Identifying the role of NMDA receptors in STP through investigation of synaptic plasticity and discovery of novel subtype-specific antagonists

Lead Research Organisation: University of Bristol
Department Name: Physiology and Pharmacology

Abstract

The purpose of the proposed research is to make new chemicals that can be used as tools to study the mechanisms by which the central nervous system (CNS) exercises its multiple functions. Communication between nerve cells occurs at specialised zones called synapses. 'Messages' are sent across the synapse by the release of a chemical called a neurotransmitter. Nerve impulses in the pre-synaptic cell are conveyed 'through' the synapse by the release of neurotransmitter which binds to specialized proteins (called receptors) situated on the surface of the recipient (or postsynaptic cell) which generate small electrical impulses when activated by neurotransmitter. One such neurotransmitter is glutamate, which is present at synapses throughout the CNS. Our previous research helped establish that glutamate can interact with a family of structurally related glutamate receptor subtypes each performing different functions in the CNS. The present work aims to design and make novel chemicals (pharmacological tools) that modulate the action of glutamate at particular glutamate receptor subtypes. One particular subtype of glutamate receptor, the N-methyl-D-aspartate receptor (or NMDAR for short), named after the chemical that selectively activates it, is a tetramer made up of two types of protein subunit known as GluN1 and GluN2. In most areas of the CNS NMDARs are comprised of two GluN1 and two GluN2 subunits. Four GluN2 subunits are known, GluN2A-D, and they can be combined with GluN1 to form tetramers of different subunit composition, which are differentially expressed in the brain. It is now well established that the strength of synaptic connections between nerve cells can be regulated (a process known as synaptic plasticity). Synaptic plasticity has been demonstrated experimentally in many areas of the brain including the hippocampus, a part of the brain known to be important in learning and memory. Two major forms of plasticity are long-term potentiation (LTP), which facilitates connections, thereby enhancing communication between neurons and long-term depression (LTD), which decreases neuronal connections and communication between neurons. NMDARs are known to be involved in the mechanisms of LTP and LTD in the hippocampus. We have recently identified that different NMDAR subtypes are differentially involved in various forms of synaptic plasticity that may underline specific forms of learning and memory. What is less clear is the precise role of individual NMDAR subtypes, due in part to the lack of specific pharmacological tools that can discriminate between NMDARs with different GluN2 subunit composition. We have developed a series of novel inhibitors that can modulate NMDAR function either by binding to the same site as glutamate (competitive inhibitors) or binding to a site(s) on the receptor away from the glutamate binding site (allosteric inhibitors). The inhibitors that we have developed have various patterns of GluN2 subunit selectivity and are leads for the development of newer compounds with improved GluN2 subunit selectivity. We plan to develop and use NMDAR inhibitors with the desired GluN2 subunit selectivity to investigate the specific roles of particular NMDAR subtypes in LTP and LTD in the hippocampus. The major focus of this work will be to investigate an early form of LTP known as short-term potentiation (STP) which we hypothesise is important for short-term memory. We will test this hypothesis with behavioural experiments in collaboration with a laboratory in Korea.

Technical Summary

NMDA receptors (NMDARs) are ligand-gated ion channels that belong to the glutamate receptor (GluR) family and are expressed throughout the central nervous system (CNS). In most areas of the CNS NMDARs are comprised of two GluN1 and two GluN2 subunits. The GluN2A-D subunits can combine with GluN1 to form functional tetramers of different subunit composition (or NMDAR subtypes), which are differentially expressed in the brain. NMDARs have been identified as key players in synaptic plasticity and many other CNS functions. We have recently discovered that different NMDAR subtypes are involved in different temporal components of long-term potentiation (LTP). Surprisingly we found that an early component of synaptic potentiation, termed short-term potentiation (STP), comprises two distinct components (which can be separated pharmacologically). This discovery is likely to impact enormously on our understanding of the roles of NMDARs in learning and memory. However, to explore this further we need more selective NMDAR antagonists. In this regard, we have recently developed a number of competitive antagonists and negative allosteric modulators of NMDARs with novel patterns of GluN2 subunit selectivity. Inhibitors with better GluN2 subunit selectivity will be developed using structure-activity relationship data combined with receptor structure based and ligand based drug design. These optimised compounds will be used for the identification of NMDAR subtypes in STP, and other forms of synaptic plasticity, and to identify the role of STP in learning and memory.

