The Bezos Center for Neural Circuit Dynamics focuses on the development and application of microscopy imaging techniques for measuring neural circuit dynamics in the functioning brain. The Bezos Center for Neural Circuit Dynamics was created by a generous gift from Jeff ’86 and MacKenzie '92 Bezos. Neural dynamics refers to the patterns of nerve cell electrical and chemical activity in which information is created, manipulated, and stored. The focus of the Bezos Center for Neural Circuit Dynamics is the development and application of microscopy imaging techniques for measuring neural circuit dynamics in the functioning brain. The Center hosts both commercial and custom-built optical instrumentation for large-scale monitoring and optogenetic perturbation of neural activity. The Bezos Center is a critical asset for Princeton researchers working toward a circuit-level understanding of computation in the nervous system. Directors David Tank - Scientific Director Stephan Thiberge - Facilities Director Faculty Research Supported by The Bezos Center Michael Berry Neural computation in the retina Carlos Brody Quantitative approaches to systems neuroscience Tim Bushman Dynamics of executive function Annegret Falkner Neural circuits for social decision-making Carolyn (Lindy) McBride Genes and neural circuits underlying behavioral evolution Mala Murthy Neural Mechanisms Underlying Acoustic Communication H. Sebastian Seung Connectivity of neurons in biological tissue David Tank Measurement and analysis of neural circuit dynamics Samuel Wang Learning rules and design principles in neural circuits Ilana Witten Neural circuits for reward learning and decision making Bezos Center Resources Bezos Center Equipment Two large FOV 2P microscopes with 2-axis rotating objective. These instruments were custom designed using commercially available optical elements and use resonant scanners to achieve 30 Hz frame rate imaging. They have a 1.2mm diameter flat field of view. The first microscope is dedicated to rat imaging, the second to other species. The later microscope has 2 modes of operation. One is the conventional 2Photon scanning approach. In this mode, the laser used is a Coherent Chameleon, In this mode the axial resolution is below 5 microns (920nm) across the FOV. In the second mode of operation, a White Dwarf laser at 5MHz and 960nm is used to implement the Light Bead Microscopy method. With this approach, up to 28 beams equally spaced along a 500um depth in the neural tissue are generated to record simultaneously the activity of all neurons within a volume of 600um in width, 1200um in length and 500um in depth at 30Hz. A Mesoscope, based on the design developed at Janelia, HHMI, with random access scanning across a 5mm diameter field of view. A 3P excitation microscope. This microscope uses a wavelength tunable Coherent Laser system (Monaco pumps an OperaF) with 1 MHz pulses at 1300 nm or 1600nm. A system for combined optical imaging and optogenetic stimulation at cellular resolution. This instrument employs separate lasers for cellular resolution 2P excitation at 1064 nm (Carbide & Orpheus-HP, Light Conversion), and simultaneous 2P excitation imaging at 920 nm (Chameleon Vision II, Coherent). All of the above imaging systems are combined with either mouse virtual reality or rat voluntary head fixation systems for head-restrained behavior during calcium imaging. The Bezos Center also includes an electronic bench, a small machine shop containing a bandsaw, a drill and mill, a diamond saw, and a laser cutter. High-throughput Transmission Electron Microscope Facility The High-throughput Transmission Electron Microscope Facility hosts four of the world’s fastest transmission electron microscopes (TEMs). Each of the four installed TEMs consists of a JEOL 1200EX II refurbished by TSS Microscopy, an automated reel-to-reel section feed by Voxa Inc (GridStageTM), a Beam deflector system by Voxa Inc (CricketTM), an Acquisition software by Voxa Inc (BladeTM) with a browser-based GUI and a Python-scriptable backend, and an AMT NanoSprint50M-AV 50 Megapixel research-grade camera system with integrated scintillator and lens system. What's New Bezos Center Light Bead Microscope In 2022, the Bezos Center initiated an application to the DOF Transformative Equipment Initiative for the creation of a Light-Bead microscope, a microscope designed by Alipasha Vaziri (Rockefeller University). The project was funded and allowed the purchase of the low repetition rate laser typically used in this type of approach. The custom elements of the microscope were received in early 2023, and the instrument completion was done in the spring.The instrument includes up to 28 beams equally spaced along a 500um depth in the neural tissue. In the current implementation of the light bead method, the instrument can record the activity of all neurons within a volume of 600um in width, 1200um in length and 500um in depth at 30Hz. Bezos Center Three-Photon microscope improvements using adaptive optics The Bezos Center has been rebuilding the 3-Photon microscope optical path to incorporate a deformable mirror in order to implement wavefront correction of the optical aberrations caused by the brain tissue when imaging at high depth. This method has allowed us to improve the signal by a factor of 2 to 5, depending on the location.