Co-localization of brain corticosteroid receptors in the rat hippocampus.

New developments in corticosteroid receptor research enabled us to perform a highly detailed study on the neuroanatomical topography of MR and GR in the rat hippocampus. Receptor immunocytochemistry was used to map the distribution of GR protein with the help of a monoclonal antibody raised against the purified rat liver GR-hormone complex. Furthermore, in situ hybridization with 35S-labeled RNA probes, which were transcribed from cDNAs complementary to either a fragment of the rat brain MR gene or to the rat liver GR gene, was applied to investigate the localization of MR and GR mRNA in the limbic brain. The pyramidal neurons of cell field Ca1 and CA2 and the granular neurons of the dentate gyrus showed marked GR immunoreactivity (GRir) as well as intense labeling of GR mRNA. The radiolabeled density of GR mRNA in cell fields CA3 and CA4 was considerable less, whereas low-to-almost-undetectable levels of GRir could be observed in these regions. MR mRNA appeared to be evenly distributed over all cell fields of the hippocampus and the dentate gyrus. The topography of GRir, GR mRNA and MR mRNA was found to agree with the cellular distribution of MR and GR binding sites in the hippocampus. Moreover, the microanatomy of MR and GR in the hippocampus appeared to overlap. Our data strongly suggest that MR and GR are co-expressed in the majority of pyramidal and granular neurons of the hippocampal formation. This assumption is based on coherence in the detection of different aspects of the receptor cycle of MR and GR.(ABSTRACT TRUNCATED AT 250 WORDS)     

Sub-neurotoxic neonatal anoxia induces subtle behavioural changes and specific abnormalities in brain group-I metabotropic glutamate receptors in rats

Anoxia in the first week of life can induce neuronal death in vulnerable brain regions usually associated with an impairment of cognitive function that can be detected later in life. We set-up a model of subneurotoxic anoxia based on repeated exposures to 100% nitrogen during the first 7 days of post-natal life. This mild post-natal exposure to anoxia specifically modified the behaviour of the male adult rats, which showed an attention deficit and an increase in anxiety, without any impairment in spatial learning and any detectable brain damage (magnetic resonance imaging and histological analysis). Post-anoxic rats showed a reduction in the expression of group-I metabotropic glutamate receptors (i.e. mGlu1 and mGlu5 receptors) in the hippocampus and cerebral cortex, whereas expression of the mGlu 2/3 receptors, the NR1 subunit of NMDA receptors, and the GluR1 subunit of alpha-amino-3-hydroxy-5-methylisoxazole-4-propionate (AMPA) receptors was unchanged. mGlu1 and mGlu5 receptor signalling was also impaired in postanoxic rats, as revealed by a reduced efficacy of the agonist (1S,3R)-1-Aminocyclopentane-1,3-dicarboxylic acid (1S,3R-ACPD) to stimulate polyphosphoinositide hydrolysis in hippocampal slices. We conclude that rats subjected to subneurotoxic doses of anoxia during the early post-natal life develop behavioural symptoms that are frequently encountered in the inattentive subtype of the attention deficit hyperactivity disorder, and that group-I mGlu receptors may be involved in the pathophysiology of these symptoms.     

Changes in glucocorticoid receptors in different regions of brain of immature and mature male rats

Specific cytosolic binding for synthetic glucocorticoid dexamethasone was studied in several brain regions (hypothalamus, hippocampus, caudate nucleus, cerebellum, cerebral cortex) of immature (3-week) and mature (26-week) male rats, intact and adrenalectomized. A significant regional difference was observed in the concentration of in vitro [3H] dexamethasone binding in the brain of adrenalectomized rats at both ages, with the highest levels in the hippocampus. A marked decrease in specific binding was observed in all brain regions of adrenalectomized mature rats as compared to immature. The dexamethasone binding was significantly lower in all brain regions of normal intact animals as compared to adrenalectomized rats in both ages.     

On the role of glucocorticoid receptors in brain plasticity

Summary 1. The mapping of glucocorticoid receptors (GR) in the rat central nervous system (CNS) has demonstrated their widespread presence in large numbers of nerve and glial cell populations also outside the classical stress regions.2. The present paper summarizes the evidence that glucocorticoids via GR in the CNS can act as lifelong organizing signals from development to aging. The following examples are given. (a) In the prepubertal and adult offspring, prenatal corticosterone treatment can produce long-lasting changes in striatal dopaminergic communication. (b) In adulthood, the evidence suggests complex regulation by adrenocortical hormones of neurotrophic factors and their receptors in the hippocampal formation. (c) In aging, the strongly GR-immunoreactive pyramidal cell layer of the CA1 hippocampal area appears to be preferentially vulnerable to neurotoxic actions of glucocorticoids, especially in some rat strains.3. Strong evidence suggests that each nerve cell in the CNS is supported by a trophic unit, consisting of other nerve cells and glial cells, blood vessels, and extracellular matrix molecules. Due to multiple actions on nerve and glial cell populations of the different trophic units, the glucocorticoids may exert either an overall trophic or a neurotoxic action. It seems likely that with increasing age, the endangering actions of glucocorticoids on nerve cells prevail over the neurotrophic ones, leading to reduced nerve cell survival in some trophic units.     