Planned Impact

Understanding how the brain works by regulating neuronal activity is a topic of great public and academic interest. The extensively studied NMDA receptors (NMDARs) play a central role in synaptic plasticity and in several neurological disorders. Better understanding of the structure of NMDARs and their properties created exciting new opportunities for rational drug design and the development of new, more potent and selective pharmacological reagents for the analysis of endogenous NMDARs and their complex interactions in their native environment in neurones. This project is thus timely and should have major impact at both academic and social levels. To deliver that impact, it is important that the research is disseminated as effectively as possible. The target audiences are academics, pharmaceutical industry, health professionals, schools and the wider public.
ACADEMICS: The various ways chemists, structural biologists, neuroscientists, neuropharmacologists, postdoctoral scientists employed on the project and the very wide range of undergraduate and postgraduate students benefit from this project are described under 'Academic beneficiaries'.
PHARMACEUTICAL INDUSTRY: Mutually beneficial collaborations with industry are an integral part of the way that we achieve our aims. DJ has worked as a consultant for Eli Lilly's project to develop kainate receptor antagonists for the treatment of a range of neurological disorders. DJ, GC and DL have been members of Lilly's Centre for Cognitive Neuroscience (CCN) since its inception in 2008. The CCN is a major collaborative venture between UK based academics and Lilly, the one remaining Pharma with a major UK presence in the area of neuroscience. DJ and GC have close links with biotechnology companies (Abcam and Tocris Biosciences). These established collaborations will be used to commercialise and distribute new research tools developed during this project (novel subunit selective competitive antagonists and negative allosteric modulators for NMDARs) to the wider research community. Furthermore, there is significant interest in NMDARs by industry as potential drug targets for epilepsy, schizophrenia, neurodegenerative disorders, migraine and chronic pain therefore our results are likely to have an impact in this area.
HEALTH PROFESSIONALS: Because NMDARs have been linked to a range of neurological disorders where effective treatment is currently unavailable, they offer inviting possibilities for therapeutic exploitation. Two channel blockers are used clinically, memantine (moderate-severe Alzheimer's) and ketamine (dissociative anaesthetic) and some inhibitors are used as a drug of abuse (ketamine and PCP). A much better understanding of NMDAR pharmacology and the role of NMDARs in physiological processes will be essential for the development of new therapies and understanding the underlying mechanisms behind the use of inhibitors as drugs of abuse. Therefore, the medical community will also benefit from any new developments in the field.
SCHOOLS: The future of science depends on enthusiastic young scientists. All applicants are involved in University open days, making explanation of science accessible to prospective students and parents. For instance, DJ has developed an interactive tutorial "drug molecules in 3D" through which school children can learn about structures of drug molecules and how they interact with their protein targets.
THE WIDER PUBLIC: All applicants are committed to engage public interest and to shape public perception about the benefits of scientific discovery. We will take every opportunity to directly engage the public and schools through initiatives coordinated by the Bristol University Centre for Public Engagement, Bristol Neuroscience and by local attractions (e.g. the science activity centre '@Bristol'). Our published findings will be promoted to the public through Bristol Neuroscience, University web sites and the University Press Office.

Publications

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Collingridge GL (2017) Antidepressant Actions of Ketamine Versus Hydroxynorketamine. in Biological psychiatry

 
Description We have discovered chemical tools (such as UBP145) that block a specific protein subunit known as GluN2D which is part of a protein complex known as the N-methyl-D-aspartate (NMDA) receptor. With the use of these tools we have shown that the GluN2D subunit is involved in the fundamental processes in a part of the brain known as the hippocampus that are thought to underlie learning and memory. We have also developed both positive and negative allosteric modulators (PAMs (e.g. UBP684) and NAMs (e.g. UBP792) that either enhance or inhibit NMDA receptor function, respectively. These may have therapeutic utility as due to their modulatory properties they do not completely block NMDAR physiological responses (NAMs) or overactivate NMDARs (PAMs) leading to neuronal cell death.
Exploitation Route The chemical tools (UBP145 and UBP684) we have produced is now available to the worldwide neuroscience community and can be used to assess the role of GluN2D-containing NMDA receptors in a variety of brain functions, in neurological disorders such as chronic pain, epilepsy and schizophrenia and in neurodegenerative disorders such as Alzheimer's disease, Parkinson's disease and ischaemic stroke. It may also be used as a template for drug design to produce treatments for some of the disorders mentioned above.
Sectors Chemicals,Education,Pharmaceuticals and Medical Biotechnology

