FIGURE 3.

Microglial metabolic reprogramming is involved in the anti‐inflammatory effect of MT1 receptor activation. (a) BV2 cells were transfected with control siRNA or Mtnr1a siRNA for 48 hr, then exposed to LPS (100 ng/ml) for 12 hr. After treatments, the culture media were collected and were used to detect the glucose consumption by the QuantiChrom™ Glucose Assay kits. The values were presented as the means ± SEM from three independent experiments. **p < 0.01, two‐way ANOVA followed by Sidak’s post‐hoc test. (b) BV2 cells were treated as described in (a) and then the culture media were collected to determine the lactate production using Lactate Assay kits. The values were presented as the means ± SEM from three independent experiments. *p < 0.05, **p < 0.01, two‐way ANOVA followed by Sidak's post‐hoc test. (c) BV2 cells were pretreated with Ramelteon (10 μM, 50 μM, 100 μM) for 12 hr and then exposed to LPS (100 ng/ml) for 12 hr. After treatments, the culture media were collected and were used to detect the glucose consumption by the QuantiChrom™ Glucose Assay kits. The values were presented as the means ± SEM from three independent experiments. *p < 0.05, **p < 0.01, two‐way ANOVA followed by Sidak’s post‐hoc test. (d) BV2 cells were treated as described in (c) and then the culture media were collected to determine the lactate production using Lactate Assay kits. The values were presented as the means ± SEM from three independent experiments. *p < 0.05, **p < 0.01, two‐way ANOVA followed by Sidak’s post‐hoc test. (e, f) Primary microglia were treated as (a) and (b), after treatments, the culture media were collected and were used to detect the glucose consumption and lactate production by the QuantiChrom™ Glucose Assay kits and Lactate Assay kits. (g‐h) Primary microglia were treated as (c) and (d), after treatments, the culture media were collected and were used to detect the glucose consumption and lactate production by the QuantiChrom™ Glucose Assay kits and Lactate Assay kits. (i) BV2 cells were pretreated with 2‐DG for 2 hr, followed by the knocking down of Mtnr1a and a sequential LPS stimulation. Next, the protein levels of iNOS, COX‐2 and GAPDH were measured using immunoblot analyses. Quantitative analyses of panel (i) were shown in panel below. The values were presented as the means ± SEM from three independent experiments. *p < 0.05, **p < 0.01, ns, no significance, two‐way ANOVA followed by Sidak’s post‐hoc test