A seasonal switch in histone deacetylase gene expression in the hypothalamus and their capacity to modulate nuclear signaling pathways.

Seasonal animals undergo changes in physiology and behavior between summer and winter conditions. These changes are in part driven by a switch in a series of hypothalamic genes under transcriptional control by hormones and, of recent interest, inflammatory factors. Crucial to the control of transcription are histone deacetylases (HDACs), generally acting to repress transcription by local histone modification. Seasonal changes in hypothalamic HDAC transcripts were investigated in photoperiod-sensitive F344 rats by altering the day-length (photoperiod). HDAC4, 6 and 9 were found to change in expression. The potential influence of HDACs on two hypothalamic signaling pathways that regulate transcription, inflammatory and nuclear receptor signaling, was investigated. For inflammatory signaling the focus was on NF-κB because of the novel finding made that its expression is seasonally regulated in the rat hypothalamus. For nuclear receptor signaling it was discovered that expression of retinoic acid receptor beta was regulated seasonally. HDAC modulation of NF-κB-induced pathways was examined in a hypothalamic neuronal cell line and primary hypothalamic tanycytes. HDAC4/5/6 inhibition altered the control of gene expression (Fos, Prkca, Prkcd and Ptp1b) by inducers of NF-κB that activate inflammation. These inhibitors also modified the action of nuclear receptor ligands thyroid hormone and retinoic acid. Thus seasonal changes in HDAC4 and 6 have the potential to epigenetically modify multiple gene regulatory pathways in the hypothalamus that could act to limit inflammatory pathways in the hypothalamus during long-day summer-like conditions.

A neuroendocrine role for chemerin in hypothalamic remodelling and photoperiodic control of energy balance.

Long-term and reversible changes in body weight are typical of seasonal animals. Thyroid hormone (TH) and retinoic acid (RA) within the tanycytes and ependymal cells of the hypothalamus have been implicated in the photoperiodic response. We investigated signalling downstream of RA and how this links to the control of body weight and food intake in photoperiodic F344 rats. Chemerin, an inflammatory chemokine, with a known role in energy metabolism, was identified as a target of RA. Gene expression of chemerin (Rarres2) and its receptors were localised within the tanycytes and ependymal cells, with higher expression under long (LD) versus short (SD) photoperiod, pointing to a physiological role. The SD to LD transition (increased food intake) was mimicked by 2 weeks of ICV infusion of chemerin into rats. Chemerin also increased expression of the cytoskeletal protein vimentin, implicating hypothalamic remodelling in this response. By contrast, acute ICV bolus injection of chemerin on a 12 h:12 h photoperiod inhibited food intake and decreased body weight with associated changes in hypothalamic neuropeptides involved in growth and feeding after 24 hr. We describe the hypothalamic ventricular zone as a key site of neuroendocrine regulation, where the inflammatory signal, chemerin, links TH and RA signaling to hypothalamic remodeling.

Thyroid hormone activation of retinoic acid synthesis in hypothalamic tanycytes.

Thyroid hormone (TH) is essential for adult brain function and its actions include several key roles in the hypothalamus. Although TH controls gene expression via specific TH receptors of the nuclear receptor class, surprisingly few genes have been demonstrated to be directly regulated by TH in the hypothalamus, or the adult brain as a whole. This study explored the rapid induction by TH of retinaldehyde dehydrogenase 1 (Raldh1), encoding a retinoic acid (RA)-synthesizing enzyme, as a gene specifically expressed in hypothalamic tanycytes, cells that mediate a number of actions of TH in the hypothalamus. The resulting increase in RA may then regulate gene expression via the RA receptors, also of the nuclear receptor class. In vivo exposure of the rat to TH led to a significant and rapid increase in hypothalamic Raldh1 within 4 hours. That this may lead to an in vivo increase in RA is suggested by the later induction by TH of the RA-responsive gene Cyp26b1. To explore the actions of RA in the hypothalamus as a potential mediator of TH control of gene regulation, an ex vivo hypothalamic rat slice culture method was developed in which the Raldh1-expressing tanycytes were maintained. These slice cultures confirmed that TH did not act on genes regulating energy balance but could induce Raldh1. RA has the potential to upregulate expression of genes involved in growth and appetite, Ghrh and Agrp. This regulation is acutely sensitive to epigenetic changes, as has been shown for TH action in vivo. These results indicate that sequential triggering of two nuclear receptor signalling systems has the capability to mediate some of the functions of TH in the hypothalamus.

