Aop: 12

Title

Each AOP should be given a descriptive title that takes the form “MIE leading to AO”. For example, “Aromatase inhibition [MIE] leading to reproductive dysfunction [AO]” or “Thyroperoxidase inhibition [MIE] leading to decreased cognitive function [AO]”. In cases where the MIE is unknown or undefined, the earliest known KE in the chain (i.e., furthest upstream) should be used in lieu of the MIE and it should be made clear that the stated event is a KE and not the MIE. More help

Chronic binding of antagonist to N-methyl-D-aspartate receptors (NMDARs) during brain development leads to neurodegeneration with impairment in learning and memory in aging

Short name
A short name should also be provided that succinctly summarises the information from the title. This name should not exceed 90 characters. More help
Binding of antagonist to NMDARs can lead to neuroinflammation and neurodegeneration

Graphical Representation

A graphical summary of the AOP listing all the KEs in sequence, including the MIE (if known) and AO, and the pair-wise relationships (links or KERs) between those KEs should be provided. This is easily achieved using the standard box and arrow AOP diagram (see this page for example). The graphical summary is prepared and uploaded by the user (templates are available) and is often included as part of the proposal when AOP development projects are submitted to the OECD AOP Development Workplan. The graphical representation or AOP diagram provides a useful and concise overview of the KEs that are included in the AOP, and the sequence in which they are linked together. This can aid both the process of development, as well as review and use of the AOP (for more information please see page 19 of the Users' Handbook).If you already have a graphical representation of your AOP in electronic format, simple save it in a standard image format (e.g. jpeg, png) then click ‘Choose File’ under the “Graphical Representation” heading, which is part of the Summary of the AOP section, to select the file that you have just edited. Files must be in jpeg, jpg, gif, png, or bmp format. Click ‘Upload’ to upload the file. You should see the AOP page with the image displayed under the “Graphical Representation” heading. To remove a graphical representation file, click 'Remove' and then click 'OK.'  Your graphic should no longer be displayed on the AOP page. If you do not have a graphical representation of your AOP in electronic format, a template is available to assist you.  Under “Summary of the AOP”, under the “Graphical Representation” heading click on the link “Click to download template for graphical representation.” A Powerpoint template file should download via the default download mechanism for your browser. Click to open this file; it contains a Powerpoint template for an AOP diagram and instructions for editing and saving the diagram. Be sure to save the diagram as jpeg, jpg, gif, png, or bmp format. Once the diagram is edited to its final state, upload the image file as described above. More help

Authors

List the name and affiliation information of the individual(s)/organisation(s) that created/developed the AOP. In the context of the OECD AOP Development Workplan, this would typically be the individuals and organisation that submitted an AOP development proposal to the EAGMST. Significant contributors to the AOP should also be listed. A corresponding author with contact information may be provided here. This author does not need an account on the AOP-KB and can be distinct from the point of contact below. The list of authors will be included in any snapshot made from an AOP. More help

Florianne Tschudi-Monnet, Department of Physiology, University of Lausanne, Switzerland, and Swiss Center for Applied Human Toxicology (SCAHT), Florianne.Tschudi-Monnet@unil.ch, corresponding author

Rex FitzGerald, SCAHT, Universität Basel, Missionsstrasse 64, CH-4055 Basel, Rex.FitzGerald@unibas.ch

Acknowledgments: The authors greatly acknowledged the contribution of Drs Anna Price and Magda Sachana who prepared the MIE and KE1-KE4 as well as the related KERs of this AOP.

Anna Price, Joint Research Centre Institute for Health and Consumer Protection Systems Toxicology Unit Via E. Fermi 2749 - 21020 - Ispra (VA) -Italy, e-mail address: PRICE Anna <Anna.PRICE@ec.europa.eu>

Magdalini Sachana, Joint Research Centre Institute for Health and Consumer Protection Systems Toxicology Unit Via E. Fermi 2749 - 21020 - Ispra (VA) -Italy, present e-mail address: "Magdalini.SACHANA@oecd.org" <Magdalini.SACHANA@oecd.org>

Point of Contact

Indicate the point of contact for the AOP-KB entry itself. This person is responsible for managing the AOP entry in the AOP-KB and controls write access to the page by defining the contributors as described below. Clicking on the name will allow any wiki user to correspond with the point of contact via the email address associated with their user profile in the AOP-KB. This person can be the same as the corresponding author listed in the authors section but isn’t required to be. In cases where the individuals are different, the corresponding author would be the appropriate person to contact for scientific issues whereas the point of contact would be the appropriate person to contact about technical issues with the AOP-KB entry itself. Corresponding authors and the point of contact are encouraged to monitor comments on their AOPs and develop or coordinate responses as appropriate.  More help
Florianne Tschudi-Monnet   (email point of contact)

Contributors

List user names of all  authors contributing to or revising pages in the AOP-KB that are linked to the AOP description. This information is mainly used to control write access to the AOP page and is controlled by the Point of Contact.  More help
  • Florianne Tschudi-Monnet

