Activity-dependent ciianges in synaptic strengtli at glutamatergic synapses are tiiougiit to contribute to tiie development of neural circuitry and many forms of experience-dependent plasticity, including learning and memory. The hippocampus, a major site of synaptic plasticity, plays a fundamental role in some forms of learning and memory, and has been implicated in a number of neurological and psychiatric disorders, including depression, epilepsy, Alzheimer's disease, and schizophrenia. In this proposal, I outline a series of experiments that will test the functional significance of key synaptic scaffolding proteins in regulating glutamate receptor function and synaptic plasticity at hippocampal Schaffer collateral-CAl synapses. This will involve making simultaneous whole cell patch clamp recordings from neurons in organotypic hippocampal slice cultures that haven been molecularly modified using lentiviral-mediated gene knockdown via shRNA and simultaneous lentiviral-mediated gene transfer. The bicistronic lentiviral vector I will use allows expression of mutant forms of a protein on the background of acute knockdown of the endogenous protein. I will specifically focus on the function of the postsynaptic scaffolding proteins of the family of the disc-large (DLG) membrane-associated guanylate kinases (MAGUKs) and their interacting partners. My previous results demonstrate that two family members of DLG-MAGUKs, PSD-95 and SAP97, regulate synaptic AMPAR function differently in terms of their activity-dependence. During the K99 training period, I found that the effects of PSD-95 on basal transmission and long-term depression are dissociable. The N-terminal domain of PSD-95 is required for dimerization and appropriate synaptic enrichment of PSD-95 but alone does not influence synaptic function. The C-terminal portion of PSD-95 serves a dual function. It is required to localize PSD-95 at the synapse and as a scaffold for critical downstream signaling proteins that are required for LTD. The specific objectives of my independent research are: (1) to analyze the signaling scaffold that is important for long-term depression (LTD), in particular, the role of the A-kinase anchoring protein 79/150 (AKAP79/150), and (2) to examine the interaction of PSD-95 with transmembrane AMPA receptor regulatory proteins (TARPs) in mediating long-term potentiation (LTP), (3) to examine the role of SAP97 in mediating LTP. Results from these experiments will begin to elucidate how dynamic interactions among different components of the postsynaptic density influence synaptic function and will address fundamental questions about how signaling specificity is achieved during different forms of synaptic plasticity
My proposal involves studying the detailed molecular mechanisms that underlie synaptic function and plasticity in the hippocampus, a brain structure that is critically important for learning and memory and one which plays an important role in a variety of brain disorders including schizophrenia, depression, Alzheimer's disease and epilepsy. Thus the proposal has great relevance to our understanding of brain processes that primarily fall under the purview of NIMH.
