2013 — 2016 |
Gray, John Alan Gray, John Alan [⬀] |
K08Activity Code Description: To provide the opportunity for promising medical scientists with demonstrated aptitude to develop into independent investigators, or for faculty members to pursue research aspects of categorical areas applicable to the awarding unit, and aid in filling the academic faculty gap in these shortage areas within health profession's institutions of the country. |
Synaptic Targeting and Regulation of Nmdar Subunits @ University of California At Davis
DESCRIPTION (provided by applicant): The overall goal of this research program is to better understand the molecular mechanisms by which N-methyl-D-aspartate receptors (NMDARs) are regulated to modulate synapse development and synaptic plasticity, and how these processes might be disrupted in complex neuropsychiatric disorders such as schizophrenia. The applicant for this K08 Mentored Clinical Scientist Research Career Development Award, Dr. John Gray, is a psychiatrist and a postdoctoral fellow with Dr. Roger Nicoll at UCSF. Dr. Gray's long-term research goals are to lead an independent research laboratory in psychiatric neuroscience at an academic institution combining cellular, molecular, electrophysiological, and genetic approaches to study synapse function with the goal of understanding how disruptions of the normal mechanisms of synapse development might underlie the pathophysiology of major neuropsychiatric disorders. Though etiological mechanisms underlying schizophrenia remain largely unknown, a convergence of pharmacologic and genetic data implicates a dysregulation of NMDAR function. NMDARs play critical roles in neurodevelopment and synaptic plasticity and subtle changes in NMDAR functioning can have wide-ranging developmental and cognitive effects. Most forebrain NMDARs contain two GluN1 and two GluN2A or GluN2B subunits, with receptor trafficking and functional properties largely dictated by the GluN2 subunit composition. Until recently the dogma in the field was that NMDARs were relatively immobile fixed structures, though it is now apparent that there is a remarkable plasticity of synaptic NMDARs. Indeed, the expression, trafficking, synaptic localization and functioning of different NMDARs subtypes are under dynamic cellular control, though the mechanisms are poorly understood. In this research plan, the roles of GluN2 subunit C-terminal tails in NMDAR synaptic targeting, trafficking, and regulation will be systematically investigated using an innovative molecular replacement approach in which native NMDARs are removed and replaced by recombinant NMDAR subunits in individual neurons. By combining his training in molecular and cellular biology, receptor pharmacology, and synaptic electrophysiology, Dr. Gray will pursue additional training in biochemical and proteomic analysis to address the following specific aims: 1) to determine the role of GluN2B-S1480 phosphorylation in NMDAR trafficking; 2) to determine the mechanism of GluN2A targeting to synapses; and 3) to determine the role of tyrosine phosphorylation in NMDAR trafficking and regulation. Successful completion of this proposal will identify novel trafficking mechanisms, new targeting motifs and new proteins important in NMDAR regulation and synaptic development and plasticity and will open new frontiers for the development of disease-modifying therapeutic approaches for schizophrenia and other neuropsychiatric disorders.
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2019 — 2020 |
Gray, John Alan Gray, John Alan [⬀] |
R21Activity Code Description: To encourage the development of new research activities in categorical program areas. (Support generally is restricted in level of support and in time.) |
The Role of Nmda Receptor Co-Agonist Site Occupancy in Synaptic Plasticity @ University of California At Davis
The brain is made up of billions of neurons that connect via trillions of synapses, the chemical junction between two neurons. Proper development and regulation of these synapses is crucial for the proper functioning of the brain. Indeed, synaptic dysfunction is thought to be the primary basis of many brain diseases, including Alzheimer?s disease, schizophrenia, autism, epilepsy, addiction, and chronic pain. In the brain, the majority of synapses important for learning and memory are chemically stimulated by glutamate. Glutamate activates a specific protein known as the N-methyl-D-aspartate (NMDA) receptor that plays an essential role in proper brain development and synapse functioning. However, two fundamental properties of the NMDA receptor remain elusive; 1) why do NMDA receptors require two chemical signals, glutamate and glycine, for activation? And 2) how can NMDA receptors regulate both the strengthening and weakening of synapses during learning and memory? Understanding these two critical functions of NMDA receptors, in addition to contributing to our basic knowledge of neurobiology, may also point to a therapeutic strategy that will directly target and possibly even reverse the core synaptic changes that cause brain disease states such as addiction and chronic pain. The goal of this proposal is to understand how NMDA receptor co-agonism influences synaptic plasticity. The central hypothesis is that co-agonist site occupancy dictates the directionality of synaptic plasticity. Recent evidence by myself and others has shown that long-term depression (LTD) does not actually require ion flow through the NMDA receptor channel, but requires only glutamate binding. These results are contrary to the long-standing view that long-term potentiation (LTP) was due to rapid, large levels of calcium influx through the receptor, and LTD was mediated by low level, repetitive increases in calcium. Instead, these new results suggest that NMDA receptors invoke intracellular signaling solely due to conformational changes upon agonist binding. Building upon these new findings, I have developed a model that predicts a fundamental role of glycine/D-serine site occupancy as a direct regulator of the directionality of synaptic plasticity. This model allows for predictions that can be rigorously tested with pharmacological approaches in slice electrophysiology (Aim 1) and the single-cell genetic manipulations (Aim 2) regularly used in my laboratory. Validation of this model will open a new frontier of NMDA receptor and synapse biology that will have a significant impact on many areas of neuroscience and may lead to the development of novel approaches to modify synaptic plasticity for the treatment of neuropsychiatric diseases.
