The NMDA-type glutamate receptor is implicated in long-term potentiation, memory formation, brain development, and the neurodegeneration associated with epilepsy, ischemia, Huntington's chorea, Alzheimer's disease and AIDS encephalopathy. The proposed research aims to study molecular diversity in neuronal NMDA receptors. Molecular diversity will be addressed at the following levels of resolution; amino acid residues, receptor domains, receptor variants, individual cells, and neural circuits. Experiments outlined in this proposal are directed at the development of a detailed molecular/functional/anatomic profile of NMDA receptor variants. This laboratory has cloned two new NMDA receptor splice variants from rat brain and has characterized their functional properties. The receptor variants differ in their agonist affinity, current amplitudes, and regulation by polyamines, zinc and protein kinase C(PKC). In one project, we will use site-directed mutagenesis to identify functionally important amino acid residues in the NR1 receptor protein. Recombinant NMDA receptor channels will be analyzed in Xenopus oocytes and human embryonic kidney 293 cells by whole cell recording and by patch clamp. Studies will focus on the receptor domains involved in polyamine and zinc potentiation, regulation by PKC and in binding of glycine and glutamate. Working hypotheses are as follows: 1) Positively charged residues within the N terminal insert N1 govern zinc and spermine potentiation, agonist potency, and current amplitude, thereby generating receptors with altered normal responsiveness and sensitivity to glutamate pathogenicity; 2) In NR1 receptors lacking the C1 insert, serine residues within the cytoplasmic loops play a critical role in regulation by PKC; 3) A glycine receptor-like motif within the N terminal domain forms part of the glycine binding site. 4) A glutamate binding protein (QBP)-like domain within the N terminal domain of NR1, just preceding TMI, is involved in the glutamate binding site. In a second project we will characterize NR1 heteromers with normal, mutationally altered and chimeric NR2 receptors for comparison in a situ receptors and to determine structure/activity relationships for this receptor subunit. In a third project we will define cell-specific and circuit-specific expression of NMDA receptor splice variants in the hippocampus, a brain region known to be vulnerable to glutamate toxicity. Individual hippocampal neurons grown in dissociated culture on coverslips will be analyzed electrophysiologically at the whole cell and single channel level for sensitivity to polyamines and to PKC. Subpopulations of NMDA splice variants and subtypes will be identified by in situ hybridization using exon-specific and splice junction-specific oligonucleotide probes. As a future direction, individual neurons will be analyzed for expression of splice variants by patch clamp methods and by the polymerase chain reaction. Findings from these studies are expected to aid in the development of new strategies for intervention in neurodegenerative disorders.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS020752-16
Application #
6539607
Study Section
Special Emphasis Panel (ZRG1-MDCN-4 (01))
Program Officer
Stewart, Randall
Project Start
1984-07-01
Project End
2004-06-30
Budget Start
2002-07-01
Budget End
2003-06-30
Support Year
16
Fiscal Year
2002
Total Cost
$489,949
Indirect Cost
Name
Albert Einstein College of Medicine
Department
Neurosciences
Type
Schools of Medicine
DUNS #
009095365
City
Bronx
State
NY
Country
United States
Zip Code
10461
Udagawa, Tsuyoshi; Swanger, Sharon A; Takeuchi, Koichi et al. (2012) Bidirectional control of mRNA translation and synaptic plasticity by the cytoplasmic polyadenylation complex. Mol Cell 47:253-66
Rodenas-Ruano, Alma; Chávez, Andrés E; Cossio, Maria J et al. (2012) REST-dependent epigenetic remodeling promotes the developmental switch in synaptic NMDA receptors. Nat Neurosci 15:1382-90
Eugenin, E A; King, J E; Hazleton, J E et al. (2011) Differences in NMDA receptor expression during human development determine the response of neurons to HIV-tat-mediated neurotoxicity. Neurotox Res 19:138-48
Nolt, Mark J; Lin, Ying; Hruska, Martin et al. (2011) EphB controls NMDA receptor function and synaptic targeting in a subunit-specific manner. J Neurosci 31:5353-64
Jitsuki, Susumu; Takemoto, Kiwamu; Kawasaki, Taisuke et al. (2011) Serotonin mediates cross-modal reorganization of cortical circuits. Neuron 69:780-92
Wang, Dan Ohtan; Martin, Kelsey C; Zukin, R Suzanne (2010) Spatially restricting gene expression by local translation at synapses. Trends Neurosci 33:173-82
Lau, C Geoffrey; Takayasu, Yukihiro; Rodenas-Ruano, Alma et al. (2010) SNAP-25 is a target of protein kinase C phosphorylation critical to NMDA receptor trafficking. J Neurosci 30:242-54
Sharma, Ali; Hoeffer, Charles A; Takayasu, Yukihiro et al. (2010) Dysregulation of mTOR signaling in fragile X syndrome. J Neurosci 30:694-702
Takayasu, Yukihiro; Takeuchi, Koichi; Kumari, Ranju et al. (2010) Caveolin-1 knockout mice exhibit impaired induction of mGluR-dependent long-term depression at CA3-CA1 synapses. Proc Natl Acad Sci U S A 107:21778-83
Philpot, Benjamin D; Zukin, R Suzanne (2010) Synapse-specific metaplasticity: to be silenced is not to silence 2B. Neuron 66:814-6

Showing the most recent 10 out of 71 publications