Methods for voltage-sensitive dye imaging of rat cortical activity with high signal-to-noise ratio

We describe methods to achieve high sensitivity in voltage-sensitive dye (VSD) imaging from rat barrel and visual cortices in vivo with the use of a blue dye RH1691 and a high dynamic range imaging device (photodiode array). With an improved staining protocol and an off-line procedure to remove pulsation artifact, the sensitivity of VSD recording is comparable with that of local field potential recording from the same location. With this sensitivity, one can record from approximately 500 individual detectors, each covering an area of cortical tissue 160 microm in diameter (total imaging field approximately 4 mm in diameter) and a temporal resolution of 1,600 frames/s, without multiple-trial averaging. We can record 80-100 trials of intermittent 10-s trials from each imaging field before the VSD signal reduces to one half of its initial amplitude because of bleaching and wash-out. Taken together, the methods described in this report provide a useful tool for visualizing evoked and spontaneous waves from rodent cortex.