The mechanism of ligand-induced (homologous) down-regulation of the glucocorticoid receptor (GR) has been studied. Dexamethasone caused a down-regulation of the levels of GR mRNA and protein both in hepatoma tissue culture cells and rat liver in vivo. The decrease in the level of rat liver GR mRNA was due to a reduced transcription rate of the GR gene, as assessed by nuclear run-on transcription experiments. The half-life of GR mRNA in hepatoma tissue culture cells was determined to be approximately 4.5 h and was unaffected by dexamethasone. In addition to the transcriptional regulation of GR gene expression, a dexamethasone-dependent posttranslational modification in the rate of GR protein turnover was observed. In the absence of dexamethasone, GR protein half life was approximately 25 h whereas it decreased to approximately 11 h in the presence of hormone. Down-regulation of GR protein occurred with a 6- to 24-h delay as compared to the decline in GR mRNA. This is most likely due to the differences in half-lives of GR mRNA and protein, respectively. Our results suggest that auto-regulation of GR by its cognate ligand is complex and occurs at both transcriptional and posttranslational levels.

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The localization and distribution of mRNA encoding the glucocorticoid receptor (GR) was investigated in tissue sections of the adult male rat brain by in situ hybridize. tion and RNA blot analysis. GR mRNA levels were measured by quantitative autoradiography with IS-and 32P-labeled RNA probes, respectively. Strong labeling was observed within the pyramidal nerve cells of the CAl and CA2 areas of the hippocampal formation, in the granular cells of the dentate gyrus, in the parvocellular nerve cells of the paraventricular hypothalamic nucleus, and in the cells of the arcuate nucleus, especially the parvocellular part. Moderate labeling of a large number of nerve cells was observed within layers II, III, and VI of the neocortex and in many thalamic nuclei, especially the anterior and ventral nuclear groups as well as several midline nuclei. Within the cerebellar cortex, strong labeling was observed all over the granular layer. In the lower brainstem, strong labeling was found within the entire locus coeruleus and within the mesencephalic raphe nuclei rich in noradrenaline and 5-hydroxytryptamine cell bodies, respectively. A close correlation was found between the distribution of GR mRNA and the distribution of previously described GR immunoreactivity. These studies open the possibility of obtaining additional information on in vivo regulation of GR synthesis and how the brain may alter its sensitivity to circulating glucocorticoids.Adrenal steroids have a wide range of actions in the central nervous system, where they have been shown to affect growth and differentiation of nerve cells as well as mental state, perception, learning, and sleep (for a review, see ref.1). Glucocorticoids, as well as other steroid hormones, exert at least a part of their action via binding to specific intracellular receptor proteins, acting as modulators of the transcriptional activity of specific networks of genes (for a review, see ref.2). Corticosterone binds to two distinct receptor systems in the rat brain (4)-the mineralocorticoid-receptor-like type I receptor and the classical glucocorticoid receptor (GR). Recently, it has been possible to map out GR immunoreactive nerve cell populations (5-9) and glucocorticoid recognition sites (10, 11) in many areas of the central nervous system by means of mouse monoclonal antibodies against the rat liver GR (12) and specific GR radioligands, respectively. High levels of glucocorticoid binding (11) and strong GR immunoreactivity (5, 8) were found within the monoaminergic neurons, in the hippocampal formation, in the corticotropinreleasing factor immunoreactive neurons, and in the mediobasal hypothalamus.Studies performed with ligand binding and quantitative immunocytochemistry techniques indicate that cellular GR levels vary as a result of, e.g., endocrine manipulations (13), stress (14, 15), and aging (16,17). By means of an in situ hybridization technique, it has been possible in the present paper to detect and map out the distribution of GR mRNA in the male rat brain. These results ...

A cDNA clone for the rat glucocorticoid receptor (GR) was used to study mechanisms of GR mRNA regulation. Treatment of rat hepatoma culture cells with 0.5 MAM dexamethasone caused a small, initial increase in the GR mRNA level after 6 hr as well as a 50% to 95% reduction of the GR mRNA level after 24 hr of incubation when studied by RNA blot hybridization. After 72 hr, the initial GR mRNA level was restored. The down-regulation of GR mRNA levels appears to be independent of protein synthesis, since it also was observed in the presence of cycloheximide. However, cycloheximide caused a 4-fold increase in intracellular levels of GR mRNA.Using an immunoprecipitation assay, we could demonstrate that the GR specifically interacts with a GR cDNA clone, which represents a 2.6-kilobase fragment of the 3' nontranslated region of the GR mRNA. RESULTSTo investigate the effect of dexamethasone on cellular GR mRNA levels, HTC cells were grown in a logarithmic phase for various times in the absence or presence of 0.5 ,uM dexamethasone,-whereafter GR mRNA was analyzed by RNA blot-hybridization analysis (Fig. 1). A reduction by a factor of =*15 in the amount of a single =7-kb GR mRNA species was seen in this experiment at 24-48 hr after treatment with dexamethasone. This decrease occurred after a slight (-2-fold) but significant increase in the GR mRNA level after 6 hr of incubation. Although the down-regulation of GR mRNA after hormone treatment for 24 hr was a consistent observation in all experiments, the degree of down-regulation caused by treatment with 0.5 AM dexamethasone varied between 50% and 95%. After 72 hr of incubation, the initial level of GR mRNA was restored (Fig. 1). The changes in the amount of GR mRNA were evaluated in relation to the same amount of total cellular RNA and to the content of 3-actin mRNA, which only exhibited slight changes (15-20%) upon hormone treatment (Fig. 1). A similar degree of down-regulation (50% to 90%o) of cellular GR mRNA was also seen in vivo in livers of rats treated with 4 mg of dexamethasone per kg of body weight for 4 days (data not shown).To investigate the importance of protein synthesis for down-regulation of GR mRNA by dexamethasone, experiments in HTC cells were performed as above in the absence or presence of the inhibitor of translation, cycloheximide. a b c d

Glucocorticoids play an important role in the normal regulation of bone remodeling; however continued exposure of bone to glucocorticoid excess results in osteoporosis. In vivo, glucocorticoids stimulate bone resorption and decrease bone formation, and in vitro studies have shown that while glucocorticoids stimulate osteoblastic differentiation, they have important inhibitory actions on bone formation. Glucocorticoids have many effects on osteoblast gene expression, including down-regulation of type I collagen and osteocalcin, and up-regulation of interstitial collagenase. The synthesis and activity of osteoblast growth factors can be modulated by glucocorticoids as well. For example, insulin-like growth factor I (IGF-I) is an important stimulator of osteoblast function, and expression of IGF-I is decreased by glucocorticoids. The activity of IGF I can be modified by IGF binding proteins (IGFBPs), and their synthesis is also regulated by glucocorticoids. Thus, glucocorticoid action on osteoblasts can be direct, by activating or repressing osteoblast gene expression, or indirect by altering the expression or activity of osteoblast growth factors. Further investigation of the mechanisms by which glucocorticoids modulate gene expression in bone cells will contribute to our understanding of steroid hormone biology and will provide a basis for the design of effective treatments for glucocorticoid-induced osteoporosis.

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Monoclinic bismuth vanadate (BiVO4), as a rising star in light-catching materials, has been researched in many fields, such as photoelectrochemical water splitting.

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