Differential effect of retinoic acid and triiodothyronine on the age-related hypo-expression of neurogranin in rat

https://doi.org/10.1016/j.neurobiolaging.2004.06.004Get rights and content

Abstract

Given the important role of retinoids and thyroid hormone for optimal brain functioning and the tenuous relationship between retinoic acid (RA) and triiodothyronine (T3) signalings, we compared the effects of RA or T3 administrations on RA and T3 nuclear receptors (RAR, RXR and TR) and on their target genes, neuromodulin (GAP43) and neurogranin (RC3) in 24-month-old rats.

Quantitative real time PCR and western blot analysis allowed us to verify that retinoid and thyroid signalings and GAP43 and RC3 expression are affected by age. By in situ hybridization we observed a decreased expression of RC3 in hippocampus, striatum and cerebral cortex.

RARβ, RXRβ/γ and GAP43 were up-regulated by RA as well as T3 treatment. The abundance of TRα/β mRNA and RC3 expression were only increased by T3 administration in the whole brain. This up-regulator effect of T3 on RC3 was only observed in the striatum.

During aging, T3 become a limiting factor alone able to correct the age-related concomitant hypo-activation of retinoid and thyroid signalings and alterations of synaptic plasticity.

Introduction

The roles of the retinoids, derivatives of the Vitamin A, in many physiological functions such as vision, immunity, cellular differentiation or proliferation or regulation of gene expression are at least in part well known [7], [55]. The action of Vitamin A is mediated through its active metabolite, retinoic acid (RA), by two classes of nuclear receptors: the retinoic acid receptors (RARα, β and γ) able to bind all-trans stereoisomer of retinoic acid (RA), and the retinoid X receptors (RXRα, β and γ), which bind the 9-cis stereoisomer. These receptors belong to the steroid/thyroid hormone nuclear receptor superfamily and are DNA-binding proteins [1], which, upon activation by specific retinoid ligands, induce gene transcription by interacting with distinct promoter sequences in the target genes predominantly in the form of RAR/RXR heterodimers [35], [42]. Recent research concerning the role of Vitamin A in the brain suggests that retinoid signaling plays an important role during mouse embryonic development [40], and that retinoid receptors contribute to specific functions in the adult central nervous system as well [32], [45]. The presence of the two classes of retinoid receptors in different adult mouse central nervous system areas [36] and the high level of cellular retinol and retinoic acid-binding proteins in the adult brain [61] indeed, indicate an involvement of retinoids in specific physiological brain functions. In the brain, RARβ is the main isoform expressed and RXRβ and RXRγ are also largely expressed at high levels [41]. Moreover, knockout mice for RARβ and RARβ–RXRγ display an alteration of synaptic plasticity as well as substantial performance deficits in a hippocampal-dependent spatial learning task [9]. Among the genes whose expression is regulated by retinoic acid, and in addition to those coding for their own receptors, there are genes of two identified neuron specific proteins involved in synaptic plasticity: neuromodulin or GAP43 [52], [53] and neurogranin or RC3 [34]. GAP43, a pre-synaptic PKC substrate, which is associated with the cytoplasmic face of the axonal growth-cone membrane, has been implicated in different forms of synaptic plasticity, including neurite outgrowth, regeneration and long term potentiation (LTP) [2], [25]. The post-synaptic PKC substrate, RC3, is a postnatal onset protein, which accumulates in forebrain dendritic spines where a high degree of plasticity is maintained throughout life [26]. An increased amount of the phosphorylated form of RC3 has been reported during the maintenance phase of LTP [8]. More recently, it has been shown that mice lacking the RC3 gene exhibited deficits in hippocampal synaptic plasticity and spatial learning impairment [46]. RC3 is mainly expressed in the hippocampus, the striatum and the cerebral cortex [26].

Although aging is associated with a clear decline in cognitive functions [20], [49], the structural and cellular bases of these changes remain poorly defined. Recently, evidence has been presented that aging leads to hypo-expression of retinoid signaling pathway in the brain in mice and rats. This hypo-expression may be the consequence of a decrease in retinoic acid bio-availability and is associated with an age-related reduction in neuron plasticity (characterized by hypo-expression of RC3). These studies showed that older rats and mice, which displayed lower level of brain RARβ and RXRβ/γ mRNA with respect to younger adults, also had severely and specifically impaired memory performance [16], [21]. Likewise, in the same studies, retinoic acid administration restored, to pre-senescent levels, the age-related deficits observed, confirming that a fine regulation of retinoid mediated gene expression seems fundamentally important for optimal brain function.

