Rapid Action of Retinoic Acid on the Hypothalamic Pituitary Adrenal Axis

Abstract

Retinoic acid (RA) is the active metabolite of vitamin A but is also used as a medication, primarily for acne in which the treatment regime lasts several months. A number of studies have indicated that treatment with RA over this time period impacts the hypothalamic-pituitary-adrenal (HPA) axis and may contribute to a number of the side-effects of the drug. No studies though have investigated the short-term, early effects RA may have on the HPA axis via the transcriptional pathways activated by the RA receptor. This study investigated the action of RA over 3 days on regulatory components of the HPA axis. Several key genes involved in glucocorticoid feedback pathways in the hippocampus, hypothalamus and pituitary were unchanged after 3-days exposure to RA. Key elements though in the adrenal gland involved in corticosterone and aldosterone synthesis were altered in particular with the Cyp11b2 gene downregulated in vivo and ex vivo. The rapid, 5 h, change in Cyp11b2 expression suggested this activation may be direct. These results highlight the adrenal gland as a target of short-term action of RA and potentially a trigger component in the mechanisms by which the long-term adverse effects of RA treatment occur.

Keywords: Cyp11b1; Cyp11b2; adrenal gland; aldosterone synthesis; corticosterone; hypothalamic pituitary adrenal axis; retinoic acid.

Figure 1

The influence of 3-day 1 mg/kg or 10 mg/kg retinoic acid (RA) treatment of the rat on mRNA expression levels in the (A) hippocampus, (B) hypothalamus and (C) pituitary gland. The Rarb positive control was the only gene to show a statistically significant change in the three tissues (p < 0.001 in all cases) indicated by *. All control values were normalized to 100%. Panel A; n = 6 RA, n = 6 control; panel B; n = 6 RA, n = 6 control and panel C; n = 5 RA, n = 5 control. * = p < 0.05 , ** = p < 0.01, **** = p < 0.0001.

Figure 2

The influence of in vivo and ex vivo RA treatment on adrenal gland Cyp11b1, Cyp11b2 and Rarb expression. (A) The relative effects of 1 mg/kg vs. 10 mg/kg dose of RA in the rat was compared on gene expression and there was no influence on Cyp11b1 at either dose (1 mg/kg RA, p = 0.882, 10 mg/kg RA, p = 0.874) while Cyp11b2 was repressed at the higher dose (1 mg/kg RA, p = 0.188, 10 mg/kg RA, p = 0.018). Rarb was strongly induced by the 10 mg/kg RA dose (p = 0.00009) but surprisingly repressed at 1 mg/kg RA (p = 0.048). (B) Five-hour treatment of the rat with RA resulted in significant induction of the Rarb positive control in the adrenal gland and Cy11b2 fell in expression (p < 0.001 and p = 0.018, respectively). (C) Five-hour treatment with RA of an ex vivo adrenal glands culture resulted in Rarb again increasing and Cyp11b2 falling in expression (p = 0.0188 and p = 0.0184, respectively). * = p < 0.05 , **** = p < 0.0001.

Figure 3

The influence of in vivo RA treatment on adrenal gland expression of CYP11B1 and CYP11B2 protein as well as serum aldosterone and corticosterone levels. (A) Western blotting and band quantitation to determine the change in CYP11B1 and CYP11B2 relative to β-actin (imaged simultaneously) following in vivo RA treatment showed a significant increase in CYP11B1 (p = 0.043) and no significant changes in CYP11B2 (p = 0.798). All control values were normalized to 100%. (B) Quantitation of blood aldosterone levels with in vivo RA treatment indicated no significant change (p = 0.221) while (C) quantitation of blood corticosterone levels indicated a significant decline in hormone after in vivo RA treatment (p = 0.049). Significance was indicated by *. N = 5 per treatment group in each case. *p < 0.05.