Molecular Mechanisms of Neurotransmitter Release: Analysis of in Vivo Protein Interactions in Synaptic Vesicle Trafficking

Investigating the molecular mechanisms of synaptic function within a single neuron is challenging because of the small size of the cells and molecules of the nervous system. However the development of sophisticated microscopy and imaging methods are making it possible to overcome this obstacle. In this project, in vivo imaging of fluorescently labeled proteins will be employed to examine the protein-protein interactions and molecular mechanisms by which electrical signals are transmitted among neurons. The studies will be done in Drosophila melanogaster, because the functional mechanisms of synaptic transmission in the fruit fly are similar to those of vertebrates. The project integrates two powerful approaches to examine how electrical signals are transmitted among neurons: analysis of electrical signaling in the nervous system and imaging of fluorescently labeled proteins. Drosophila strains expressing appropriate pairs of fluorescently labeled versions of proteins that function at synapses will be analyzed for binding interactions in single living cells. The interactions will be detected using Fluorescence Resonance Energy Transfer (FRET). Electrophysiological analysis of synaptic transmission in the transgenic fly strains will be used to confirm that they are functioning normally.

This project will have a broad impact. It will engage a group of young investigators from several cultures in a collaborative research and training program requiring a high level of intellectual interaction. The project will provide opportunities for scientists at all stages, from senior research associate to undergraduate, and will promote the development of women and minorities in science. Many research materials generated thus far have already been disseminated for related research applications in laboratories throughout the world. Finally, the materials and methods developed in this project are integrated into an advanced cellular and molecular neuroscience laboratory course providing undergraduates with intensive exposure to current neuroscience research methods.