One of the central goals of the BRAIN initiative is to develop and disseminate methods for imaging activity in large populations of neurons at high speeds. Of the available techniques, one of the most promising is light sheet microscopy. This technique, first developed for functional neuronal imaging in my laboratory, allows rapid whole-brain imaging in small model organisms and recording from tens of thousands of neurons in explanted tissue from the mouse. In practice, it is approximately one hundred fold faster than other widely-used tools for imaging the brain, while delivering gentle illumination that reduces photodamage to permit long-term recording. The mouse is the most widely-used model organism for investigating neural circuits and the basis of behavior. However, light sheet microscopy has a number of technical limitations which heretofore have prevented its widespread application to studies in the awake behaving mouse. Because the illumination is at a 90? angle relative to the imaging axis, the resulting geometric obstruction has made it difficult to employ light sheet for imaging samples like the surface of the mouse brain. Elaborations of light sheet microscopy can avoid this geometric obstruction, but at the cost of compromising light efficiency at moderate numerical aperture, such as when visualizing large cortical areas or when imaging through a microendoscope in deep brain. Here, I propose a new imaging technique to address these concerns. Based on the principle of ?re-imaging,? we will develop a method to visualize optical planes that are parallel (or near- parallel) to the axis of the microscope objective, nearly orthogonal to the classical plane of focus of a lens system. We will demonstrate this principle in two practical assemblies, one to support volumetric imaging of superficial cortex at high speed, and the other to support deep-brain imaging through a microendoscope.
The mechanisms of most neurological diseases, and the drugs used to treat them, are poorly understood. This application proposes development of new imaging technologies allowing whole circuits to be watched ?in action? at single-cell resolution. This project will demonstrate and document these powerful tools for the use of a large number of investigators, whose work will generate new insights into the mechanisms of disease.