Corticosterone regulation of brain and lymphoid corticosteroid receptors

Circulating lymphocytes are often used as a model for brain corticosteroid receptor regulation in clinical disease states, although it is not known if lymphoid receptors are regulated in a similar manner as brain receptors. In the present study the regulation of brain (hippocampus, frontal cortex, hypothalamus and striatum), lymphoid (circulating lymphocytes, spleen and thymus) and pituitary glucocorticoid receptors in response to alterations in circulating corticosterone levels was examined. Seven days following adrenalectomy, type II corticosteroid receptors (i.e. glucocorticoid receptors) were significantly increased in the hippocampus, frontal cortex and hypothalamus, but not in any other tissues. Administration of corticosterone (10 mg/kg) for 7 days significantly decreased type II as well as type I (i.e. mineralocorticoid receptors) receptors in the hippocampus. Type II receptors in the frontal cortex, circulating lymphocytes and spleen were also significantly decreased by chronic corticosterone treatment. Immobilization stress (2 h a day for 5 days) failed to alter receptor density in any of the tissues. These results demonstrate that homologous regulation of corticosteroid receptors by corticosterone does not invariably occur in all tissues and emphasize the complex degree of regulation of these receptors. However, the simultaneous downregulation of both hippocampal and lymphocyte glucocorticoid receptors by corticosterone provides support for the hypothesis that circulating lymphocytes do reflect some aspects of brain glucocorticoid receptor regulation.     

Developmental changes of glucocorticoid receptors in the rat pancreas

Glucocorticoids are known to play a role in the maturation of the exocrine pancreas. The exact mechanism of glucocorticoid action in pancreatic ontogeny is, however, not clear. The present study characterized and quantitated the binding of [3H]dexamethasone to cytosol fractions from pancreata of rats at various ages. Trunk blood samples from these rats were also checked for levels of free and bound corticosterone. Specific and saturable bindings for dexamethasone were found in pancreatic cytosol fractions from newborn suckling and adult rats. Competition studies showed a preference for steroids with glucocorticoid activity. Specific binding was relatively low in pancreatic cytosol from newly born and 1-day old pups. A significant rise was seen after day 15. Cytosolic binding capacities were greatest from pancreata obtained from pups at weaning (3rd to 5th weeks). Values then declined toward the adult level. Scatchard analysis revealed a single class of binding sites with a dissociation constant (Kd) of 7.3 (+/- 1.1) X 10(-8) M and number of binding sites equalled to 1.29 (+/- 0.18) X 10(-13) mole/mg of cytosolic protein in adult rat pancreas. Pancreata from 25- and 15-day old rats had Kds of 3.4 (+/- 0.8) X 10(-8) M and 2.7 (+/- 0.7) X 10(-8) M with the number of binding sites equal to 1.77 (+/- 0.21) X 10(-13) mole/mg protein and 1.31 (+/- 0.16) X 10(-13) mole/mg protein respectively. Total plasma corticosterone concentration was low before day 10. It rose significantly by day 15, peaked at day 25, and then declined after weaning. About 5-15% of corticosterone during weaning and about 20-30% before and after weaning were in the free form. The peak level of dexamethasone binding corresponded to an increase in the plasma corticosterone level during weaning. This suggests a close relationship between plasma corticosterone levels and pancreatic glucocorticoid receptors. Both may, therefore, play a role in pancreatic development in the rat.     

Ginsenoside RG1 and corticosteroid receptors in rat brain

Old (28 months) male Wistar rats were treated chronically for two weeks with ginsenoside Rg1 or with vehicle delivered via sc implanted Alzet mini-pumps (rate of ginsenoside release 2.4 micrograms/0.5 microliter/h). The number of Type 1 corticosterone-preferring receptor sites (CR) and Type 2 glucocorticoid receptors (GR) was measured in the cytosol of hippocampus tissue of rat brain with an in vitro binding assay. In old rats the Bmax of Type 1 CR and Type 2 GR was reduced by 51.5% and 28.3% respectively. Following the two week treatment with Rg1 the Bmax of Type 1 CR increased by 60% and a receptor concentration was reached which was 21% lower than that observed in the young control animals. Minor differences in affinity of steroid binding to both receptor systems were observed in the groups of rats. The possible binding of ginsenosides to brain corticosteroid receptors in vitro was investigated as well. The inclusion of a 500 fold molar excess of Rg1 in hippocampus cytosol did not displace 3H-corticosterone from its soluble receptor sites. The affinity of Rg1 with these sites in vitro is therefore negligible. In conclusion, the binding capacity of Type 1 CR and Type 2 GR is reduced in the hippocampal brain region of aged rats. Upon chronic infusion of ginsenoside Rg1, only Type 1 CR capacity is restored towards the level observed in young control animals. This finding suggests that in old rats the ginsenoside enhances the CORT signal via Type 1 CR on the function of the hippocampus, which is a limbic brain structure involved in cognition, mood and affect.     

