A Toolbox for Spatiotemporal Analysis of Voltage-Sensitive Dye Imaging Data in Brain Slices

PLOS ONE, Dec 2019

Voltage-sensitive dye imaging (VSDI) can simultaneously monitor the spatiotemporal electrical dynamics of thousands of neurons and is often used to identify functional differences in models of neurological disease. While the chief advantage of VSDI is the ability to record spatiotemporal activity, there are no tools available to visualize and statistically compare activity across the full spatiotemporal range of the VSDI dataset. Investigators commonly analyze only a subset of the data, and a majority of the dataset is routinely excluded from analysis. We have developed a software toolbox that simplifies visual inspection of VSDI data, and permits unaided statistical comparison across spatial and temporal dimensions. First, the three-dimensional VSDI dataset (x,y,time) is geometrically transformed into a two-dimensional spatiotemporal map of activity. Second, statistical comparison between groups is performed using a non-parametric permutation test. The result is a 2D map of all significant differences in both space and time. Here, we used the toolbox to identify functional differences in activity in VSDI data from acute hippocampal slices obtained from epileptic Arx conditional knock-out and control mice. Maps of spatiotemporal activity were produced and analyzed to identify differences in the activity evoked by stimulation of each of two axonal inputs to the hippocampus: the perforant pathway and the temporoammonic pathway. In mutant hippocampal slices, the toolbox identified a widespread decrease in spatiotemporal activity evoked by the temporoammonic pathway. No significant differences were observed in the activity evoked by the perforant pathway. The VSDI toolbox permitted us to visualize and statistically compare activity across the spatiotemporal scope of the VSDI dataset. Sampling error was minimized because the representation of the data is standardized by the toolbox. Statistical comparisons were conducted quickly, across the spatiotemporal scope of the data, without a priori knowledge of the character of the responses or the likely differences between them.

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A Toolbox for Spatiotemporal Analysis of Voltage-Sensitive Dye Imaging Data in Brain Slices

et al. (2014) A Toolbox for Spatiotemporal Analysis of Voltage-Sensitive Dye Imaging Data in Brain Slices. PLoS ONE 9(9): e108686. doi:10.1371/journal.pone.0108686 A Toolbox for Spatiotemporal Analysis of Voltage- Sensitive Dye Imaging Data in Brain Slices Elliot B. Bourgeois 0 Brian N. Johnson 0 Almedia J. McCoy 0 Lorenzo Trippa 0 Akiva S. Cohen 0 Eric D. Marsh 0 Shashank M. Dravid, Creighton University, United States of America 0 1 Department of Pathology, Brigham and Women's Hospital and Harvard Medical School , Boston , Massachusetts, United States of America, 2 Department of Pediatrics, Division of Pediatric Neurology, The Children's Hospital of Philadelphia, Philadelphia, Pennsylvania, United States of America, 3 Department of Biostatistics and Computational Biology, Dana-Farber Cancer Institute, Boston, Massachusetts, United States of America, 4 Department of Neurology, Division of Pediatric Neurology, The Perelman School of Medicine at the University of Pennsylvania , Philadelphia, Pennsylvania , United States of America Voltage-sensitive dye imaging (VSDI) can simultaneously monitor the spatiotemporal electrical dynamics of thousands of neurons and is often used to identify functional differences in models of neurological disease. While the chief advantage of VSDI is the ability to record spatiotemporal activity, there are no tools available to visualize and statistically compare activity across the full spatiotemporal range of the VSDI dataset. Investigators commonly analyze only a subset of the data, and a majority of the dataset is routinely excluded from analysis. We have developed a software toolbox that simplifies visual inspection of VSDI data, and permits unaided statistical comparison across spatial and temporal dimensions. First, the threedimensional VSDI dataset (x,y,time) is geometrically transformed into a two-dimensional spatiotemporal map of activity. Second, statistical comparison between groups is performed using a non-parametric permutation test. The result is a 2D map of all significant differences in both space and time. Here, we used the toolbox to identify functional differences in activity in VSDI data from acute hippocampal slices obtained from epileptic Arx conditional knock-out and control mice. Maps of spatiotemporal activity were produced and analyzed to identify differences in the activity evoked by stimulation of each of two axonal inputs to the hippocampus: the perforant pathway and the temporoammonic pathway. In mutant hippocampal slices, the toolbox identified a widespread decrease in spatiotemporal activity evoked by the temporoammonic pathway. No significant differences were observed in the activity evoked by the perforant pathway. The VSDI toolbox permitted us to visualize and statistically compare activity across the spatiotemporal scope of the VSDI dataset. Sampling error was minimized because the representation of the data is standardized by the toolbox. Statistical comparisons were conducted quickly, across the spatiotemporal scope of the data, without a priori knowledge of the character of the responses or the likely differences between them. - Funding: Funding for this work was provided by the US National Institutes of Health to EBB (F32 NS080556), EDM (K02 NS065975 and R01 NS082761), ASC (R01 HD059288 and R01 NS069629), and Jeffrey A. Golden (R01 NS046616). The funder had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript. Competing Interests: The authors have declared that no competing interests exist. Voltage-sensitive dye imaging (VSDI) provides unparalleled resolution to assay the emergent properties of complex neural network ensembles. Transmembrane potential can be recorded with relative ease using VSDI, compared to conventional electrophysiological methods, and the spatial resolution of VSDI is limited only by the number of pixels in the VSDI camera. VSDI studies have yielded new insight into the function of native neural networks [1,2,3], and VSDI is increasingly applied to models of neurological disease to identify dysfunctional neural circuits [4,5]. VSDI is typically used to record activity evoked by trains of electrical stimuli, but more recently VSDI has been used to record activation patterns that result from optogenetic stimulation [6,7]. While the chief advantage of VSDI is the ability to broadly record spatiotemporal activity, there are no tools available to conduct quantitative, statistical comparison of two groups of VSDI recordings, across the full spatiotemporal range of the acquired activity. Traditionally, VSDI analysis is conducted on a small subset of the dataset, in hand-selected regions-of-interest (ROIs). First, temporal signals are visually inspected to evaluate possible differences between groups. If differences are apparent, the investigator conducts a quantitative comparison by averaging fluorescence values over a spatial and temporal ROI in each slice, and (...truncated)


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Elliot B. Bourgeois, Brian N. Johnson, Almedia J. McCoy, Lorenzo Trippa, Akiva S. Cohen, Eric D. Marsh. A Toolbox for Spatiotemporal Analysis of Voltage-Sensitive Dye Imaging Data in Brain Slices, PLOS ONE, 2014, Volume 9, Issue 9, DOI: 10.1371/journal.pone.0108686