Status

The status section is used to provide AOP-KB users with information concerning how actively the AOP page is being developed, what type of use or input the authors feel comfortable with given the current level of development, and whether it is part of the OECD AOP Development Workplan and has been reviewed and/or endorsed. “Author Status” is an author defined field that is designated by selecting one of several options from a drop-down menu (Table 3). The “Author Status” field should be changed by the point of contact, as appropriate, as AOP development proceeds. See page 22 of the User Handbook for definitions of selection options. More help
Author status OECD status OECD project SAAOP status
Open for citation & comment WPHA/WNT Endorsed 1.13 Included in OECD Work Plan
This AOP was last modified on June 04, 2021 10:20
The date the AOP was last modified is automatically tracked by the AOP-KB. The date modified field can be used to evaluate how actively the page is under development and how recently the version within the AOP-Wiki has been updated compared to any snapshots that were generated. More help

Revision dates for related pages

Page Revision Date/Time
Inhibition, NMDARs September 16, 2017 10:14
N/A, Neurodegeneration February 23, 2021 05:07
Decreased, Calcium influx June 13, 2018 08:26
Binding of antagonist, NMDA receptors June 13, 2018 08:23
Reduced levels of BDNF April 04, 2019 09:21
Cell injury/death September 11, 2020 08:27
Neuroinflammation February 23, 2021 04:56
Impairment, Learning and memory March 16, 2020 09:20
Binding of antagonist, NMDA receptors leads to Inhibition, NMDARs June 13, 2018 08:48
Inhibition, NMDARs leads to Decreased, Calcium influx April 07, 2018 04:32
Decreased, Calcium influx leads to BDNF, Reduced June 13, 2018 09:00
BDNF, Reduced leads to Cell injury/death November 29, 2016 20:07
Cell injury/death leads to Neuroinflammation November 07, 2019 09:36
Neuroinflammation leads to N/A, Neurodegeneration February 23, 2021 05:47
N/A, Neurodegeneration leads to Impairment, Learning and memory April 07, 2018 04:29
N/A, Neurodegeneration leads to Neuroinflammation June 13, 2018 09:35
Lead November 29, 2016 18:42

Abstract

In the abstract section, authors should provide a concise and informative summation of the AOP under development that can stand-alone from the AOP page. Abstracts should typically be 200-400 words in length (similar to an abstract for a journal article). Suggested content for the abstract includes the following: The background/purpose for initiation of the AOP’s development (if there was a specific intent) A brief description of the MIE, AO, and/or major KEs that define the pathway A short summation of the overall WoE supporting the AOP and identification of major knowledge gaps (if any) If a brief statement about how the AOP may be applied (optional). The aim is to capture the highlights of the AOP and its potential scientific and regulatory relevance More help

This AOP is an extension of AOP 13 linking NMDAR chronic inhibition during brain development to impairment of learning and memory. It links chronic NMDA receptors inhibition during brain development to Adverse Outcomes, i.e. neurodegeneration in hippocampus and cortex with amyloid plaque deposition and tau hyperphosphorylation and impairment of learning and memory, which are considered as hallmark of Alzheimer's disease. It introduces another KE, Neuroinflammation, which is involved in several neurodegenerative diseases. With Neuroinflammation and Neurodegeneration, this AOP connects to AOP 48, where in adult brain, « neuroinflammation » leads to « Neurodegeneration » ; « Neurodegeneration » leads to « Decreased neuronal network function », which finally leads to « Impairement of learning and memory ». Both neurodegeneration and cognitive deficits are observed in Alzheimer’s pathology. But as neurodegenerative diseases are complex and multifactorial, the authors proposed two Adverse outcomes: one at the organism level « Impairment of learning and memory», and one at the organ level, « neurodegeneration ». Both are regulatory endpoints. This AOP integrates in the network of AOPs relative to neurotoxicity testing.

This AOP is based on the hypothesis of Landrigan and coworkers (2005) proposing an early origin of neurodegenerative diseases in later life. The chemical initiator known to block NMDARs and used in this AOP for the empirical support is lead (Pb), which is a well-known developmental neurotoxicant. In epidemiological studies of adults, cumulative lifetime lead exposure has been associated with accelerated decline in cognition (Bakulski et al., 2012), suggesting that long term exposure to lead during brain development or occupational exposure in adulthood increases the risk to develop a neurodegenerative disease of Alzheimer's type. The long latency period between exposure and late-onset of neurodegeneration and cognitive deficits gives a very broad life-stage applicability, where developmental exposure has consequences in the aging brain. Such a long temporal delay between exposure and adverse outcome is a real difficulty and challenge for neurotoxicity testing. As the Key Event « Neuroinflammation » appears to play a crucial role in the neurodegenerative process, the authors propose to include the measurement of this apical KE in the battery of regulation-required neurotoxicity testing.

Background (optional)

This optional subsection should be used to provide background information for AOP reviewers and users that is considered helpful in understanding the biology underlying the AOP and the motivation for its development. The background should NOT provide an overview of the AOP, its KEs or KERs, which are captured in more detail below. Examples of potential uses of the optional background section are listed on pages 24-25 of the User Handbook. More help

Summary of the AOP

This section is for information that describes the overall AOP. The information described in section 1 is entered on the upper portion of an AOP page within the AOP-Wiki. This is where some background information may be provided, the structure of the AOP is described, and the KEs and KERs are listed. More help

Events:

