This BRAIN EAGER will support an interdisciplinary team of investigators to jointly develop and deploy optimized light sheet microscopy and novel genetically encoded probes to image synaptic function in the intact nervous system. This team will optimize a novel 3D imaging microscope, in which multi-photon light sheet illumination is combined with light field microscopy to permit a single snapshot to capture the full 3D image, enabling the team to visualize these probes with unprecedented speed and coverage. They will also develop new genetically encoded glutamate and calcium probes, targeted to defined synaptic compartments, to optimize signal magnitude and report synaptic activity with high sensitivity and fidelity. These innovations will be exploited to synergistically visualize synaptic structure and monitor glutamate and calcium dynamics in the intact Drosophila central nervous system.
Although these tools could be used in a variety of settings in the vertebrate or the invertebrate nervous system, these will be first applied to address the fundamental relationship between sleep and synaptic plasticity. Although sleep is ancient, the essential biological function of this behavior remains a great mystery of science. This project will explore the exciting possibility that a fundamental function of sleep, operating at the level of individual neurons and synapses, is the homeostatic modulation of synaptic strength. Addressing this hypothesis has been beyond our capabilities because visualizing neural activity in the central nervous system during sleep-wake behavior has been limited in both speed and resolution. Through a combination of new genetically encoded probes reporting synaptic structure and activity and cutting-edge imaging approaches, this project will permit the imaging of synapses over time without perturbing the nervous system or the sleep-wake cycle. These test experiments will advance our knowledge of the complex, fundamental, and poorly understood signaling systems that orchestrate the homeostatic control of synaptic strength, and their modulation during sleep behavior. The education and outreach activities of the research team will be intimately linked with their research programs, and will include a research project with local inner-city Los Angeles high school students investigating sleep and circadian behavior. In addition, a new undergraduate course will be developed exploring the biological functions of sleep.
