantz et al., 2014, Matsumura et al., 2001, Faro et al., 2013) (as described in KE: NMDARs, Binding of agonist). It is well established in the existing literature that activation of NMDARs leads to the intra-cellular Ca2+ overload and based on this assumption it can be suggested that an exposure to GLF leads to increased intra-cellular calcium levels.

Uncertainties and Inconsistencies
Addresses inconsistencies or uncertainties in the relationship including the identification of experimental details that may explain apparent deviations from the expected patterns of concordance. More help

A case of a 59-yr-old woman who ingested a herbicide containing glufosinate was suffering from severe intoxication, however, she did not develop convulsions, which experimentally occurs in rats treated with GLF (Koyama et al., 1994) and is described in other human cases (Watanabe and Sano 1998).

Known modulating factors

This table captures specific information on the MF, its properties, how it affects the KER and respective references.1.) What is the modulating factor? Name the factor for which solid evidence exists that it influences this KER. Examples: age, sex, genotype, diet 2.) Details of this modulating factor. Specify which features of this MF are relevant for this KER. Examples: a specific age range or a specific biological age (defined by...); a specific gene mutation or variant, a specific nutrient (deficit or surplus); a sex-specific homone; a certain threshold value (e.g. serum levels of a chemical above...) 3.) Description of how this modulating factor affects this KER. Describe the provable modification of the KER (also quantitatively, if known). Examples: increase or decrease of the magnitude of effect (by a factor of...); change of the time-course of the effect (onset delay by...); alteration of the probability of the effect; increase or decrease of the sensitivity of the downstream effect (by a factor of...) 4.) Provision of supporting scientific evidence for an effect of this MF on this KER. Give a list of references.  More help
Response-response Relationship
Provides sources of data that define the response-response relationships between the KEs.  More help
Information regarding the approximate time-scale of the changes in KEdownstream relative to changes in KEupstream (i.e., do effects on KEdownstream lag those on KEupstream by seconds, minutes, hours, or days?). More help
Known Feedforward/Feedback loops influencing this KER
Define whether there are known positive or negative feedback mechanisms involved and what is understood about their time-course and homeostatic limits. More help

Domain of Applicability

A free-text section of the KER description that the developers can use to explain their rationale for the taxonomic, life stage, or sex applicability structured terms. More help

NMDARs have been shown to regulate calcium ion flow in a variety of species including zebrafish and rats (Horzmann and Freeman, 2016, el Nasr et al., 1990).


List of the literature that was cited for this KER description. More help

Berman FW, LePage KT, Murray TF., Domoic acid neurotoxicity in cultured cerebellar granule neurons is controlled preferentially by the NMDA receptor Ca(2+) influx pathway. Brain Res., 2002, 924: 20-29.

el Nasr, M. S., Peruche, B., Roβberg, C., Mennel, H.-D. & Krieglstein, J. 1990. Neuroprotective effect of memantine demonstrated in vivo and in vitro. European Journal of Pharmacology, 185, 19-24. DOI:

Faro LR, Ferreira Nunes BV, Alfonso M, Ferreira VM, Durán R., Role of glutamate receptors and nitric oxide on the effects of glufosinate ammonium, an organophosphate pesticide, on in vivo dopamine release in rat striatum. Toxicology., 2013, Sep 15, 311: 154-61.

Giordano G, White CC, McConnachie LA, Fernandez C, Kavanagh TJ, Costa LG., Neurotoxicity of domoic Acid in cerebellar granule neurons in a genetic model of glutathione deficiency. Mol Pharmacol. 2006., 70: 2116-2126.

Giordano G, White CC, Mohar I, Kavanagh TJ, Costa LG., Glutathione levels modulate domoic acid-induced apoptosis in mouse cerebellar granule cells. Toxicol Sci., 2007, 100: 433-444.

Higley MJ, Sabatini BL., Calcium signalling in dendritic spines. Cold Spring Harb Perspect Biol., 2012, 4: a005686.

Horzmann, K. A. & Freeman, J. L. 2016. Zebrafish get connected: investigating neurotransmission targets and alterations in chemical toxicity. Toxics, 4.

Hu, E., Mergenthal, A., Bingham, C. S., Song, D., Bouteiller, J. M. & Berger, T. W. 2018. A Glutamatergic Spine Model to Enable Multi-Scale Modeling of Nonlinear Calcium Dynamics. Front Comput Neurosci, 12, 58. DOI: 10.3389/fncom.2018.00058.

Hyrc, K., Handran, S. D., Rothman, S. M. & Goldberg, M. P. 1997. Ionized intracellular calcium concentration predicts excitotoxic neuronal death: observations with low-affinity fluorescent calcium indicators. J Neurosci, 17, 6669-77. DOI: 10.1523/jneurosci.17-17-06669.1997.

Koyama K, Andou Y, Saruki K, Matsuo H., Delayed and severe toxicities of a herbicide containing glufosinate and a surfactant. Vet Hum Toxicol., 1994, 36: 17-8.

Lantz Stephen R , Cina M. Mack , Kathleen Wallace, Ellen F. Key , Timothy J. Shafer , John E. Casida., Glufosinate binds N-methyl-D aspartate receptors and increases neuronal network activity in vitro. NeuroToxicology, 2014, 45: 38–47.

Lefebvre KA, Robertson A. Domoic acid and human exposure risks: a review. Toxicon. 2010 Aug 15;56(2):218-30.

Liu F, Patterson TA, Sadovova N, Zhang X, Liu S, Zou X, Hanig JP, Paule MG, Slikker W Jr, Wang C., Ketamine-induced neuronal damage and altered N-methyl-D-aspartate receptor function in rat primary forebrain culture. Toxicol Sci., 2013, 131: 548-557.

Luscher C. and Robert C. Malenka. NMDA Receptor-Dependent Long-Term Potentiation and Long-Term Depression (LTP/LTD). Cold Spring Harb Perspect Biol., 2012;4:a005710.

Matsumura N, Takeuchi C, Hishikawa K, Fujii T, Nakaki T., Glufosinate ammonium induces convulsion through N-methyl-D-aspartate receptors in mice. Neurosci Lett., 2001, 304(1-2): 123-5.

Michaels, R. L. and Rothman, S. M. 1990. Glutamate neurotoxicity in vitro: antagonist pharmacology and intracellular calcium concentrations. J Neurosci, 10, 283-92. DOI: 10.1523/jneurosci.10-01-00283.1990.

Palygin, O., Lalo, U. & Pankratov, Y. 2011. Distinct pharmacological and functional properties of NMDA receptors in mouse cortical astrocytes. British Journal of Pharmacology, 163, 1755-1766. DOI: 10.1111/j.1476-5381.2011.01374.x.

Qiu S, Currás-Collazo MC., Histopathological and molecular changes produced by hippocampal microinjection of domoic acid. Neurotoxicol Teratol., 2006, 28: 354-362.

Watanabe T, Sano T., Neurological effects of glufosinate poisoning with a brief review. Hum Exp Toxicol. 1998, 17: 35-9.