Molecular Initiating Events (MIE)
An MIE is a specialised KE that represents the beginning (point of interaction between a stressor and the biological system) of an AOP. More help
Key Events (KE)
This table summarises all of the KEs of the AOP. This table is populated in the AOP-Wiki as KEs are added to the AOP. Each table entry acts as a link to the individual KE description page.  More help
Adverse Outcomes (AO)
An AO is a specialised KE that represents the end (an adverse outcome of regulatory significance) of an AOP.  More help
Sequence Type Event ID Title Short name
1 MIE 201 Binding of antagonist, NMDA receptors Binding of antagonist, NMDA receptors
2 KE 195 Inhibition, NMDARs Inhibition, NMDARs
3 KE 52 Decreased, Calcium influx Decreased, Calcium influx
4 KE 381 Reduced levels of BDNF BDNF, Reduced
5 KE 55 Cell injury/death Cell injury/death
6 KE 188 Neuroinflammation Neuroinflammation
7 AO 352 N/A, Neurodegeneration N/A, Neurodegeneration
8 AO 341 Impairment, Learning and memory Impairment, Learning and memory

Relationships Between Two Key Events (Including MIEs and AOs)

This table summarises all of the KERs of the AOP and is populated in the AOP-Wiki as KERs are added to the AOP. Each table entry acts as a link to the individual KER description page.To add a key event relationship click on either Add relationship: events adjacent in sequence or Add relationship: events non-adjacent in sequence.For example, if the intended sequence of KEs for the AOP is [KE1 &gt; KE2 &gt; KE3 &gt; KE4]; relationships between KE1 and KE2; KE2 and KE3; and KE3 and KE4 would be defined using the add relationship: events adjacent in sequence button.  Relationships between KE1 and KE3; KE2 and KE4; or KE1 and KE4, for example, should be created using the add relationship: events non-adjacent button. This helps to both organize the table with regard to which KERs define the main sequence of KEs and those that provide additional supporting evidence and aids computational analysis of AOP networks, where non-adjacent KERs can result in artifacts (see Villeneuve et al. 2018; DOI: 10.1002/etc.4124).After clicking either option, the user will be brought to a new page entitled ‘Add Relationship to AOP.’ To create a new relationship, select an upstream event and a downstream event from the drop down menus. The KER will automatically be designated as either adjacent or non-adjacent depending on the button selected. The fields “Evidence” and “Quantitative understanding” can be selected from the drop-down options at the time of creation of the relationship, or can be added later. See the Users Handbook, page 52 (Assess Evidence Supporting All KERs for guiding questions, etc.).  Click ‘Create [adjacent/non-adjacent] relationship.’  The new relationship should be listed on the AOP page under the heading “Relationships Between Two Key Events (Including MIEs and AOs)”. To edit a key event relationship, click ‘Edit’ next to the name of the relationship you wish to edit. The user will be directed to an Editing Relationship page where they can edit the Evidence, and Quantitative Understanding fields using the drop down menus. Once finished editing, click ‘Update [adjacent/non-adjacent] relationship’ to update these fields and return to the AOP page.To remove a key event relationship to an AOP page, under Summary of the AOP, next to “Relationships Between Two Key Events (Including MIEs and AOs)” click ‘Remove’ The relationship should no longer be listed on the AOP page under the heading “Relationships Between Two Key Events (Including MIEs and AOs)”. More help

Network View

The AOP-Wiki automatically generates a network view of the AOP. This network graphic is based on the information provided in the MIE, KEs, AO, KERs and WoE summary tables. The width of the edges representing the KERs is determined by its WoE confidence level, with thicker lines representing higher degrees of confidence. This network view also shows which KEs are shared with other AOPs. More help

Stressors

The stressor field is a structured data field that can be used to annotate an AOP with standardised terms identifying stressors known to trigger the MIE/AOP. Most often these are chemical names selected from established chemical ontologies. However, depending on the information available, this could also refer to chemical categories (i.e., groups of chemicals with defined structural features known to trigger the MIE). It can also include non-chemical stressors such as genetic or environmental factors. Although AOPs themselves are not chemical or stressor-specific, linking to stressor terms known to be relevant to different AOPs can aid users in searching for AOPs that may be relevant to a given stressor. More help
Name Evidence Term
Lead

Life Stage Applicability

Identify the life stage for which the KE is known to be applicable. More help
Life stage Evidence
During brain development

Taxonomic Applicability

Latin or common names of a species or broader taxonomic grouping (e.g., class, order, family) can be selected. In many cases, individual species identified in these structured fields will be those for which the strongest evidence used in constructing the AOP was available in relation to this KE. More help
Term Scientific Term Evidence Link
human Homo sapiens Low NCBI
Monkey Monkey High NCBI
rat Rattus norvegicus High NCBI
mouse Mus musculus Moderate NCBI
zebrafish Danio rerio Moderate NCBI

Sex Applicability

The authors must select from one of the following: Male, female, mixed, asexual, third gender, hermaphrodite, or unspecific. More help

Overall Assessment of the AOP

This section addresses the relevant biological domain of applicability (i.e., in terms of taxa, sex, life stage, etc.) and WoE for the overall AOP as a basis to consider appropriate regulatory application (e.g., priority setting, testing strategies or risk assessment). The goal of the overall assessment is to provide a high level synthesis and overview of the relative confidence in the AOP and where the significant gaps or weaknesses are (if they exist). Users or readers can drill down into the finer details captured in the KE and KER descriptions, and/or associated summary tables, as appropriate to their needs.Assessment of the AOP is organised into a number of steps. Guidance on pages 59-62 of the User Handbook is available to facilitate assignment of categories of high, moderate, or low confidence for each consideration. While it is not necessary to repeat lengthy text that appears elsewhere in the AOP description (or related KE and KER descriptions), a brief explanation or rationale for the selection of high, moderate, or low confidence should be made. More help

