Fig. 3.

Intravital calcium imaging of mice with labeled PNNs. (a) A representative GCaMP6s-labeled field of view (FOV) with markings of some of the detected active components. The $290\times 290\mu \mathrm{m}$ FOV was captured at 30 Hz, with the depicted image being 1000 frames average for display purposes only. Inset – 30 frames averaged image of ${\mathrm{PNN}}^{+}$ cells (WFA-Atto590; magenta) overlaying the GCaMP image (grayscale). (b) Representative $\mathrm{\Delta }F/F$ traces of the marked cells with a PNN (magenta) and without a PNN (black). (c) Representative $\mathrm{\Delta }F/F$ traces of ${\mathrm{PNN}}^{-}$ (left; black) and ${\mathrm{PNN}}^{+}$ (right; magenta) neurons at different cortical depths (measured from pia) in a mouse injected with WFA-FITC and RCaMP7. Vertical lines in the traces indicate the timing of whisker stimulation and are scaled to 300% $\mathrm{\Delta }F/F$ change. (d) A representative FOV of ${\mathrm{PV}}^{+}$ cells expressing labeled with WFA-Alexa594 (magenta) and GCaMP7f (gray) with marking of some of the detected active components (as detected by CaImAn), with their corresponding $\mathrm{\Delta }F/F$ traces of the marked cells with a PNN (magenta) and without a PNN (black) (see Video 5, MOV, 1.52 MB [URL: https://doi.org/10.1117/1.NPh.10.1.015008.s5]). (e) The calcium transient event rate in the barrel cortex was similar between ${\mathrm{PNN}}^{+}$ and ${\mathrm{PNN}}^{-}$ PV cells, while the mean AUCs of the $\mathrm{\Delta }F/F$ traces were significantly higher in ${\mathrm{PNN}}^{+}$ PV cells compared to ${\mathrm{PNN}}^{-}$ cells in the same mouse ($n=3$ mouse, 147 ${\mathrm{PNN}}^{+}$ cells and 1025 ${\mathrm{PNN}}^{-}$ cells, unpaired two-tailed student $t$-test, $t=2.6066$, $p=0.0092$). Moreover, the variance of the mean AUCs of ${\mathrm{PNN}}^{-}$ cells was significantly larger compared to ${\mathrm{PNN}}^{+}$ PV cells (Levene’s test, $p=0.0092$). Data were averaged to 1 fps for display purposes only. Scale bars are $50\mu \mathrm{m}$.