A central goal of neuroscience is to understand how animal and human behaviors are generated by the circuits of the brain. Progress towards this goal has relied in part on the ability to record the activity of neuronal populations. To record densely from a local circuit, one of the most widely-used techniques is calcium imaging. Calcium imaging by two-photon microscopy currently allows investigators to record from hundreds of neurons simultaneously;while a major improvement over past approaches, it is nevertheless a small fraction of the total number of neurons in almost any local circuit. The difficulty of determining which neurons first compute a particular """"""""decision"""""""" or pattern of activity is, in my view, the most important technical barrier to a comprehensive understanding of how neuronal circuits drive behavior. Recently, we developed a method, called Objective-Coupled Planar Illumination (OCPI) microscopy, to perform fast fluorescence imaging of whole tissue volumes. Instead of collecting data from one pixel at a time, it uses a thin sheet of light to collect entir images at once. OCPI microscopy has allowed us to record from approximately ten thousand neurons simultaneously at high speeds and signal-to-noise ratio. For our scientific work on the olfactory system, OCPI microscopy has revealed what the sensory periphery detects, how chemical stimuli are encoded, the spatial arrangement of circuits in the brain, and even how different individuals perceive the world. Here I propose to extend the domain of applicability of OCPI microscopy.
In aim 1, we will develop new methods to see deeper into the living brain.
In aim 2, we will develop procedures to """"""""tag"""""""" individual neurons based on their physiological properties.
In aim 3, we will develop and share algorithms to process the terabyte-sized datasets produced by OCPI microscopy.
These aims will allow OCPI microscopy to address new questions and to be disseminated widely throughout the neuroscience community.
The mechanisms of most neurological diseases, and the drugs used to treat them, are poorly understood. This application proposes new imaging technologies allowing whole circuits or brains to be watched in action at single-cell resolution. These tools will shed new light on the fundamental circuit mechanisms of disease.
|Xu, Pei Sabrina; Lee, Donghoon; Holy, Timothy E (2016) Experience-Dependent Plasticity Drives Individual Differences in Pheromone-Sensing Neurons. Neuron 91:878-92|
|Liang, Xitong; Holy, Timothy E; Taghert, Paul H (2016) Synchronous Drosophila circadian pacemakers display nonsynchronous CaÂ²âº rhythms in vivo. Science 351:976-81|
|Fu, Xiaoyan; Yan, Yuetian; Xu, Pei S et al. (2015) A Molecular Code for Identity in the Vomeronasal System. Cell 163:313-23|
|Hammen, Gary F; Turaga, Diwakar; Holy, Timothy E et al. (2014) Functional organization of glomerular maps in the mouse accessory olfactory bulb. Nat Neurosci 17:953-61|
|Xu, Pei Sabrina; Holy, Timothy E (2013) Whole-mount imaging of responses in mouse vomeronasal neurons. Methods Mol Biol 1068:201-10|
|Tolokh, Illya I; Fu, Xiaoyan; Holy, Timothy E (2013) Reliable sex and strain discrimination in the mouse vomeronasal organ and accessory olfactory bulb. J Neurosci 33:13903-13|
|Turaga, Diwakar; Holy, Timothy E (2012) Organization of vomeronasal sensory coding revealed by fast volumetric calcium imaging. J Neurosci 32:1612-21|
|Turaga, Diwakar; Holy, Timothy E (2010) Image-based calibration of a deformable mirror in wide-field microscopy. Appl Opt 49:2030-40|