RN46A-B14

1,25-dihydroxyvitamin D represses SERT and degradation (MAO-A) gene expression in cultured rat serotonergic neuronal cell lines

To date, low brain serotonin and active vitamin D metabolite (1,25D) have been linked to atypical behavior such as autism and depression. It was previously demonstrated that 1,25D can cause TPH2 expression in rat serotonergic neuronal cells. However, other serotonin pathway components have not been evaluated.

To determine if vitamin D affects the major route of serotonin reuptake and degradation, the influence of 1,25D on the expression of SERT and MAO-A was assessed by quantitative real-time PCR in differentiated serotonergic rat raphe RN46A-B14 cells. Cells in culture were challenged with 1, 10, and 100 nM 1,25D, and mRNA was isolated, reverse transcribed to cDNA and probed for SERT and MAO-A expression. Figure 1a illustrates that SERT is unaffected by 1 and 100 nM 1,25D-treatment, but a concentration of 10 nM 1,25D significantly represses SERT mRNA expression by 59%. Thus, the response of SERT to 1,25D is biphasic, reminiscent of the U-shaped curves generated when plotting circulating 25-hydroxyvitamin D levels versus disease and death in humans. Strikingly, MAO-A displays an identical regulatory pattern of expression when challenged with a range of 1,25D concentrations in differentiated serotonergic rat raphe RN46A-B14 cells (Fig. 1b). MAO-A is similarly unaffected by 1 and 100 nM 1,25D-treatment, but a concentration of 10 nM 1,25D significantly represses MAO-A mRNA expression by 51%. Therefore, two neurally expressed genes exhibit a similar profile of regulation by 1,25D, and the data suggest that certain levels of the vitamin D hormone appear to be capable of suppressing serotonin reuptake and catabolism.

Fig. 1. Effect of 1,25D on selected serotonin degrading pathway and control mRNAs in a serotonergic neuronal cell line (Sabir M S, Haussler M R, et al., 2018).

Pomegranate Derivative Urolithin a Enhances Vitamin D Receptor Signaling to Amplify Serotonin-Related Gene Induction by 1,25-Dihydroxyvitamin D

The active metabolite of vitamin D (1,25D) up-regulates genes through the vitamin D receptor (VDR). Urolithin A, a pomegranate metabolite, induces mitophagy through AMPK. The two together could alter the expression of neuroendocrine genes.

To assess whether urolithin A also increases 1,25D/VDR driven transcription on native chromatin, Livingston et al. quantified CYP24A1 mRNA using qPCR. 1,25D induced CYP24A1 12.6 fold in HEK293 cells (Fig. 2A). The presence of 10 µM urolithin A increased this induction to 3.3 fold. Urolithin A alone had no effect, consistent with its inability to bind the VDR directly. They next asked whether urolithin A produced similar effects in HEK293 cells (Fig. 2A) and RN46A-B14 cells (Fig. 2B). In RN46A-B14 cells, 1,25D induced cyp24a1 8 fold, which was augmented in the presence of urolithin A to a total induction of 100 fold. They also tested resveratrol as a second nutraceutical compound that has been shown to activate AMPK. Resveratrol augmented 1,25D/VDR-mediated transcription, but to a lesser and non-significant degree. They also observed that resveratrol decreased the potency of urolithin A in the presence of both compounds (Fig. 2B, far right treatment group). Urolithin A is the only nutraceutical they tested that can significantly and potently augment VDR-driven gene induction on native chromatin at a half-maximal concentration of 10 μM.

Fig. 2. Effect of 1,25D and UroA and/or Res on CYP24A1 mRNA concentrations in cultured cells (Livingston S, Mallick S, et al., 2020).