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. 2018 Feb 2;8:2242. doi: 10.1038/s41598-018-20563-2

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© The Author(s) 2018

Open Access This article is licensed under a Creative Commons Attribution 4.0 International License, which permits use, sharing, adaptation, distribution and reproduction in any medium or format, as long as you give appropriate credit to the original author(s) and the source, provide a link to the Creative Commons license, and indicate if changes were made. The images or other third party material in this article are included in the article’s Creative Commons license, unless indicated otherwise in a credit line to the material. If material is not included in the article’s Creative Commons license and your intended use is not permitted by statutory regulation or exceeds the permitted use, you will need to obtain permission directly from the copyright holder. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/.

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Whole-mount immunolabelling and optical tissue clearing in the rat brain. (A) The intact rat brain before (i) after (ii) immunolabelling for tyrosine hydroxylase (TH) and tissue clearing. (B) 3D rendering of TH-immunoreactivity (ir) in the rat brain imaged in the horizontal plane and viewed from the ventral surface (z depth = 4 mm). (C) Hypothalamic TH-ir neurons viewed in the sagittal plane from −2.6 mm to −4 mm Bregma. Insets (D,E): (D) TH-ir neurons in periventricular nuclei viewed from the dorsal surface with high magnification renderings of neurons in the horizontal plane (D i) and rotated to the sagittal (D ii) plane. (E) TH-ir neurons within a portion of the dorsomedial and arcuate hypothalamic nuclei projected into the coronal plane. (F) 3D rendering and a high resolution projected image (F i) of TH-ir fibers in the cortex. (G) TH-positive neurons within the substantia nigra and ventral tegmental area of the midbrain. (H) TH-positive neurons of the A5 group located in the ventrolateral pons viewed in the horizontal plane (i) and rotated into the sagittal plane (ii). R = rostral, C = caudal, D = dorsal, V = ventral, oc = optic chiasm, ME = median eminence, 3 V = third ventricle.