The N-methyl-D-aspartate receptor (NMDAR) has been extensively studied due to its critical role in activity-dependent synaptic plasticity and excitotoxicity. Defining the signaling cascades regulated by the NMDAR is essential for a mechanistic understanding of NMDAR function, but the intracellular signals initiated by NMDARs are not well characterized. Our laboratory has undertaken a systematic analysis of the proteins associated with the cytoplasmic tails of NMDARs with the rationale that such proteins would be ideally positioned to transduce calcium influx through the NMDAR channel to downstream signaling pathways. The multidomain protein PSD-95 appears to be an organizing center for these transducing elements, binding to NMDARs as well as to signaling molecules such as two GTPase activating proteins (GAPs). One of these, SynGAP, is a recently described Ras-specific GAP; the other, SpanGAP, is a novel GAP likely to regulate Rap small GTPases. This proposal will focus on the potential role of SpanGAP and Rap in NMDAR signaling, examining the relative contribution of this pathway (compared with the Ras pathway) in the NMDAR signaling cascade. Dominant negative mutants and antisense strategies will be employed to dissect specific transduction mechanisms in cultured primary neurons. Effects will be analyzed by using a variety of transcriptional reporters as well as phospho-specific antibodies against MAP kinases and CREB. The results of this study may shed light on novel mechanisms contributing to NMDAR-mediated synaptic plasticity and excitotoxicity.
