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Once a standard protocol and equipment were developed, activity of individual neurons to a visual stimulus in a targeted area of the visual cortex could be conducted. Neuronal activity was measured in GCaMP6-expressing neurons from selected cortical layers from four separate transgenic mouse lines. For more information on these how neurons in specific layers and areas were targeted in these transgenic lines, see the Transgenic Line Catalog in Documentation.

Video recordings of the visual cortex were taken during presentation of the various visual stimuli. The data processing of these raw data movies involved motion correction and image segmentation to identify the sets of pixels representing distinct cells. These segmentation masks were used to extract traces from each neuron so that cellular activity over time can be analyzed. The goal was to relate the activity of each cell in the field of view back to the visual stimulus that was viewed by the mouse.

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