The aim of this AOP is to capture the KEs and KERs that occur after chronic binding of antagonist to NMDA receptors in neurons of hippocampus and cortex during brain development and that lead to neurodegeneration with impairment in learning and memory in later life.  Neurodegenreation with accumulation of amyloid plaques and hyperphosphorylated tau, as well as cognitive deficit are associated with Alzheimer-type neurodegeneration. Currently, the hypothesis of Landrigan et al., (2005) of developmental origins of neurodegenerative diseases has been demonstrated in monkeys, in rats, mice  and in zebrafish following Pb treatment (Zawia and Basha, 2005; Basha and Reddy, 2010; Bihaqi et al., 2014a; Bihaqi et al., 2014b ; Lee and Freeman, 2014). There is strong agreement that Alzheimer's disease is progressive and that neurodegeneration is occuring mainly in hippocampus and cortex, associated with cognitive deficits (Schoemaker et al., 2014). This AOP uses the MIE and several KEs of the AOP 13 entitled "Binding of antagonist to N-methyl-D-aspartate receptors (NMDARs) during brain development induces impairment of learning and memory abilities ", with an additional KE: neuroinflammation and two AOs: an AO at the organ level: Neurodegeneration in hippocampus and cortex and an AO at the organism level: Impairment of learning and memory.  Impairment of learning and memory is the same AO as in AOP 13, but the point is that this AO is detected when the brain is aging, and it is due to neurodegeneration with accumulation of amyloid peptides and tau hyperphosphorylation. The recent review by Tartatglione and coworkers (2016) is a very good summary of the challenges and experimental studies described in this AOP.

Developmental Pb exposure has adverse effects on cognitive functioning that can persist into adulthood and may be exacerbated with aging (Schneider et al., 2013). Such delayed effects may be due to epigenetic effects of developmental Pb exposure on DNA methylation mediated at least in part through dysregulation of methyltransferases observed often at the lowest level of exposure (Schneider et al., 2013). In addition, key neurodevelopmental events, such as neural differentiation, cell migration and network formation may be modulated by Pb exposure, predisposing the brain for alterations in higher brain functions, such as learning and memory, and this at different ages (for review, see Aschner et al., 2017). The fact that neuroinflammation triggered during early brain development was shown to cause Alzheimer-like pathology when aging (Krstic et al., 2012), suggests that chronic neuroinflammation may play a causal role in cognitive decline in aging. A recent report described a mechanistic link between chronic inflammation and aging microglia; and a causal role of aging microglia in neurodegenerative cognitive deficits: A sirtuin 1 (SIRT1) deficiency was observed in aging microglia, leading to a selective activation of IL1-b transcription mediated through hypomethylation of IL-1b proximal promoter exacerbating aging or tau-associated cognitive deficits (Cho et al. 2015). Taken together, these data suggest that Pb-induced neuroinflammation during brain development may underlie the delayed effects on cognitive deficits in aging, as depicted in the proposed AOP

Domain of Applicability

The relevant biological domain(s) of applicability in terms of sex, life-stage, taxa, and other aspects of biological context are defined in this section. Biological domain of applicability is informed by the “Description” and “Biological Domain of Applicability” sections of each KE and KER description (see sections 2G and 3E for details). In essence the taxa/life-stage/sex applicability is defined based on the groups of organisms for which the measurements represented by the KEs can feasibly be measured and the functional and regulatory relationships represented by the KERs are operative.The relevant biological domain of applicability of the AOP as a whole will nearly always be defined based on the most narrowly restricted of its KEs and KERs. For example, if most of the KEs apply to either sex, but one is relevant to females only, the biological domain of applicability of the AOP as a whole would be limited to females. While much of the detail defining the domain of applicability may be found in the individual KE and KER descriptions, the rationale for defining the relevant biological domain of applicability of the overall AOP should be briefly summarised on the AOP page. More help

This AOP is not sex dependent. Regarding the life stage applicability, MIE induced during brain development can have consequences when brain is aging, according to the hypothesis proposed by Landrigan and coworkers (2005). However, it is also possible that the AO does not depend exclusively on developmental exposure, since cumulative occupational exposure also decreased cognitive functions in aging (Stewart et al., 2006).

Essentiality of the Key Events

An important aspect of assessing an AOP is evaluating the essentiality of its KEs. The essentiality of KEs can only be assessed relative to the impact of manipulation of a given KE (e.g., experimentally blocking or exacerbating the event) on the downstream sequence of KEs defined for the AOP. Consequently evidence supporting essentiality is assembled on the AOP page, rather than on the independent KE pages that are meant to stand-alone as modular units without reference to other KEs in the sequence.The nature of experimental evidence that is relevant to assessing essentiality relates to the impact on downstream KEs and the AO if upstream KEs are prevented or modified. This includes: Direct evidence: directly measured experimental support that blocking or preventing a KE prevents or impacts downstream KEs in the pathway in the expected fashion. Indirect evidence: evidence that modulation or attenuation in the magnitude of impact on a specific KE (increased effect or decreased effect) is associated with corresponding changes (increases or decreases) in the magnitude or frequency of one or more downstream KEs.When assembling the support for essentiality of the KEs, authors should organise relevant data in a tabular format. The objective is to summarise briefly the nature and numbers of investigations in which the essentiality of KEs has been experimentally explored either directly or indirectly. See pages 50-51 in the User Handbook for further definitions and clarifications.  More help

Table: Essentiality of KEs

2 Support for Essentiality of KEs

Defining Question

Are downstream KEs and/or the AO prevented if an upstream KE is blocked ?

