All-optical electrophysiology in brain slice for drastically increased throughput

Project Summary Severe psychiatric disordersincluding schizophreniabipolar disorderand major depressive disorder are prevalentchronic and debilitating brain diseases that result in intense suffering and reduced quality of lifeThere is a profound lack of new medicines for psychiatric disorderswhich remain a major area of unmet medical needDiscovery of more effective therapies for mental illness has been limited by poor understanding of disease biologya lack of model systems that can predict clinical resultsand the dearth of tools to rapidly study relevant electricalsynapticand circuit level phenotypes that underlie many psychiatric disordersIn this proposalwe aim to address these challengesOne powerful platform for studying brain function in intact neural circuits is rodent brain slicecells buried deep in the brain can be studiedbut maintain many of their native connections and propertiesHoweverit has been challenging to comprehensively analyze neuronal properties and connectivity in brain slicessince traditional methodology based on patch clampthe piercing of cells with microelectrodesis extremely labor intensive and lacks sufficient neuronal throughputA promising solution to this problem is optogeneticswherein which light is used to stimulate and record neuronal activityThe Optopatch platform recently developed at Q State Biosciencescomprised of engineered optogenetic proteinscustom microscopesand softwareenables simultaneous stimulationblue lightand recordingred lightof electrical activity fromneurons in a dish with one millisecond temporal resolutionsingle cell spatial resolution and high signal to noise ratioPatterned blue light can be used to stimulate one or more neurons while recording from all synaptic partnersHencethe Optopatch platform can be used to study single cell excitabilitysynaptic transmissionand network behaviorHere we propose to create a custom microscope for harnessing the Optopatch system in brain slices and demonstrate applicability of the system to compound screeningSince background fluorescence and light scattering are many times higher in tissue than in dishes of cultured neuronsa new microscope design is required for high speed voltage imagingWe will employ reconfigurablepatterned illumination to selectively excite a small subset of cellsreducing the background to manageable levels by minimizing the total amount of light deliveredWe will also drive high expression of Optopatch proteins in a small fraction of neurons to obtain large signals with minimal cross talk from neighboring cellsFinallywe will confirm that the system can be used to efficiently determine firing behavior of different types of neurons and measure pharmacological responsesUltimatelythe system we propose to develop has potential to benefit patients with mental illness by enabling newcircuit based therapeutic discovery approaches