Genetic Analysis of Neuromuscular Synaptic Structure and Function

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The central questions that guide my research are: How do synapses function? How do they develop? We are motivated to study these questions to better understand how the nervous system and the brain give rise to behaviors in both health and disease. To address our questions my research team combines classical and molecular genetics in Drosophila melanogaster with developmental analysis and measurements of synaptic physiology. Due to the high degree of conservation at the cellular, genetic and molecular level, the information we generate studying flies translates to a more general understanding of how neurons develop and function. Our basic approach is to use perturb genes in a systematic way and then determine how the synapses respond. The main technical approaches we use are fly genetics electrophysiology of synaptic transmission, imaging technologies, including confocal, live-cell, and electron microscopy; we also do molecular biology and biochemistry as required. NSERC Discovery Grants have continuously supported our exploration within this framework. In this grant, using newly available genetic tools, we will examine the role of developmental competition and co-operation on synaptic physiology. At mammalian NMJs, synaptic competition occurs when more than one neuron innervates a target muscle and subsequent interactions between neurons leads to one `winner' and the withdrawal of the other nerve terminals. However, at arthropod NMJs, including Drosophila, it is normal for two or more functionally specialized motor neurons to innervate the same muscle cell, often in very close proximity; they appear to co-operate to innervate the target cell. We now have the ability to selectively drive gene expression in one or the other of these MNs and ask how the NMJ responds physiologically and developmentally. As one example, we will kill one of the neurons by driving apoptosis-inducing genes and determine how the remaining motor neuron develops and functions in the absence of the partner neuron. In a second set of experiments, we will control electrical excitability using K+ channel expression. Finally, we will target synaptic activity by reducing synaptic transmission in one of the two motor neurons by expressing well known regulators, of the vesicle cycle. In each case, we will characterize the function and morphology of the untreated motor neuron, in the absence of its partner. Based upon preliminary data, we will design a forward screen, to harness the real advantage of Drosophila genetics, to uncover molecules crucial to determining communication between two innervating neurons. Since errors in synaptic development and function are associated with a wide variety of neurological, cognitive and behavioural diseases the proposed experiments will yield new data that will help us understand the basic development and function of synapses; the knowledge gained will further inform our concept of neurodevelopmental and neurophysiological disorders.