High (Strong)

Moderate

Low (Weak)

Direct evidence from specifically designed experimental studies illustrating essentiality for at least one of the important KEs (e.g. stop/reversibility studies, antagonism, knock out models, etc.)

Indirect evidence that sufficient modification of an expected modulating factor attenuates or augments a KE leading to increase in KE down or AO

No or contradictory experimental evidence of the essentiality of any of the KEs

KE1

NMDARs inhibition

STRONG

Activation of NMDAR results in LTP, which is related to increase synaptic strength and memory formation in hippocampus (Johnston et al., 2009).

KE2

Calcium influx decreased

STRONG

In CNS, many intracellular responses to modified calcium level are mediated by calcium/calmoduline-regulated protein kinases (Wayman et al., 2008). Mice with a mutation of calmoduline kinase II, which is abundantly found in hippocampus, have shown spatial learning impairment (Silva et al., 1992)

KE3

Release of BDNF, reduced

STRONG

BDNF serves essential function in synaptic plasticity (Poo, 2001) and is crucial for learning and memory processes (Lu et al., 2008). Precursor form of BDNF and mature BDNF are decreased in the preclinical stages of Alzheimer's disease (Peng et al., 2005)

KE4

Cell Injury/death, increased

STRONG

Several studies dealing with postnatal administration of NMDAR antagonists such as MK 801, ketamine or ethanol have shown a devastating cell apoptotic degeneration in several brain areas of animal models resulting in learning deficits (Creeley and Olney, 2013)

KE5

Neuroinflammation

MODERATE

Rationale: Rats treated with Pb from PND 24 to 80 showed a neuroinflammatory response associated with neuronal death in hippocampus and LTP impairment. These effects were significantly reversed by administration of minocycline, an antibiotic known to block microglial reactivity (Liu et al., 2012), demonstrating the essentiality of neuroinflammation for neurodegeneration in hippocampus and impairment of memory processes. In addition, the fact that neuroinflammation triggered during brain development by a systemic immune challenge caused Alzheimer's like pathology (Krstic et al., 2012), showed the central role of neuroinflammation in this pathology. In addition, in a mouse model of Alzheimer's disease, the blockade of microglial cell proliferation and the shifting of the microglial inflammatory profile to an anti-inflammatory phenotype by inhibiting the colony-stimulating factor 1 receptor on microglial cells, prevented synaptic degeneration and improved cognitive functions (Olmos-Alonso et al., 2016). This latter experiment has not been done during brain development. But the hypothesis is that a chronic neuroinflammation during a prolonged period increased the risk to develop an Alzheimer's neurodegenerative disease in aging (Krstic and Knuesel, 2013).

However, as other mechanisms such epigenetic modifications can lead to accumulation of amyloid plaques- and tau hyperphosphorylation-related neurodegeneration, and due to some inconsistencies of anti-inflammatory treatments as protection against the neurodegenerative process, the essentiality of Neuroinflammation was considered as moderate.

AO (at organ level)

Neurodegeneration in

 hippocampus and cortex

STRONG

Several studies descibed Pb-induced accumulation of amyloid peptides and hyperphosphorylated and Pb-induced cell injury/deathin hippocampus or decrease in hippocampal volume, what are all well accepted landmarks of Alzheimer's pathology (Lloret et al., 2015). As described in AOP 48, neurodegeneration can lead to "Decreased neuronal network function" which in turn leads to "impairment of learning and memory", which is also considered as a hallmark of Alzheimer's pathology (Schoemaker et al., 2014).

However, there is some controversy about the relationship between increased accumulation of amyloid plaques and increased cognitive deficits:  Lichtenstein and coworkers (2010) described that accumulation of amyloid plaques reaches a plateau, whereas a temporal relationship is observed between increased microglial activation, widespread degeneration (decreased hippocampal volume) and increased cognitive deficits. Therefore the essentiality for accumulation of amyloid and tau to cognitive deficits should be considered as moderate. But, as cell injury/death in hippocampus and cortex or decrease in hippocampal volume due to widespread neurodegeneration is strongly associated to impairment in learning and memory, the essentiality of this KE has been rated as strong.

 AO (at organism level)

Impairment of learning and memory

STRONG

Neurodegenerative diseases are complex and multifactorial, depend on gene-environment interactions, and have a slow temporal evolution (Sherer et al., 2002; Steece-Collier et al., 2002; Tsang and Soong, 2003); Mutter et al., 2004). A direct association between Pb exposure during brain development and Alzheimer's pathology is not supported by epidemiological studies. However, two studies reported that past adult exposure is linked with neurodegeneration (Stewart et al., 2006) and decline in cognitive function (Schwartz et al., 2000), effects which were observed long after exposure ceases. Tibia lead levels were good predictors of these delayed effects. Another study showed an association between lead exposure early in life with cognitive and behavioral consequences in early adulthood (Agency for toxic substances, 1997). Despite the lack of specific epidemiological evidence, the principle of delayed effects occuring long after exposure, as well as strong evidence from experimental studies (for review, see Chin-Chan et al., 2015) suggest that long-term exposure to environmental toxicants such as Pb during brain development or exposure later in life can be considered as a risk factor for the development of neurodegenerative diseases in aging.