 
Description UBP145, a pharmacological tool that can be used to selectively block GluN2D-containing NMDA receptors and UBP684, a positive allosteric modulator of NMDA receptors, are now sold to the worldwide neuroscience community by a UK company, Hello Bio, which is based in Bristol. Thus, our work is beginning to have economic impact.
First Year Of Impact 2016
Sector Chemicals,Pharmaceuticals and Medical Biotechnology
Impact Types Economic

 
Title UBP145 selective GluN2D NMDA receptor antagonist 
Description UBP145 is a selective GluN2D NMDAR antagonist that can be used to assess the role of GluN2D in brain function. For example it has been used to understand the role of GluN2D in hippocampal synaptic plasticity. 
Type Of Material Technology assay or reagent 
Year Produced 2016 
Provided To Others? Yes  
Impact UBP145 has been used to assess brain function and has been used to show that GluN2D-containing NMDA receptors contribute to a form of short term plasticity in the hippocampus. UBP145 is now sold to the worldwide neuroscience community as a research tool via a company called Hello Bio, which is a neurochemical supplier based in Bristol. 
URL http://www.hellobio.com/ubp145.html?picat=&q=UBP145
 
Title UBP684 a positive allosteric modulator of N-Methyl_D-aspartate receptors 
Description UBP684 can be used as a tool to understand the consequences of potentiating the activity of NMDA receptors in the central nervous system. 
Type Of Material Technology assay or reagent 
Year Produced 2017 
Provided To Others? Yes  
Impact Tool is the first allosteric modulator for NMDARs that potentiates NMDARs containing GluN2A, GluN2B, GluN2C or GluN2D and is now on sale at Hello Bio which is a neurochemical supplier. 
URL https://www.hellobio.com/ubp684.html?picat=&q=ubp684
 
Description DTM collaboration 
Organisation University of Nebraska System
Department Department of Pharmacology and Experimental Neuroscience
PI Contribution Provide medicinal chemistry support to the project, involved in the design and synthesis of new ligands for the NMDA receptor. From 2008 onward this collaboration has led to the following publications: PUBMED ids: 22269804, 22155206, 22111545, 21975018, 21204415, 20858708, 19684252
Collaborator Contribution My partner is involved in the pharmacological evaluation of the compounds synthesized by my research team on NMDA receptor subtypes. The data produced at Nebraska was and still is essential for the success of the project funded by the MRC.
Impact The following publications have arisen from this collaboration since 2008: PUBMED ids: 22269804, 22155206, 22111545, 21975018, 21204415, 20858708, 19684252 A grant application to the National Institute of Mental Health in which my collaborator and I were joint principal investigators has been awarded (see further funding section). This is a multi-disciplinary collaboration involving organic and medicinal chemists, pharmacologists and neuroscientists.
 
Description International conference on glutamate receptors 
Form Of Engagement Activity A talk or presentation
Part Of Official Scheme? No
Geographic Reach International
Primary Audience Other academic audiences (collaborators, peers etc.)
Results and Impact Talked sparked lively discussion and opened up the possibility of new collaborations with scientists in the audience.

No notable impacts
Year(s) Of Engagement Activity 2014
URL http://conferences-on-aminoacidergic-transmission.eu/index.php/conference-2014
 
Description School open day 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach National
Primary Audience Schools
Results and Impact Open day for school students to decide where to apply for undergraduate studies
Year(s) Of Engagement Activity 2016
 
Description Visit of school pupils to School of Physiology, Pharmacology and Neurosceicne 
Form Of Engagement Activity Participation in an open day or visit at my research institution
Part Of Official Scheme? No
Geographic Reach Local
Primary Audience Schools
Results and Impact One of my postdoctoral workers, Erica Burnell was involved in two physiology lab outreach sessions where ~50 school children from a variety of age groups from schools in the local area visited the university and performed experiments using cardiac (ECG) and respiratory (vitalograph) equipment. They had the opportunity to use equipment they haven't come across before, leading to a discussion of the techniques used and their role in understanding cardiac and respiratory physiology. The students also had the chance to talk to PhD students and postdoctoral workers about any questions they may have about going to university. This session sparked the interest of pupils in scientific research and applying to University to undertake a science degree.
Year(s) Of Engagement Activity 2016