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. 2023 Mar 31;14:1800. doi: 10.1038/s41467-023-36808-2

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© The Author(s) 2023

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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Fig. 5 — A Experimental timeline for DiD behavioral studies. B Average alcohol intake in DiD, two-tailed Student’s unpaired t-tests, p_2h = 0.0042, p_4h = 0.0069C Cumulative alcohol intake in DiD, two-tailed Student’s unpaired t-test, p = 0.0017. D Average alcohol preference in DiD, two-tailed Student’s unpaired t-tests, p_2h = 0.0841, p_4h = 0.0685. E Blood alcohol concentration immediately following 4h of DiD, two-tailed Student’s unpaired t-test. F Social preference in the 3-chamber sociability test. G Social novelty preference in the 3-chamber sociability test, p_sexXDiD = 0.008. H Percent time in center of open field, p_DiD = 0.0019, p_sex < 0.0001. I Center entries open field, p_DiD < 0.0001. J Total distance in open field, p_DiD = 0.0013, p_sex = 0.0366. K Mean velocity in open field, p_DiD = 0.0013, p_sex = 0.0366. L Acoustic startle behavior, females (10 trials/dB), two-way repeated measures ANOVA. M Acoustic startle behavior, males (10 trials/dB), two-way repeated measures ANOVA followed by Holm-Sidak’s post-hoc, p_DiD = 0.0821, p_DiDXdB = 0.0055. N Acoustic startle behavior, 120 dB only (10 trials), p_DiD = 0.0164, p_sex = 0.0034. O Experimental timeline for DiD qPCR studies. P Average alcohol intake in DiD, two-tailed Student’s unpaired t tests, p_2h = 0.1589, p_4h = 0.1012. Q Cumulative alcohol intake in DiD, two-tailed Student’s unpaired t-test, p = 0.0448. R Average alcohol preference in DiD, two-tailed Student’s unpaired t tests, p_2h = 0.0054, p_4h = 0.1964. S Relative expression of Htr2c mRNA in Hb, p_DiD = 0.0005, p_sex = 0.0902, p_DiDXsex = 0.1012. T Relative expression of Htr2c mRNA in BNST. For panels 3B-3N, Females: water n = 8 mice, DiD n = 8 mice; Males: water n = 8 mice, DiD n = 8 mice. For panels 3P–3R, Females: n = 10 mice; Males: n = 10 mice. For panels 3S and 3T, Females: water n = 10 mice/3 replicates, DiD n = 10 mice/3 replicates; Males: water n = 10 mice/3 replicates, DiD n = 10 mice/3 replicates. Unless otherwise stated, statistical comparisons were performed using two-way ANOVA followed by Holm-Sidak’s post-hoc. All data are presented as mean ± SEM. & denotes effect of DiD (&p < 0.05, &&p < 0.01, &&&p < 0.001, &&&&p < 0.0001), @ denotes effect of sex (@p < 0.05, @@p < 0.01, @@@p < 0.001, @@@@p < 0.0001), # denotes interaction (sex × DiD or DiD × stimulus strength, #p < 0.01, ##p < 0.01), and * denotes post-hoc p < 0.05, ** denotes post-hoc p < 0.01. Source data are provided as a Source Data file. Created with Biorender.com.