 

Evidence Assessment

The biological plausibility, empirical support, and quantitative understanding from each KER in an AOP are assessed together.  Biological plausibility of each of the KERs in the AOP is the most influential consideration in assessing WoE or degree of confidence in an overall hypothesised AOP for potential regulatory application (Meek et al., 2014; 2014a). Empirical support entails consideration of experimental data in terms of the associations between KEs – namely dose-response concordance and temporal relationships between and across multiple KEs. It is examined most often in studies of dose-response/incidence and temporal relationships for stressors that impact the pathway. While less influential than biological plausibility of the KERs and essentiality of the KEs, empirical support can increase confidence in the relationships included in an AOP. For clarification on how to rate the given empirical support for a KER, as well as examples, see pages 53- 55 of the User Handbook.  More help

1. Concordance of dose-response and temporal concordance between KEs and the AO

It is difficult to analyze the dose-response relationships between the different KEs, (i) because of the long temporal delay between MIE and AOs ; (ii) because no study has analyzed them simultaneously, and (iii)  because of the difficulties in extrapolating in vitro to in vivo data. As the apical KEs and AO occur and can be measured years after exposure, even when Pb blood level has returned to normal, measurement of bone Pb content has been proposed as a measurement of historical Pb exposure in adults (Bakulski et al., 2012, 2014).  The following table gives an overview of the doses/concentrations and exposure duration at which the different KEs were measured.

KE1

KE2

KE3

KE4

K5

AO at organ level

AO at organism level

NMDAR inhibition

Calcium influx, decreased

BDNF release, decreased

Cell injury/death

Neuroinflammation

Neurodegeneration with amyloid plaques and tau hyperphosphorylation

Impairment of learning and memory

 

Pb 2.5-5 mM acute

inhibits

NMDAR whole cell and channel current in hippocampal neurons

 

(Alkondon et al., 1990)

Pb 100 nM 1h-24h

decrease Ca2+ in embryonic rat hippocampal neurons

 

(Ferguson et al., 2000)

No direct evidence

 

Pb 2mM in drinking water 3 weeks before mating till weaning (PND 21) resulting in

at PND 21

Pb blood 108.8 mg/L

Pb hippoc. 0.253 mg/g

at PND 91

Pb blood 39.27 mg/L

Pb hippoc. 0.196 mg/g

 

about 35% decrease in synapses in hippocampus

 

about 30% decrease of hippocampal neurons

 

(Xiao et al., 2014)

 

In vivo

0.22 ppm (together with As and Cd) from gestational day 5 till day 180

 

in adulthood: IL-1b, TNF-a, IL-6 increased 2x

 

Ashok et al., 2015

 

Rats exposed to Pb 100 ppm for 8 weeks (from PND 24 to 80) caused at the end of treatment microglial activation in hippocampus.

(Liu et al., 2012

 

 

In vitro 

10-6-10-4 M for 10 days

in 3D cultures of fetal rat brain cells

 

microglial and astrocyte reactivities

 

(Zurich et al., 2002)

 

co-cultures of hippocampal neurons with microglial cells treated with Pb (50 micomol/L for 48h) caused microglial activation and upregulation of IL-1beta, TNF-alpha and i_NOS

(Liu et al., 2012)

Monkeys exposed to

Pb 1.5 mg/kg/day

from birth to 400 days

 

at 23 years of age

 

Tau accumulation

Overexpression of amyloid-beta protein precursor and of amyloid-beta

enhanced pathologic neurodegeneration

 

(Bihaqi et al., 2011; Bihaqi and Zawia, 2013)

 

Mice exposed to

Pb 0.2% in drinking water from PND 1-20 or from PND 1-20 + From 3-7 months

 

at 700 days of age

 

elevated protein and mRNA for tau

and

aberrant site-specific tau hyperphosphorylation

 

(Bihaqi et al., 2014)

 

Human Tg-SWDI APP transgenic mice , PB 50 mg/kg by gavage for 6 weeks exhibit increase AB in CSF, cortex and hippocampus and increased amyloid plaque load (Gu et al., 2012)

 

Rats exposed to Pb 100 ppm for 8 weeks (from PND 24 to 80) caused at the end of treatment neuronal death in hippocampus.

(Liu et al., 2012)

 

Mice exposed to

Pb 0.2% in drinking water from PND 1-20 or from PND 1-20 and from 3-7 months

 

Tested at 700 days of age

 

Decrease in cognitive functions (Morris water maze, Y maze testing for spatial memory and memory, a hippocampal formation-dependent task)

 

(Bihaqi et al., 2014b)

 

Rats exposed to Pb 100 ppm for 8 weeks (from PND 24 to 80) reduced hippocampal LTP level at the end of the treatment

(Liu et al., 2012)

 

 

Human Tg-SWDI APP transgenic mice , PB 50 mg/kg by gavage for 6 weeks showed an impaired spatial learning (Gu et al., 2012)

 