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2019 — 2021 |
Gray, John Alan [⬀] Gray, John Alan [⬀] |
R01Activity Code Description: To support a discrete, specified, circumscribed project to be performed by the named investigator(s) in an area representing his or her specific interest and competencies. |
Function and Regulation of Postsynaptic Serine Racemase @ University of California At Davis
Though etiological mechanisms underlying schizophrenia remain largely unknown, a convergence of pharmacologic and genetic data implicates a dysregulation of N-methyl-D-aspartate receptor (NMDAR) function. NMDARs are ligand-gated ion channels that play critical roles in neurodevelopment and synaptic plasticity and subtle changes in NMDAR functioning can have wide-ranging developmental and cognitive effects. Thus, there is a critical need for a detailed understanding of the molecular mechanisms involved in the regulation of NMDARs which will yield insights into the pathophysiology of schizophrenia and facilitate the development of novel therapeutic strategies. Unlike all other neurotransmitter receptors, NMDARs have an absolute requirement for the binding of two different agonists in order to be activated. In addition to glutamate released from the presynaptic terminal, NMDARs require a co-agonist, which can be either glycine or D-serine. At most forebrain synapses, D-serine is the primary NMDAR co-agonist and D-serine deficiency has been implicated in the pathophysiology of schizophrenia. However, our understanding of the mechanisms regulating the availability of synaptic D-serine remains quite limited. Even the cellular source of D-serine is fiercely debated. D-serine is synthesized in the brain by the enzyme serine racemase (SR) that converts L-serine to D-serine. Original studies placed both SR and D-serine in astrocytes, and activity-dependent release of D-serine from astrocytes was venerated as the prototypical gliotransmitter. However, a growing literature using more selective antibodies and mouse genetics has challenged this astrocytic role in D-serine regulation and pointed to a primarily neuronal source. In this proposal, conditional SR knock-out mice were injected with viral constructs into the hippocampus to remove SR in only a few individual neurons. By comparing the effects of SR deletion to adjacent control neurons, we have obtained preliminary data that provides the first rigorously- controlled evidence for a functional role of neuronal D-serine. This preliminary data supports our central hypothesis that synaptic NMDAR activity and function is regulated by postsynaptic SR and D-serine release. In Specific Aim 1, the cell-autonomous effects of postsynaptic SR deletion will be characterized and the source of the remaining synaptic co-agonist will be determined. In Specific Aim 2, we will examine in detail the effects of postsynaptic SR deletion on both postsynaptic and presynaptic NMDARs. In Specific Aim 3, we will determine the functional role of postsynaptic SR, using an innovative molecular replacement approach to replace endogenous SR with characterized recombinant SR mutants to explore the function and regulation of postsynaptic SR. Successful completion of this research program will provide a better understanding of the molecular mechanisms regulating co-agonist availability at synaptic NMDARs, and how these processes might be disrupted in disease states, helping to lead towards our long-term goal of developing novel disease-modifying approaches to treat schizophrenia and other complex neuropsychiatric disorders.
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