Next generation high-throughput random access imaging, in vivo

点击翻译按钮获取中文摘要

 DESCRIPTION (provided by applicant): The goal of this proposal is to develop new methods for high speed monitoring of sensory-driven synaptic activity across all inputs to single living neurons in the context of the intact cerebral cortex. Although our focus is on understanding how synaptic inputs are integrated across a single neuron embedded in an intact circuit, the next generation random access imaging technology we propose is more broadly applicable for monitoring multi-cellular activity representing large intra-and inter areal neuronal networks. The approach improves on the speed and sensitivity of current random-access technology by nearly 2 orders of magnitude, enabling high- throughput interrogation of up to 104 independent locations within a fraction of a millisecond and compatible with imaging using next generation voltage sensitive indicators. In Aim 1 we propose to generate a comprehensive structural map that will allow random access scanning of all excitatory and inhibitory synapses on functionally defined pyramidal cell types expressing a genetically encoded Ca+2 indicator. The data generated in this Aim will be used to develop image segmentation algorithms to quickly convert structural images of the dendritic tree and the associated synapses into a 3D location map with grid coordinates for sparse sampling of activity patterns at known locations using a fast random access imaging approach described in Aim 2. In Aim 2 we will construct and develop an imaging system allowing high throughput, random addressing within 10-100 ms of approximately 10,000 locations corresponding to all excitatory synapses and other functionally relevant dendritic and somal sites on a single neuron. In Aim 3 we will test and validate the utility of our approach.