2. Strength, consistency and association of AO and MIE

The accepted molecular mechanism of action of the chemical initiator Pb is inhibition of NMDARs (Alkondon et al., 1990; Gavazzo et al., 2001, 2008; Guilarte et al., 1992; Omelchenko et al., 1997) and several experimental studies in rat, monkey and zebrafish linked chronic exposure to Pb during brain development to Alzheimer's-like neurodegeneration with cognitive deficits (Zawia and Basha, 2005; Basha and Reddy, 2010; Bihaqi et al., 2014a; Bihaqi et al., 2014 b; Lee and Freeman, 2014). This AOP is defined by a single environmental chemical, Pb. However, other NMDAR antagonists used as general anesthetics (MK 801, phenylcyclidine, ketamine) applied during brain development may also lead to functional impairments in cognitive domains relevant to memory. The effects of these anesthetics on brain function appear to have a delayed onset, and can be very long-lasting if not permanent. In general, longer durations, higher concentrations and longer or repeated exposures tend to exacerbate impairments (for review, see Walters and Paule, 2017). The mechanisms underlying anesthetic-induced neurotoxicity are unclear, but several hypotheses have been proposed: impairment of mitochondrial integrity and function, dysregulation of intracellular calcium and neuroinflammation have all been implicated (Lei et al., 2012). Some of these mechanisms are common to the KEs described in this AOP, suggesting that such delayed effects on memory processes can be a general consequence of developmental brain exposure to NMDAR inhibitors. However, no studies have yet reported that these other NMDAR inhibitors cause amyloid plaque deposition or tau hyperphosphorylation associated with Alzheimer-like neurodegeneration when aging.

Interestingly, memantine, a NMDAR antagonist used in the treatment of Alzheimer's disease, was shown to improve cognitive functions (for review, see Dekundy, 2006). This might be considered as a discrepancy with the described AOP considering Pb as an antagonist of NMDAR and its potential risk to cause cognitive deficits and amyloid plaque accumulation, which are hallmarks of Alzheimer's disease. However, memantine antagonism of NMDAR is quite different (low affinity and voltage-dependent) and the window of exposure differs completely, since memantine is applied in aged patients when the disease has broken out; whereas the risk of delayed neurodegeneration described in this AOP is due to NMDAR inhibition during brain development.

3. Biological Plausibility, and empirical support

 

Defining Question

High /Strong

Moderate

Low/weak

Support for Biological Plausibility of KERs

Is there a mechanistic (i.e. structural or functional) relationship between KEup and KEdown consistent with established biological knowledge?

Extensive understanding of the KER based on extensive previous documentation and broad acceptance

The KER is plausible based on analogy to accept biological relationship but scientific understanding is not completely established

There is empirical support for a statistical association between KEs but the structural or functional relationship between them is not understood

MIE to KE inhibition of NMDARs

 

Extensive understanding

Limited conflicting data

 

 

KE NMDAR inhibition to KE calcium influx, decreased

 

Extensive understanding

Limitied conflicting data

 

 

KE calcium influx, decreased to KE release of BDNF, decreased

 

Extensive understanding

Limited conflicting data

 

 

KE release of BDNF, decreased to KE Cell  Injury/death

 

Extensive understanding

Limited conflicting data

 

 

KE Cell injury/death to KE Neuroinflammation

 

 

The general mechanisms linking cell injury/death to neuroinflammation is well accepted. However, it is mainly descibed in adult brain. However, a neuroinflammatory response was found following Pb exposure of 3D cultures during synaptogenesis and myelination (Zurich et al., 2002). A controversy exists about apoptosis and neuroinflammation, but some empirical evidences has been provided.

 

The fact that cell injury/deat leads to neuroinflammation and that neuroinflammation leads to neurodegeneration is known as avicious circle and is involved in neurodegenerative diseases, suggesting that neuroinflammation exacerbates the neurodegenerative process (Griffin et al., 1998; 2006)

 

 

 

KE Neuroinflammation to AO Neurodegeneration in Hippocampus and cortex

 

 

In adult, the early involvement of neuroinflammation in the neurodegenerative process is widely accepted.

 

In immature brain, one study in mice link gestational induction of neuroinflammation to late neurodegeneration with accumulation of aberrant amyloid and tau (Kristic et al., 2012).

 

 

 

There is in vitro experimental data following Pb exposure linking neuroinflammation to extensive neuronal death in immature cells.

In vivo, There are several studies linking early Pb exposure to late neurodegeneration in several species. However, the mechanisms involved is epignenetic modifications of genes involved in the amyloid cascade. Such epigenetic modifications may be due to ROS released by the neuroinflammatory process (Bolin et al., 2006).

Therefore the link may be indirect and needs further analyses.

 

AO Neurodegeneration in hippocampus and cortex to KE Neuroinflammation 

Concept of vicious circle where neuroinflammation lead to neurodegeneration and vice versa (Griffin et al., 1998, 2006) 

 

 

 There are no specific empirical data for the chemical initiator Pb.

 AO Neurodegeneration in hippocampus and cortex  to AO Impairment of learning and memory

 

 The role of hippocampus in memory processes is well accepted. Alterations of LTP in hippocampus of rats exposed to Pb has been described (Liu et al., 2012), as well as preferential accumulation of hyperphosphorylated tau in frontal cortex of mice exposed during development to Pb. These mice exhibited cognitive deficit when aging (Bihaqi et al., 2014b).

