The role of nitric oxide signaling in synaptic plasticity

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Learning and memory has at its core modifications to tiny spine like structures that exist on a special subset of neurons in the brain known as "spiny neurons." These spines must move, adapt, grow and retract in response to chemical neurotransmitters in the brain, a process that as a whole is believed to be the cellular manifestation of learning, known as synaptic plasticity. The spines themselves are thought to serve as basic units of memory storage. While much work has focused directly on the chemical signals that mediate communication in the brain and the ion channels that are opened as a result, little is know of the specific signaling pathways that control the actual generation, shape and loss of individual spines. This proposal aims to shed light on this process by focusing on a specific molecule (Nitric Oxide) that is produced in these spines following ion channel opening. Nitric Oxide is unique in that it can directly alter the proteins responsible for dynamic changes to spine morphology. This proposal is driven by the hypothesis that Nitric Oxide signaling regulates synaptic plasticity by altering the density and morphology of spines through a process called S-nitrosylation.**Ion channels link a cell with the environment that surrounds it. These channels allow the cell to interpret its surroundings by responding to neurotransmitters and propagating signals from the environment outside of the cell inward. These signals mediate changes in the structure and shape of the cell itself and facilitate spine growth and retraction. One class of ion channel of interest to this proposal is characterized by its ability to respond to a neurotransmitter called glutamate. Glutamate signaling can increase the efficiency of communication between cells in the brain by both increasing the number and stability of new spines as well by pruning excess or unnecessary spines, leaving a more efficient communication network. Glutamate-mediated opening of the "NMDA" type ion channel results in production of Nitric Oxide that modifies the proteins in spines through a chemical reaction called S-Nitrosylation. This event alters the function of near-by proteins and as a result the dynamic properties of the spines they inhabit. While it is estimated that 50% of proteins in the brain are altered by S-nitrosylation, a comprehensive analysis of the role it plays in synaptic plasticity has never been performed.**Using powerful mouse genetics we will control the level of Nitric Oxide synthesized in spines by altering the genetic composition of the NMDA-type ion channel and thus its ability to open and close. Using live imaging microscopy techniques coupled with fluorescent probes, we will monitor the flux of ions through NMDA-type channels as well the amount Nitric Oxide subsequently produced in cultured brain slices and isolated neurons from these mice. This will allow us to determine how the level of Nitric Oxide correlates with dynamic changes to spine morphology. We will then monitor what proteins are S-nitrosylated by Nitric Oxide under conditions of spine growth, spine stabilization and spine retraction using Mass Spectrometry to analyze changes to proteins composition. This will form the basis of a novel molecular pathway that underlies the processes of synaptic plasticity. Moreover, this research program will further our understanding neural architecture and the means by which cells of the brain communicate.