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. 2017 Jul 21;11:50. doi: 10.3389/fncir.2017.00050

Figure 1.

Figure 1

Neuropil and somatic response variability and dependence to contrast. (A) Selecting cell somata and local neuropil patches. An example of a typical field of view (FOV, left). The orange area is magnified on the right. Cell radii in our experiments were generally ≤5 μm. A local neuropil patch was defined around each cell body as an annulus with inner radius 7 μm and outer radius 15 μm. Any annulus pixels belonging to or being close to other cells (i.e., within 7 μm of the cell center), vessels, or poorly stained regions were excluded by visual inspection to minimize contamination of the neuropil signal by sources other than neuropil. In the example, the green outline shows a selected cell soma (green outline only for visualization). Its local neuropil patch is outlined in red. Note that nearby neurons were excluded from the definition of the neuropil ROI (annulus) and are shown in blue (outline only). The pixel resolution is 1.2 × 1.2 μm. (B) Gaussian fit of the point spread function (PSF) of our microscope measured by imaging a single spherical fluorophore with a radius of 0.1 μm. (C) Example of the average response of cells and neuropil-patches (annular patch with radii of 7–15 μm) in a single FOV to 100% contrast. “NP”: mean neuropil response. “Uncor”: mean uncorrected somatic response. “Cor”: mean somatic response after neuropil contamination correction (see Section Materials and Methods) (D) Mean evoked fluorescence responses to gratings of 100, 40, and 15% contrast, respectively, derived from all visually responsive cells in 7 anesthetized (AN) and 11 awake (AW) FOV's, respectively. (E) Ratio of mean somatic vs. mean neuropil responses (pooled from all FOV data). These ratios are plotted after correcting for neuropil contamination. Note that the relative strength of cell vs. neuropil ΔF/F responses decreased in the lightly anesthetized state. This is largely due to the fact that neuropil responses markedly increase in the lightly anesthetized state. NP, neuropil; AW, awake state; AN, anesthetized state. Statistics are across FOV's.