Project Summary To further our understanding of the function of neural circuits, there is a need for new tools that can collect simultaneous measurements from large populations of neurons involved in a common neural computation and provide precise functional modulation. Optical imaging in awake animals expressing calcium or voltage indicators provides real-time functional and spatial information from individual neurons within local neural circuits. The limitations of current imaging technology include small fields of view encompassing single brain regions, and the requirement for head fixation, which prevents naturalistic behavior. In addition, most optical imaging systems do not allow for simultaneous high- resolution functional imaging in combination with spatially-localized optogenetic modulation. To meet this challenge, we propose to develop an optical device (`3D-FAST') that allows for rapid, real- time volumetric neural recording and precise optical stimulation. By pairing miniature arrays of micropatterned LED emitters with the axial focusing capabilities of electrowetting lens technologies, we will achieve duplex recording and stimulation of many thousands of neurons. Through utilization of novel 3D-printed scaffolding, we will be able to create modular, expandable, customizable lens arrays that allow for recording of large-scale bi-directional neural interfaces for closed-loop modulation of neural circuits. We will create two versions of the 3D-FAST device. 3D-FAST-GECI that will provide fast, volumetric imaging at 30 Hz for GCaMP imaging, and 3D-FAST-GEVI that will provide frame rates of 1 kHz at a single plane combined with a slower, high resolution volume scan. Initially, the devices will be tested in the anesthetized, and then awake, freely moving mouse for experiments using GCaMP, voltage indicators ASAP and Voltron, and finally combining with localized optogenetics for closed loop feedback. In sum, these experiments will demonstrate the unique capabilities of the 3D-FAST technology. Rapid, imaging of improved voltage indicators will be paired with spatially-restricted light delivery for optogenetic neural modulation for the first time in a freely moving mouse. The optical imaging properties will be compared with ground-truth two-photon microscopy, and the functional consequence of neuromodulation will be dissected through behavioral assays. The 3D-FAST tool will bring novel capabilities to measuring and modulating large populations of neurons in freely-moving animals, to better understand the neural computations that underlie behavior. In addition, this body of work will lay the ground for future development of fully implantable optical recording and modulating units for use in freely-moving, untethered naturalistic behavior experiments.

Project Narrative Understanding how neural circuits function requires precise spatiotemporal monitoring in intact animals. Optical techniques can provide both spatial and temporal precision, but are limited in by the penetration of light into tissue and the requirement for head fixation, precluding naturalistic movement. We have designed a modular, scalable optical device that uses novel hardware and computational strategies to allow for rapid, volumetric closed-loop neural recording and modulation in freely-moving animals.