Figure 5.

ROIs can be generated using noisy, low spatial resolution datasets. A. anatomical location of recording overlaps with the preBötC, which has been identified as lying 500 µm caudal to the VIIn (Ruangkittisakul et al., 2005). Two juxtaposed ROIs are likely generated by the same cell (arrow). In the 4 datasets analyzed, the number of respiration modulated ROI traces remained at approximately 10% of the total number of ROIs identified (inset). B. Traces sorted to match ROI numbering reveal closely matched activity in 2 adjacent ROIs (arrows), suggesting that one cell has been misidentified as 2. Despite the fact that traces 8 and 9 are likely generated from the same cell, the two traces do not match exactly (boxes). C. Effects of high- and band-pass filtering on the optical recording from one ROI (dotted box, 2^nd optical trace from the bottom, Figure 5 B). Top trace: the raw optical signal, obtained by calculating the mean of pixels within the ROI for each frame in the data series. High-pass filtering (via subtraction of the trace low-pass filtered using a 14 s moving average) eliminates slow time-course fluorescence fluctuations (middle trace); subsequent low-pass filtering (by taking a 0.6 s moving average of the signal) gives rise to a band-pass fitered signal in which high-frequency fluctuations in brightness associated with arc-lamp jitter is attenuated (bottom trace), but in which the underlying Ca²⁺ signal is retained. The effect of low-pass filtering on one Ca²⁺ peak associated with an inspiratory burst (dotted box, left) is shown on an expanded time-scale (right panel).