The evidence for a beneficial role of vitamin A in multiple sclerosis.

Vitamin A is an essential nutrient with important roles in immunological responses and in brain development. Its main metabolite is retinoic acid (RA), which is responsible for the neuroimmunological functions related to vitamin A. In the brain, RA is known to have interactions with other nuclear receptor-mediated signalling pathways. RA is involved in plasticity, regeneration, cognition and behaviour. In the peripheral blood, RA plays a major role both in increasing tolerance and in decreasing inflammation, through balancing T-lymphocyte populations. It is likely that RA synthesis may be manipulated by complex cross-talk among cells during infection and inflammation. The role of vitamin A in multiple sclerosis (MS) could be dual: at the same time as it decreases inflammation and increases tolerance of autoimmunity, it may also help in brain protection. The present review discusses the beneficial effects that vitamin A might have for controlling MS, although it must be clearly stated that, at the present time, there is no clear indication for using vitamin A as a treatment for MS. However, the results from the present review should encourage clinical trials with vitamin supplementation as a potential treatment or as an add-on option. Vitamin A acts in synergy with vitamin D, and the immunological homeostasis ensured by these vitamins should not be unbalanced in favour of only one of them.

Fenretinide treatment prevents diet-induced obesity in association with major alterations in retinoid homeostatic gene expression in adipose, liver, and hypothalamus.

The synthetic retinoid, Fenretinide (FEN), inhibits obesity and insulin resistance in mice and is in early clinical trials for treatment of insulin resistance in obese humans. We aimed to determine whether alterations in retinoic acid (RA)-responsive genes contribute to the beneficial effects of FEN. We examined the effect of FEN on 3T3-L1 adipocyte differentiation and alterations in gene expression in C57Bl/6 and retinaldehyde dehydrogenase (RALDH) 1 knockout (KO) mice fed a high-fat (HF) diet. FEN completely inhibited adipocyte differentiation by blocking CCAAT/enhancer-binding protein (C/EBP) α/peroxisome proliferator-activated receptor (PPAR) γ-mediated induction of downstream genes and upregulating RA-responsive genes like cellular retinol-binding protein-1. In mice fed an HF diet, RA-responsive genes were markedly increased in adipose, liver, and hypothalamus, with short-term and long-term FEN treatment. In adipose, FEN inhibited the downregulation of PPARγ and improved insulin sensitivity and the levels of adiponectin, resistin, and serum RBP (RBP4). FEN inhibited hyperleptinemia in vivo and leptin expression in adipocytes. Surprisingly, hypothalamic neuropeptide Y expression was completely suppressed, suggesting a central effect of FEN to normalize hyperglycemia. Moreover, FEN induced RA-responsive genes in RALDH1 KO mice, demonstrating that FEN can augment RA signaling when RA synthesis is impaired. We show that FEN-mediated beneficial effects are through alterations in retinoid homeostasis genes, and these are strong candidates as therapeutic targets for the treatment of obesity and insulin resistance.

A paradoxical teratogenic mechanism for retinoic acid.

Retinoic acid, an active metabolite of vitamin A, plays essential signaling roles in mammalian embryogenesis. Nevertheless, it has long been recognized that overexposure to vitamin A or retinoic acid causes widespread teratogenesis in rodents as well as humans. Although it has a short half-life, exposure to high levels of retinoic acid can disrupt development of yet-to-be formed organs, including the metanephros, the embryonic organ which normally differentiates into the mature kidney. Paradoxically, it is known that either an excess or a deficiency of retinoic acid results in similar malformations in some organs, including the mammalian kidney. Accordingly, we hypothesized that excess retinoic acid is teratogenic by inducing a longer lasting, local retinoic acid deficiency. This idea was tested in an established in vivo mouse model in which exposure to excess retinoic acid well before metanephric rudiments exist leads to failure of kidney formation several days later. Results showed that teratogen exposure was followed by decreased levels of Raldh transcripts encoding retinoic acid-synthesizing enzymes and increased levels of Cyp26a1 and Cyp26b1 mRNAs encoding enzymes that catabolize retinoic acid. Concomitantly, there was significant reduction in retinoic acid levels in whole embryos and kidney rudiments. Restoration of retinoic acid levels by maternal supplementation with low doses of retinoic acid following the teratogenic insult rescued metanephric kidney development and abrogated several extrarenal developmental defects. This previously undescribed and unsuspected mechanism provides insight into the molecular pathway of retinoic acid-induced teratogenesis.

