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. 2020 Mar 31;14:286. doi: 10.3389/fnins.2020.00286

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Copyright © 2020 Iba, Kim, Florio, Mante, Adame, Rockenstein, Kwon, Rissman and Masliah.

This is an open-access article distributed under the terms of the Creative Commons Attribution License (CC BY). The use, distribution or reproduction in other forums is permitted, provided the original author(s) and the copyright owner(s) are credited and that the original publication in this journal is cited, in accordance with accepted academic practice. No use, distribution or reproduction is permitted which does not comply with these terms.

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Double immunohistochemical analysis of p38γ cellular localization in non-Tg and α-syn Tg mouse brains. Vibratome sections of murine brains were double immunofluorescence labeled and analyzed with Apotome II mounted in a Carl Zeiss AxioImager Z1 microscope. Far right (detail): enlarged image of the area contained in the dotted square. (A) p38γ (red) and the pre-synaptic marker synaptophysin (green), colocalization between the two markers is in yellow; (B) image analysis showing high colocalization of p38γ to synapses in non-Tg mice; (C) p38γ (red) and the neuronal marker NeuN (green); (D) image analysis showing increased localization of p38γ to neurons in α-syn Tg mice; (E) p38γ (red) and the astroglial cell marker GFAP (green); (F) image analysis showing minimal localization of p38γ to astroglia in mouse brains; (G) p38γ (red) and α-syn (green); (H) image analysis showing high colocalization of p38γ to α-syn aggregates in the cytoplasm (Lewy body-like) in α-syn Tg mice. (I) α1-syntrophin (red) and pre-synaptic marker synaptophysin (green); (J) image analysis showing low colocalization of α1-syntrophin to pre-synaptic site in mice brains; (K) α1-syntrophin (red) and α-synuclein (green); (L) image analysis showing high localization of α1-syntrophin with α-syn in α-syn Tg mice; Scale bars are 10 μm in the standard panels and 5 μm in the zoomed panels. n = 6 per group (^∗∗p < 0.01, ^∗∗∗p < 0.001). Image analysis of p38γ and synaptic proteins in α-syn Tg mouse brains. The single labeled channels from above were further analyzed to calculate the% area of the neuropil occupied by (M) synaptophysin, (N) α-syn, (O) p38γ, and (P) α1-syntrophin immunofluorescence in non-Tg and Tg mice. (Q) Linear regression analysis was performed to ascertain the correlation coefficient between the levels of p38γ in the neuropil (% area) and synaptophysin, α-syn, and α1-syntrophin in non-Tg and Tg mice. n = 6 per group (**p < 0.01, ***p < 0.001).