 

 

Quantitative Understanding

Some proof of concept examples to address the WoE considerations for AOPs quantitatively have recently been developed, based on the rank ordering of the relevant Bradford Hill considerations (i.e., biological plausibility, essentiality and empirical support) (Becker et al., 2017; Becker et al, 2015; Collier et al., 2016). Suggested quantitation of the various elements is expert derived, without collective consideration currently of appropriate reporting templates or formal expert engagement. Though not essential, developers may wish to assign comparative quantitative values to the extent of the supporting data based on the three critical Bradford Hill considerations for AOPs, as a basis to contribute to collective experience.Specific attention is also given to how precisely and accurately one can potentially predict an impact on KEdownstream based on some measurement of KEupstream. This is captured in the form of quantitative understanding calls for each KER. See pages 55-56 of the User Handbook for a review of quantitative understanding for KER's. More help

With an Adverse Outcome occurring after such a long delay after the MIE, it is extremely difficult to make a quantitative link, since the AO can occur when serum Pb levels have returned to normal. Bakulski and coworkers (2012) therefore proposed measuring Pb bone content as an index of historical Pb exposure. Similarly, Schwartz and coworkers (2000) showed that tibia Pb levels were good predictors of delayed cognitive decline of former organolead workers. Thus, Pb blood level is rather representative of acute exposure, whereas Pb bone level represents long-term accumulation (Dorsey et al., 2006).

Regarding the KER "cell injury/death to neuroinflammation", it is accepted that neuronal injury may be sufficient to trigger a neuroinflammatory response. But, because of the neuroprotective or neuroreparative potential of neuroinflammation, it is possible that the consequences of neuroinflammation will be in a first step positive, with microglia expressing the M2 phenotype. After an exposure arrest and a temporal delay (Sandström et al., 2014), or in the presence of cell death (Nakajima and Kohsaka, 2004; Hanish and Kettenmann, 2007), microglia can acquire the M1 neurodegenerative phenotype. Therefore, it is rather the qualitative phenotype of neuroinflammation that will induce neurodegeneration. However, a possible correlation of increased microglial reactivity, measured by PET, and a decrease in hippocampal volume, measured by MRI, suggests, in advanced Alzheimer's disease, a possible link between the intensity of neuroinflammation and the neurodegenerative consequences (Lichtenstein et al., 2010).

Considerations for Potential Applications of the AOP (optional)

At their discretion, the developer may include in this section discussion of the potential applications of an AOP to support regulatory decision-making. This may include, for example, possible utility for test guideline development or refinement, development of integrated testing and assessment approaches, development of (Q)SARs / or chemical profilers to facilitate the grouping of chemicals for subsequent read-across, screening level hazard assessments or even risk assessment. While it is challenging to foresee all potential regulatory application of AOPs and any application will ultimately lie within the purview of regulatory agencies, potential applications may be apparent as the AOP is being developed, particularly if it was initiated with a particular application in mind. This optional section is intended to provide the developer with an opportunity to suggest potential regulatory applications and describe his or her rationale.To edit the “Considerations for Potential Applications of the AOP” section, on an AOP page, in the upper right hand menu, click ‘Edit.’ This brings you to a page entitled, “Editing AOP.” Scroll down to the “Considerations for Potential Applications of the AOP” section, where a text entry box allows you to submit text. In the upper right hand menu, click ‘Update AOP’ to save your changes and return to the AOP page or 'Update and continue' to continue editing AOP text sections.  The new text should appear under the “Considerations for Potential Applications of the AOP” section on the AOP page. More help

This AOP aims at giving a conceptual framework to mechanistically understand an apical hazard, which can occur long after initial exposure; this hazard is not captured in standard regulatory neurotoxicity testing.

The KE "neuroinflammation", which is shared with other AOPs, appears to play an early and central role in the neurodegenerative process (Eikelenboom et al., 2000; Whitton, 2007; Krstic et al., 2012). Neuroinflammation is observed in most neurodegenerative diseases including Alzheimer's disease  (Whitton, 2007 ; Tansey and Goldberg, 2009 ; Niranjan, 2014 ; Verkhratsky et al., 2014). Neuroinflammation can also be triggered by several classes of toxicants (Monnet-Tschudi et al., 2007). Any toxicant able to trigger a neuroinflammatory response expressing the neurodegenerative phenotype should be considered as a risk factor for neurodegenerative diseases. Therefore, testing for toxicant-induced neuroinflammation should be used as an endpoint in regulatory toxicology. The standard neurotoxicity testing does not require measurement of any marker of neuroinflammation, except for fuel additives, where testing for a potential increase in glial fibrillary acidic protein (GFAP), as marker of astrocyte reactivity, is mandatory according to US EPA (40 CFR 79 67).

The evolution of regulation towards mechanistically-driven approaches for supporting hazard identification implies also the development of in vitro testing. Three-dimensional cultures, prepared from fetal rat brain cells, exhibiting an histotypic organisation comprising all types of brain cells (specifically microglial cells and astrocytes, as effector cells of neuroinflammation) and allowing long-term maintenance for repeated exposure and for studying the evolution of neuroinflammatory phenotypes, are already available (Alépée et al., 2014; Monnet-Tschudi et al., 2007 ; Sandström et al., 2014). Similar 3D cultures prepared from human pluripotent stem cells are in development (Schwartz et al., 2015; Sandström et al., 2017).

References

List the bibliographic references to original papers, books or other documents used to support the AOP. More help

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