Fig. 3. Testosterone levels in serum in males after fear acquisition are decreased by osanetant while estradiol in females’ serum is increased 330 min after its administration compared to vehicle-treated mice, with no effect of the drug on progesterone, corticosterone, deoxycorticosterone, or dehydrocorticosterone.

All mice received osanetant (5 mg/Kg, ip) or vehicle 30 min after fear acquisition, and trunk blood was collected either 30 min or 330 min after the injection. Estradiol measures were only determined 330 min after the injection. Dashed lines indicate the relatively basal concentration for an independent group of animals that did not receive any treatment. a–c Testosterone concentration in males (p = 0.001, treatment₃₃₀min: p = 0.007), proestrous females, and metestrous females. d–f Serum corticosterone concentration in males (p = 0.000), proestrous females (p = 0.000), and metestrous females (p < 0.000). g–i Circulating progesterone concentration in serum in males (p = 0.000), proestrous females, and metestrous females. j–l Males (p < 0.000), proestrous (p < 0.000), and metestrous (p < 0.000) females’ serum dehydrocorticosterone (DHC) concentration. m–o Concentration of circulating deoxycorticosterone (DOC) in serum in males (p = 0.000), proestrous (p < 0.000) and metestrous females (p < 0.000). p Serum estradiol concentration after FA and 330 min after receiving osanetant in proestrous females (p = 0.022). q Serum estradiol concentration after FA and 330 min after receiving osanetant in metestrous females. Basal levels of the estradiol experiment were replicated in Supplementary Fig. 3e with similar results. Males vehicle₃₀: n = 11; males osanetant₃₀: n = 10; males vehicle₃₃₀: n = 16; males osanetant₃₃₀: n = 16; proestrous females vehicle₃₀: n = 8; proestrous females osanetant30: n = 8; proestrous females vehicle₃₃₀: n = 7; proestrous females osanetant₃₃₀: n = 8; metestrous females vehicle₃₀: n = 8; metestrous females osanetant₃₀: n = 8; metestrous females vehicle₃₃₀: n = 8; metestrous females osanetant₃₃₀: n = 8. Proestrous females vehicle₃₃₀ for estradiol determination n = 5; proestrous females osanetant₃₃₀ for estradiol determination n = 7; metestrous females vehicle₃₃₀ for estradiol determination n = 7, metestrous females osanetant₃₃₀ for estradiol determination n = 6. Data are mean ± SEM. *p ≤ 0.05 **p ≤ 0.01 ***p ≤ 0.001 vs its vehicle. When with lines, it indicates specific comparisons. ⁺p ≤ 0.05 ⁺⁺p ≤ 0.01 ⁺⁺⁺p ≤ 0.001 vs baseline. The statistic was Wald’s χ² with pairwise comparisons between groups for testosterone, corticosterone, progesterone, DHC, and DOC. Estradiol was analyzed using non-parametric Kruskal–Wallis’ χ².