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. 2022 Sep 2;15:976349. doi: 10.3389/fnmol.2022.976349

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Copyright © 2022 Tsuneoka, Atsumi, Makanae, Yashiro and Funato.

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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Cell typing using two rounds of ISH combined with Nissl staining. Two-round ISH visualized Esr1, Hrh1, Cart, neurotensin, Moxd1, and Tac2 mRNAs. (A) Wide-field merged image of the medial preoptic area (MPOA). The rectangle indicates the region shown in (B–D). Scale bar: 100 μm. (B) Merged image of six different mRNAs and fluorescent Nissl staining (gray). (C) Drawing of each cell position and cell type. Colors indicate different cell types shown at the bottom. (D) Separate images of each mRNA and fluorescent Nissl staining. Scale bar: 25 μm. BNSTpr, bed nucleus of the stria terminalis, principal nucleus; cMPOA, central part of MPOA; dmMPOA, dorsomedial part of MPOA; MPNc, medial preoptic nucleus (MPN), core part; MPNl, MPN, lateral part; vlMPOA, ventrolateral MPOA.