Data comparable to those obtained in aged animals have recently been obtained in Vitamin A deficient animals, i.e. hypo-expression of retinoid nuclear receptors and RC3 as well as memory impairment. Surprisingly, RA administration to depleted animals failed to fully restore the Vitamin A deficient-related hypo-expression of RC3, and had no effect on associated cognitive deficit [18], [22]. The hypo-activity of the retinoid pathway, induced by Vitamin A deficiency, is accompanied by a hypo-activity of T3 signaling, which becomes a limiting factor, because it impedes RA from exerting its modulator effect [32].

Numerous studies have reported the close relationship between retinoid and thyroid signaling [10], [54], [58]. Like retinoic acid, triiodothyronine (T3) mediates its effect through binding to nuclear receptors (TR), which are ligand-inducible transcription factors. RXR is the common partner for the formation of RAR, or TR functional heterodimers, indicating that the retinoid and thyroid signaling pathways converge through the direct interaction of their respective nuclear receptors [38], [60]. Interestingly, the RC3 gene is well known to be at once a T3 and RA responsive gene [44].

Interestingly, alterations in thyroid function have also been frequently reported as arising with age [12], [15]. Most of the thyroid disorders described in elderly people are related to central hypothyroidism. Indeed, serum basal and free T3 concentrations have been shown to be inversely correlated with age [13], [19].

Taking these data into account, the aim of the present paper is to investigate the involvement of thyroid signaling in the function of retinoic acid and in age-related neurobiological alterations. First, the status of aged animals related to retinoid and thyroid pathways in brain was determined. Then, effects of RA or T3 administration on RA and T3 nuclear receptor expression were compared in the whole brain of aged rats. Finally, in order to fully understand the consequences on the brain functioning, we studied the expression of neuromodulin (GAP43) and neurogranin (RC3). In the whole brain, the levels of mRNA and proteins were evaluated by using respectively real time PCR and western blot analysis. On the other hand, RC3 mRNA were also quantified by an in situ hybridization study in three regions of brain enriched in RC3: hippocampus, striatum and cerebral cortex [34].

Section snippets

Experimental design

The study was conducted in accordance with the European Community's Council Directives (861609/EEC). All the experiments conformed with the Guidelines on the Handling and Training of Laboratory Animals. Weanling male Wistar rats were purchased from Harlan (Gannat, France). They were maintained in a room with a constant airflow system, controlled temperature (21–23 °C) and a 12 h light: dark cycle. The rats had ad libitum access to food and tap water. They were randomly divided into two groups

Serum retinol and triiodothyronine concentrations

A significant reduction (−20%) in serum retinol concentration was observed in aged animals relative to controls (0.91 ± 0.06 μmol/L versus 1.16 ± 0.06 μmol/L). The concentration of triiodothyronine also significantly decreased with aging even if this reduction was less than for retinol (1.11 ± 0.02 nmol/L versus 1.00 ± 0.02 nmol/L).

Expression of retinoic acid and triiodothyronine nuclear receptors and target genes (GAP43 and RC3) in the whole brain

The results are summarized in Table 1.

A significant decrease in the level of retinoic acid nuclear receptors was observed in the brain of aged animals. The amount of

Status of aged rats

In order to study the effect of T3 on brain retinoic acid and triiodothyronine signaling in aged rats, the function of these pathways was first examined in 24 months old rats. Together, all the results obtained in this experiment established the decline in the brain of retinoid but also thyroid signaling with aging. Several authors have evoked similar results in rats and mice [16], [17]. Further data shown that knockout mice for RARβ or RARβ/RXRγ exhibited a long term potentiation (LTP) deficit

Acknowledgements

This research was supported by the Institut National de la Recherche Agronomique (INRA) and by the Conseil Régional d’Aquitaine. The authors wish to thank L. Caune for animal care.

References (61)

  • A. Guadaño-Ferraz et al.

    Transcriptional induction of RC3/neurogranin by thyroid hormone: differential neuronal sensitivity is not correlated with thyroid hormone receptor distribution in the brain

    Brain Res Mol Brain Res

    (1997)
  • P. Higueret et al.

    Retinoic acid decreases retinoic acid and triiodothyronine nuclear receptor expression in the liver of hyperthyroidic rats

    FEBS Lett.

    (1992)
  • M. Husson et al.

    Expression of neurogranin and neuromodulin is affected in the striatum of vitamin A deprived rats

    Mol Brain Res

    (2004)
  • M.A. Iñiguez et al.

    Characterization of the promoter region and flanking sequences of the neuron-specific gene RC3 (neurogranin)

    Brain Res Mol Brain Res

    (1994)
  • W. Krezel et al.

    Differential expression of retinoid receptors in the adult mouse central nervous system

    Neuroscience

    (1999)
  • M. Maden

    Role and distribution of retinoic acid during CNS development

    Int Rev Cytol

    (2001)
  • D.J. Mangelsdorf et al.

