Fig. 4. In the fetal hypothalamus, diurnal rhythms of GR and REVERBα are anti-phasic.

a Working model of reverse erythroblastoma alpha (REVERBα)-mediated regulation of rhythmic glucocorticoid receptor (GR) protein amount and prenatal programming of the offspring’s stress response. b, c Circadian profile of GR (b) and REVERBα (c) protein in fetal hypothalami obtained from naive mothers euthanized every 6 h (at Zeitgeber time (ZT) 1, 7, 13 and 19) during gestational day (GD) 15.5 to 16.5, (GR n = 6 ZT1, n = 5 ZT7, n = 5 ZT13, n = 5 ZT19 and REVERBα n = 6 for all time points). Representative Western blot results are shown; samples were run in duplicates and α-Tubulin was used as loading control. d Gr mRNA expression in fetal hypothalami from naive pregnant mothers euthanized at ZT1 (n = 7) and ZT13 (n = 8). e GR binding to glucocorticoid receptor binding elements (GREs) assessed in fetal hypothalami from naive pregnant mothers (n = 4 for both time points). f, g RevErbα mRNA expression in wild-type (n = 10 ZT1 and n = 12 ZT13) (f) and brain and muscle aryl hydrocarbon receptor nuclear translocator-like 1 (Bmal1^−/−) (g) fetal hypothalamus (n = 5 ZT1 and n = 4 ZT13) from naive mothers. qPCRs for gene expression and GR binding assays were run in duplicates. All data are expressed as means ± SEM. In b, c significant circadian rhythmicity was determined by CircWave v1.4 which fits a standard cosine function to an averaged de-trended data assuming a period of 24 h, a p < 0.05 was considered significant (^, b r² = 0.37, p = 0.05; c r² = 0.38, p = 0.028). In f data were analyzed by two-sided unpaired t-test (f t = 4.35, df=20, *p = 0.0003). Source data are provided as a Source Data file.