Mechanisms of Excitatory Synapse Formation in Vitro

Johns Hopkins University, Baltimore MD

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Abstract

DESCRIPTION (provided by applicant): The formation and maintenance of excitatory synapses in the central nervous system is crucial to the development of the brain as well as to the ongoing plasticity that underlies learning, memory and adaptation. The neuronal pentraxin Narp (Neuronal Activity Regulated Pentraxin) has been implicated in the aggregation of AMPA type glutamate receptors at excitatory synapses in the brain and spinal cord because exogenously applied Narp can cluster AMPA type glutamate receptors on cultured neurons. Narp is especially intriguing for neuronal function because its expression is tightly regulated by electrical (synaptic) activity. Therefore Narp serves as a potential link between brain activity and synapse formation. Such a link may well be impaired in diseases that affect the formation of new memories such as Alzheimer's disease. In the present proposal, we examine the role and mode of action of endogenous Narp and its related molecule NP1 in excitatory synapse formation by utilizing novel, dominant negative Narp mutants (dnNarp) that selectively bind endogenous Narp and prevent its accumulation at synapses. Neurons transfected with these mutants are deficient in excitatory but not inhibitory synapse formation. In addition we have developed techniques to isolate and characterize the receptor to which Narp binds at excitatory synapses and localizes its function. One possible candidate for this receptor is another activity regulated molecule termed Neuritin. The molecular basis for the interaction between Narp and its receptor and Narp and the AMPA class of glutamate receptors will be investigated using a series of deletion and point mutations in both HEK 293 cells and neurons. Finally the mechanism whereby the NMDA class of glutamate receptors is aggregated at excitatory synapses will be investigated. Our experiments indicate that both Narp and an as yet uncharacterized molecule are involved in this process. The mechanism of Narp's involvement in NMDA receptor aggregation and the identity of any additional molecules involved will be sought.