NMDA receptors have been implicated in the induction of various forms of synaptic plasticity including processes that may underlie some types of learning and memory. Excessive activation of NMDA receptors is in part responsible for neurodegeneration that occurs after ischemia or brain trauma. These receptors may also play a role in the development and occurrence of seizure activity, in chronic neurodegenerative diseases, and in psychoses. NMDA receptors are targets for neuroprotective and anticonvulsant drugs. The receptors are activated by glutamate and the co-agonist glycine. In addition, NMDA receptors contain recognition sites for a large number of endogenous modulators and non-competitive antagonists. Studies in our laboratory have focused on-the modulation of NMDA receptors by the endogenous polyamine spermine, which has a number of macroscopic effects on the receptor that may involve three discrete polyamine binding sites. The properties of ifenprodil have also been studied. Ifenprodil is a novel noncompetitive antagonist that discriminates subtypes of NMDA receptors and does not share the in vivo side effects associated with traditional NMDA antagonists. The properties of recombinant NMDA receptors are dependent on their subunit composition. The binding sites for many of the modulators, including polyamines, appear to be located on the NR1 subunit. In addition, multiple properties of the receptor including sensitivity to polyamines, glycine, histamine, ifenprodil, and Mg2+ are influenced by the type of NR2 subunit present in NR1/NR2 receptors. The goals of the present proposal are to identify amino acid residues in the NR1 and NR2 subunits that form binding sites for polyamines, to identify the structural features and amino acid residues in NR2 subunits that control sensitivity to various modulators, to determine the role of carbohydrate moieties in modulation by polyamines and other agents, and to investigate the properties of receptors containing two different types of NR2 subunits. These studies will involve site-directed mutagenesis of NR1 subunits, construction of chimeric NR2 subunits, and studies of effects of lectins, together with voltage-clamp recording of NMDA receptors expressed in Xenopus oocytes. The mutagenesis studies will be centered around regions of the NR1 subunit that show homology with bacterial polyamine binding proteins. Studies of chimeric NR2 subunits will initially involve NR2A and NR2B but will be expanded to include subunits containing portions of NR2C and NR2D. The proposed studies are designed to increase our knowledge of the properties and regulation of glutamate receptors and their roles in synaptic transmission and plasticity. This may ultimately contribute to our understanding of some neurological diseases and the work may have implications for the design and use of novel pharmaceuticals targeted to NMDA receptors to treat central nervous system disorders.
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