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[Preprint]. 2023 Oct 10:2023.10.06.561261. [Version 1] doi: 10.1101/2023.10.06.561261

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This article is a US Government work. It is not subject to copyright under 17 USC 105 and is also made available for use under a CC0 license.

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Figure 10: — A. FLEX-RAM labeled neurons were filled with neurobiotin during slice electrophysiology recordings (see Fig 8 for details). Following electrophysiology recordings, slices were post-fixed in 4% PFA for 24-hours and then 1x PBS until immunostaining. Slices were stained for ChAT (to visualize cholinergic neurons) and streptavidin (to visualize neurobiotin filled cells). Following immunostaining, slices were imaged on a confocal microscope to visualize FLEX-RAM, ChAT, and streptavidin.

B. Confocal images were reconstructed using Imaris software. Reconstructed neurons from an appetitive odor activated (APP) VP cholinergic neuron (n = 10, left) and an aversive odor activated (AV) VP cholinergic neuron (n = 6, right), with a convex hull analysis. A convex hull is a 3D polyhedron that encompasses all distal points of the reconstructed neuron. Scale bar is 10 μm.

C – D. Surface area and volume measurements of the convex hull. The convex hull from APP VP cholinergic neurons are significantly smaller in (C) surface area and (D) volume. * p < 0.05.

E – G. APP VP cholinergic neurons are smaller and have fewer linear dimension features vs AV VP cholinergic neurons. APP VP cholinergic neurons exhibit a significantly smaller (E) dendrite area, (F) dendrite volume, and (G) max length of reconstructed segment. * p < 0.05.

H. Sholl analysis of reconstructed APP and AV VP cholinergic neurons. Despite exhibiting reduced morphometric properties, APP VP cholinergic neurons display greater proximal soma complexity in a sholl analysis. APP VP cholinergic neurons display a significantly greater number of branch points at 5 and 10 μm from the soma. * p < 0.05.

I. Although AV VP cholinergic neurons display less branching vs. APP VP cholinergic neurons, AV neurons are overall lengthier. A larger percentage of AV VP cholinergic neurons are able to reach greater distances away from the soma vs. APP VP cholinergic neurons.