A major challenge for the advance of biology and medicine is to develop new ways of studying how proteins function in complex networks in cells and how cellular circuits operate to coordinate functions of the organism. Progress here requires molecularly focused methods for dynamic detection and manipulation that can be used in the living organism. An attractive approach is to use light as both input and output to probe signaling proteins in vivo. The UC Berkeley Nanomedicine Development Center for the Optical Control of Biological Function has been at the forefront of re-engineering proteins to be sensitive to light so that they can be rapidly switched on and off in select cells in vivo. We have developed light-gated ion channels and receptors and used these in cultured neurons, retina, and in vivo zebrafish and in the rodent eye. The development of optically controlled proteins opens the door for applications in medical research, including understanding the function and development of neural circuits, and their relation to behavior, and to re-engineering native cells to switch their state in response to light-a control that could be useful in cell replacement therapies and which could help restore vision in animal models of certain blinding diseases. For applications in both cell culture and in vivo, we require the ability to pinpoint optical stimulation in a microscope that can image fluorescent indicators of neural activity at high resolution. The optical stimulation needs to be spatially and temporally flexible, but at the same time well-focused in 3D and of sufficient intensity to enable us to mimic physiological patterns of activity in specific neurons, in specific regions of the brain, in some cases during behavioral assays. We request funds for an Optical Stimulation Microscope, which combines the high resolution high quality imaging of the Olympus FV1000 microscope with a novel approach to optical stimulation that uses LCOS-SLMs for 1P and 2P to achieve illumination that is shaped in 3D and which can stimulate structures as small as a piece of a dendrite, as large as a whole cell or nearby group of cells and ranging up to cells that are dispersed in the field of view. This unique Optical Stimulation Microscope will fill a void at UC Berkeley. The Optical Stimulation Microscope will enable us to study the development and function of neural circuits, the integration into the adult brain of transplanted neurons for cell replacement therapy and the restoration of light sensitivity to retinas that have lost their photoreceptor cells in attempts to restore vision to models of blindness. PHS 398/2590 (Rev. 09/04, Reissued 4/2006) Page 1 Continuation Format Page PUBLIC HEALTH RELEVANCE: A major challenge for the advance of biology and medicine is to develop new ways of probing proteins in intact cellular circuits to learn how they operate and to repair their function. We have re-engineered proteins to be sensitive to light so that they can be remote-controlled and request funding for a unique Optical Stimulation Microscope that can point light to selectively manipulate the activity of select neurons in the living organism. The Optical Stimulation Microscope will enable us to study the development and function of neural circuits, the integration into the adult brain of transplanted neurons for cell replacement therapy and the restoration of light sensitivity to retinas that have lost their photoreceptor cells in attempts to restore vision to models of blindness.