The current proposal describes experimental approaches to determine how neurons regulate structural and functional maturation of active zones (AZs), a key signaling hub where synaptic communication occurs. Although membrane trafficking mechanisms are highly conserved across cells, additional synapse-specific regulation has evolved to mediate rapid Ca2+- dependent synaptic vesicle (SV) fusion at specialized presynaptic AZs that are precisely aligned to postsynaptic receptors. Multiple evolutionarily conserved proteins are found at AZs, including RIM, RIM binding protein, Syd-1, Liprin-?, ELKS/CAST/Bruchpilot (BRP), Bassoon/Piccolo/Fife and Unc13. Previous studies in our lab demonstrated that the hundreds of AZs formed by a single glutamatergic motoneuron in Drosophila have a heterogeneous distribution of synaptic strength, with neighboring AZs often showing >50-fold differences in the probability of release (Pr) of SVs. We found that AZ maturation drives increased synaptic strength occur over a multi- day developmental period, with newly formed AZs developing as weak Pr sites before maturing into high Pr AZs through the coordinated accumulation of a core set of proteins. In the current application, we will determine how neurons regulate structural and functional maturation of AZs, and how variations in these processes drive synaptic diversity. The mechanisms regulating AZ maturation fall into two broad categories: those that control cell-wide availability of key building blocks to growing AZs (Aim 1) and those that affect capture and retention of new material at individual AZs (Aim 2). We will determine whether specific AZ proteins are produced and transported in excess of their incorporation into growing AZs, or whether their availability at the synaptic terminal is rate-limiting for AZ maturation. In addition, we will characterize the efficiency of material capture at individual AZs throughout the AZ maturation cycle. Finally, we will examine how material availability and capture differ in tonic and phasic motoneurons that innervate the same postsynaptic muscle, but display striking differences in their AZ organization and SV release properties (Aim 3). These studies will provide new insights into how synaptic strength develops across the cohort of AZs of a neuron, as well as how synaptic diversity can be more broadly controlled across neuronal subclasses.

Public Health Relevance

This research will define how the basic module for information transfer in the brain ? the synaptic active zone ? forms and matures during development. Given that maturation and plasticity of synapses is a fundamental feature of neuronal signaling and circuit remodeling, it is critical to define molecular mechanisms that control this process. Identification of pathways mediating active zone development and maturation will allow analysis of their role in presynaptic plasticity, as well as how disruptions in these components contribute to brain diseases.

National Institute of Health (NIH)
National Institute of Neurological Disorders and Stroke (NINDS)
Research Project (R01)
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Neurodifferentiation, Plasticity, and Regeneration Study Section (NDPR)
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Miller, Daniel L
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Massachusetts Institute of Technology
Schools of Arts and Sciences
United States
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