Photoperiodic expression of two RALDH enzymes and the regulation of cell proliferation by retinoic acid in the rat hypothalamus.

Retinoic acid (RA) has been found to regulate hypothalamic function, but precisely where it acts is unknown. This study shows expression of retinaldehyde dehydrogenase (RALDH) enzymes in tanycytes that line the third ventricle in an area overlapping with the site of hypothalamic neural stem cells. The influence of RA was examined on the proliferation of progenitors lining the third ventricle using organotypic slice cultures. As has been shown in other regions of neurogenesis, RA was found to inhibit proliferation. Investigations of the dynamics of RALDH1 expression in the rat hypothalamus have shown that this enzyme is in tanycytes under photoperiodic control with highest levels during long versus short days. In parallel to this shift in RA synthesis, cell proliferation in the third ventricle was found to be lowest during long days when RA was highest, implying that RALDH1 synthesized RA may regulate neural stem cell proliferation. A second RA synthesizing enzyme, RALDH2 was also present in tanycytes lining the third ventricle. In contrast to RALDH1, RALDH2 showed little change with photoperiodicity, but surprisingly the protein was present in the apparent absence of mRNA transcript and it is hypothesized that the endocytic tanycytes may take this enzyme up from the cerebrospinal fluid (CSF).

Photoperiod regulates vitamin A and Wnt/ß-catenin signaling in F344 rats.

In seasonal mammals, growth, energy balance, and reproductive status are regulated by the neuroendocrine effects of photoperiod. Thyroid hormone (TH) is a key player in this response in a number of species. A neuroendocrine role for the nutritional factor vitamin A has not been considered, although its metabolic product retinoic acid (RA) regulates transcription via the same nuclear receptor family as TH. We hypothesized that vitamin A/RA plays a role in the neuroendocrine hypothalamus alongside TH signaling. Using a reporter assay to measure RA activity, we demonstrate that RA activity levels in the hypothalamus of photoperiod-sensitive F344 rats are reduced in short-day relative to long-day conditions. These lower RA activity levels can be explained by reduced expression of a whole network of RA signaling genes in the ependymal cells around the third ventricle and in the arcuate nucleus of the hypothalamus. These include genes required for uptake (Ttr, Stra6, and Crbp1), synthesis (Raldh1), receptor response (RAR), and ligand clearance (Crapb1 and Cyp26B1). Using melatonin injections into long-day rats, we show that the probable trigger of the fall in RA is melatonin. Surprisingly we also found RPE65 expression in the mammalian hypothalamus for the first time. Similar to RA signaling genes, members of the Wnt/ß-catenin pathway and NMU and its receptor NMUR2 are also under photoperiodic control. Our data provide strong evidence for a novel endocrine axis, involving the nutrient vitamin A regulated by photoperiod and melatonin and suggest a role for several new players in the photoperiodic neuroendocrine response.

Photoperiodic regulation of retinoic acid signaling in the hypothalamus.

Both retinoic acid (RA) and thyroid hormone (TH) regulate transcription via specific nuclear receptors. TH regulates hypothalamic homeostasis and active T3 is generated by deiodinase enzymes in tanycytes surrounding the third ventricle. However, RA has not been previously considered in such a role. Data presented here highlights novel parallels between the TH and RA synthetic pathways in the hypothalamus implying that RA also acts to regulate hypothalamic gene expression and function. Key elements of the RA cellular signaling pathway were shown to be regulated in the rodent hypothalamus. Retinoid synthetic enzymes and the retinol transport protein Stra6 were located in the cells lining the third ventricle allowing synthesis of RA from retinol present in the CNS to act via RA receptors and retinoid X receptors in the hypothalamus. Photoperiod manipulation was shown to alter the expression of synthetic enzymes and receptors with lengthening of photoperiod leading to enhanced RA signaling. In vitro RA can regulate the hypothalamic neuroendocrine peptide adrenocorticotrophic hormone. This work presents the new concept of controlled RA synthesis by hypothalamic tanycytes giving rise to possible involvement of this system in endocrine, and possibly vitamin A, homeostasis.