    The RXR heterodimers and orphan receptors

    Cell

    (1995)
  • S. Mani et al.

    Failure to express GAP-43 during neurogenesis affects cell cycle regulation and differentiation of neural precursors and stimulates apoptosis of neurons

    Mol Cell Neurosci

    (2001)
  • V. Pallet et al.

    Aging decreases retinoic acid and triiodothyronine nuclear expression in rat liver: exogenous retinol and retinoic acid differentially modulate this decreased expression

    Mech Aging Dev

    (1997)
  • P.A. Piosik et al.

    RC3/neurogranin structure and expression in the caprine brain in relation to congenital hypothyroidism

    Brain Res Mol Brain Res

    (1995)
  • D.E. Saunders et al.

    Reversal of alcohol's effects on neurite extension and on neuronal GAP43/B50, N-myc, and c-myc protein levels by retinoic acid

    Brain Res Dev Brain Res

    (1995)
  • A. Aranda et al.

    Nuclear hormone receptors and gene expression

    Physiol Rev

    (2001)
  • J. Bernal et al.

    Perspectives in the study of thyroid hormone action on brain development and function

    Thyroid

    (2003)
  • P. Borel et al.

    Comparison of the postprandial plasma vitamin A response in young and older adults

    J Gerontol A Biol Sci Med Sci

    (1998)
  • T. Casoli et al.

    Cellular distribution of GAP-43 mRNA in hippocampus and cerebellum of adult rat brain by in situ RT-PCR

    J Histochem Cytochem

    (2001)
  • P. Chambon

    A decade of molecular biology of retinoic acid receptors

    FASEB J

    (1996)
  • Chin WW, Yen PM. Molecular mechanisms of nuclear thyroid hormone action. In: Braverman LE, editor. Totowa;...
  • A.M. Chuo et al.

    Thyroid dysfunction in elderly patients

    Ann Acad Med Singapore

    (2003)
  • G. Cizza et al.

    Central hypothyroidism is associated with advanced age in male Fischer 344/N rats: in vivo and in vitro studies

    Endocrinology

    (1992)
  • M. Coustaut et al.

    The influence of dietary vitamin A on triiodothyronine, retinoic acid, and glucocorticoid receptors in liver of hypothyroid rats

    Br J Nutr

    (1996)
  • Cited by (28)

    • Normalization of hippocampal retinoic acid level corrects age-related memory deficits in rats

      2020, Neurobiology of Aging
      Citation Excerpt :

      Indeed, RAR and its nuclear receptors (RXR) are involved in dendritic spine formation (Chen and Napoli, 2008) and in synaptic plasticity, notably in the hippocampus (Aoto et al., 2008; Chen et al., 2014; Chiang et al., 1998). In accordance with this, we previously showed that RA in aged rodents improves synaptic plasticity, that is, long-term potentiation and expression of genes involved in synaptic plasticity (Etchamendy et al., 2001; Féart et al., 2005b). RA also facilitates neuronal survival and neurite outgrowth and induces neuronal differentiation (Chen and Napoli, 2008; Christie et al., 2010; Takahashi et al., 1999).

    • RXR heterodimers orchestrate transcriptional control of neurogenesis and cell fate specification

      2018, Molecular and Cellular Endocrinology
      Citation Excerpt :

      Mechanistically, retinoid signaling exerts its patterning function, at least in part, through the regulation of Hox genes (Marshall et al., 1996), however the complex RAR:RXR-dependent epigenomic reorganization of the Hox cluster during neurogenesis has just begun to be clarified (Kashyap et al., 2011; De Kumar et al., 2015). In addition to the essential role of RA in neurogenesis, retinoids have been also shown to be involved in the maintenance of neuronal plasticity in the olfactory system and in the hippocampus and downregulation of RAR expression was detected during aging, resulting in impairment in spatial memory (Enderlin et al., 1997; Feart et al., 2005). An other heterodimer implicated in the CNS, LXR:RXR has been mainly implicated in the control of cholesterol homeostasis, lipogenesis, and inflammation.

    • Glycerophospholipids and glycerophospholipid-derived lipid mediators: A complex meshwork in Alzheimer's disease pathology

      2011, Progress in Lipid Research
      Citation Excerpt :

      In fact, several studies show that DHA can specifically bind to and directly activate RXR, leading to the regulation of the expression of genes usually under the control of retinoic acid [141–143]. These results are potentially important for the treatment or prevention of AD since the retinoic acid signaling pathway is involved in the maintenance of synaptic plasticity and memory in aged animals [144–147]. Indeed, a link between the metabolism of retinoids and late-onset AD was recently proposed [148,149].

    View all citing articles on Scopus
    View full text