Fig. 3

Acute melatonin treatment upregulated Nrf2/HO-1/GCLM expression in the acute ethanol-treated rat pups and in HT22 and BV2 cells that were exposed to ethanol. a Western blot analysis of nuclear/cytosolic Nrf2 expression using Nrf2, HO-1, and GCLM antibodies in the rat pups. The bands were quantified using Sigma Gel software, and the differences are presented in a histogram. β-Actin was used as a loading control. n = 10 pups/group, and the number of experiments = 3. b Representative immunofluorescence results of Nrf2 (FITC, DAPI, Blue) and c, d immunohistochemical results of Nrf2 and HO-1 respectively in the cortices and CA1 regions of the hippocampi in the rat pups. n = 5 pups/group, and the number of experiments = 3. Magnification × 20. Scale bar = 50 μm. e, f Western blots and the densitometric analysis of the Nrf2, HO-1, and GCLM expression in the HT22 and BV2 cells respectively that were subjected to Nrf2 siRNA and treated with ethanol (100 mM) and melatonin (100 μM) for 12 h. β-actin was used as a loading control. The data are expressed as the mean ± SEM, and the number of experiments = 3. The data are presented relative to control values. Significance = P < 0.05. σ Significantly different from the control saline-treated rat pups; Φ significantly different from the ethanol-treated rat pups. Similarly for in vitro studies, σ significantly different from the non-treated HT22 and BV2 cells; Φ significantly different from the ethanol-exposed HT22 and BV2 cells, and ω significantly different from the ethanol + melatonin-exposed HT22 and BV2 cells