Software and hardware to study brain function during behavior using laser scanning microscopy.

The past decade has seen major advances in the tools available to neuroscientistsmaking it possible to ask increasingly specific questions regarding which neurons and circuits are correlated withnecessary forand sufficient forspecific behavioral or computational functionsTwo photon laser scanning microscopyTPLSMis a widespread tool that allows three dimensional imaging and photo stimulation deep within intact brain tissue at neuron resolutionNumerous microscope manufacturers supply turnkey TPLSM systems that provide basic functionality for controlling the microscope during experimentsbut no commercially available system allows the intricate experiments required to test the complex hypotheses of interest in systems neuroscience todayVidrio s mission is to address this gap with its software productScanImagein a way that is stableflexibleand of commercial qualityExperimentalists need to performclosed loopexperiments in which salient neurons are selected for temporally specific manipulation based on ongoing experimental resultsSuch capabilities are necessary because it is difficult to predict in advance which neurons will be engaged by the function under study and because neuronsinvolvement in computations are often restricted to specific phases of a taskThis functionality requires software new scanning modes with real time analysis and controlIn our Phaseapplication we laid out a complete software development plan for enabling closed loop TPLSM experimentsWe demonstrated the feasibility of our approach in our Phaseworkwhich comprised integrating advanced laser scanning capabilities with online CPU based and FPGA based data processingHere we propose to complete our plan in Phaseby implementing the remaining key features needed for the next generation of systems neuroscience experimentsFirstit is critical to be able to image one set of cells while simultaneously manipulating another setinDIn Phaseworkwe added control for two laser scanning paths to achieve X Y independencehere in phasewe will add Z control to achieveD targeting withum accuracySecondwe will implement algorithms to enable interactive identification and motioncompensated targeting of salient neuronsby capitalizing on the high levels of data parallelism achieved by the CPU and FPGA implementations in PhaseworkPresenting researchers with ongoing maps of the correlations between neuronal activity in the field of view and their own quantitative definition of salience will allow them to rapidly select neurons for manipulationThirdwe will make this functionality easily accessible by designing and fabricating a reasonably priced ScanImage optimized computer hardware device that integrates microscope controldata acquisitionand real time image analysisSuccessful completion of this project will enable Vidrio to market ScanImage in a financially self sustaining mannerthus empowering researchers to elucidate how functionally defined subpopulations of neurons mediate specific information processing functions at key moments during behaviorin healthy animals and in animal models of neurological diseases By enabling researchers to measure and manipulate neural tissuetwo photon laser scanning microscopyTPLSMhas become an indispensable tool for neurobiologyused by hundreds of research laboratories to study normal brain function and brain disordersOur TPLSM control softwareScanImagehas become widely adopted by researchers wanting to exert finer control over the microscope in order to conduct cutting edge experiments for testing advanced hypotheses regarding brain functionHere we propose to build on successful Phasework to enhance ScanImage such that researchers canmeasure and manipulate large numbers of individual neurons with unprecedented specificity and independence inDperform real time analyses and automatically motion correctandpurchase ScanImage as part of an integrated optimized software hardware platform