NMDA receptors are involved in several critical functions of the CNS such as cellular mechanisms of learning, pain perception, motor patterns, experience-dependent synapse formation, and others. These receptors are also involved in epilepsy, narcotic adaptation, and neuronal cell death following ischemia, head and spinal cord injury, and HIV infection. Presently the roles that the various NMDA receptor subtypes play in these diverse actions are unknown. We propose to develop subtype-selective antagonists to facilitate the study of NMDA receptor subtypes in normal and abnormal CNS function. We propose to develop a new category of NMDA receptor antagonist- """"""""cleft-binding"""""""" antagonists. Our previous antagonist development studies, as well as our molecular modeling studies, have lead us to the conclusion that antagonists of greater subtype-selectivity will require large side groups that can interact with the unique amino acid residues that lay outside the primary antagonist binding site. Such antagonists have side-groups that project further out into the cleft formed between the two major lobes (S1 and S2) that comprise the glutamate binding domain of the NMDA receptor. Thus, we are targeting antagonists that are capable of interacting with the more variable regions of the receptor. The objectives of this project are to: 1) Synthesize and evaluate novel NMDA receptor antagonists that define the structural requirements for binding within the cleft domain of the different NR2 subunits. 2) Test our recently developed molecular models of NR2B and NR2C glutamate binding sites by making point mutations and one chimera that are predicted to have specific alterations in antagonist selectivity. 3) The results from novel antagonists (Aim 1) and point mutations (Aim 2) will be used to test, and if necessary, refine our molecular models of NR2B and NR2C glutamate binding sites. We will then construct NR2A and NR2D molecular models. We will then use the refined models, as well as the structure-activity information, to generate new subtype-selective cleft-binding antagonists. This process would include using automated, computer-assisted routines, as well as, visually-aided design. Given the high homology in secondary, but not primary, structure for the various glutamate receptors, we feel this unexplored approach to antagonist design has significant implications for developing subtype selective antagonists within each of the glutamate receptor families.
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