Age-related changes in the dog hypothalamic-pituitary-adrenocortical system: neuroendocrine activity and corticosteroid receptors.

Aging is associated with a progressive dysfunctioning of the hypothalamic-pituitary-adrenocortical (HPA) axis. We have studied the response of the HPA axis to stress and a hormonal (ovine corticotropin releasing factor (o-CRF) challenge in young (1.5-2 years) and aged (greater than 11 years) dogs. Compared to the young dogs, the aged animals displayed an increased basal concentration of both ACTH and cortisol. In addition, in response to an o-CRF challenge (1 microgram/kg i.v.) or an electric footshock (1 mA, alternatively on/off for 2 s) or immobilization (45 min) stress, the aged dogs showed significantly larger increments in ACTH and cortisol. Following the challenge test, the young and aged dogs reached their respective basal hormone levels at the same time, except for the o-CRF test. In the latter case, in contrast to the young controls, the aged dogs still showed an increased plasma cortisol level compared to the pre-challenge basal hormone concentration. Concerning the effect of aging on the brain and hypophyseal corticosteroid receptors, a selective decline (minus 50-75%) in mineralocorticoid receptor (MR) was observed in all measured brain regions (dorsal and ventral hippocampus, septum, hypothalamus) and anterior pituitary, whereas no change was found in brain glucocorticoid receptor (GR) number. The GR level in the anterior pituitary was even increased by 70%. In light of the role that MR and GR seem to play in the regulation of the HPA axis, it is concluded that the diminished MR number in the aged dog brain may underly the increased basal hormone levels and the elevated responsiveness of the HPA axis in these animals. The observation that the stress-induced elevations of cortisol and ACTH were not prolonged at senescence suggests that the GR-mediated negative feedback action of glucocorticoids is not altered, which is in line with the unchanged brain GR numbers in the aged dogs.     

Estrogens up-regulate type I and type II glucocorticoid receptors in brain regions from ovariectomized rats

The effects of 1-4 days of estradiol (E2) treatment on type I and type II glucocorticoid receptors (GCR) were determined in cytosolic fractions from brain regions of ovariectomized rats. Four days after E2 administration, type I GCR increased in septum, amygdala, hypothalamus and hippocampus, but decreased in the anterior pituitary. Type II GCR increased in septum and hypothalamus only. For both receptor types, changes occurred earlier in septum (1 day) than in the other regions. The E2 increment was due to an increase in Bmax, without changes in Kd. The up-regulation of type II GCR by E2 was also confirmed immunocytochemically in four nuclei of the septal area. In a parallel study, E2 receptors were determined in nuclear and cytosol fractions from the same regions analyzed for GCR. In rats receiving E2, estrogen receptors decreased in cytosol and increased in nuclei from septum, amygdala, hypothalamus and anterior pituitary, but did not change in hippocampus. The results suggest that GCR in certain neuroendocrine regions are regulated by E2, without taking into account whether the areas involved contain high (anterior pituitary), moderate (septum, hypothalamus, amygdala) or low (hippocampus) levels of E2 receptors. Our model may shed light on sex differences in GCR and on E2 regulation of glucocorticoid action in brain and the pituitary.     

Specific hydroxylations determine selective corticosteroid recognition by human glucocorticoid and mineralocorticoid receptors

The ligand binding domains of the human mineralocorticoid receptor (hMR) and glucocorticoid receptor (hGR) display a high sequence homology. Aldosterone and cortisol, the major mineralocorticoid and glucocorticoid hormones, are very closely related, leading to the cross-binding of these hormones to both receptors. The present study reports on the mechanism by which hMR and hGR are activated preferentially by their cognate hormones. We found that the ability of corticosteroids to stimulate the receptor's transactivation function is depending on the stability of the steroid-receptor complexes. In the light of a hMR structural model we propose that contacts through the corticosteroid C21 hydroxyl group are sufficient to stabilize hMR but not hGR and that additional contacts through the C11- and C17-hydroxyl groups are required for hGR.     

Effects of brain natriuretic peptide on pituitary-adrenal activation in rats

The aim of this study was to characterize the effects of brain natriuretic peptide (BNP) on the hypothalamo-pituitary-adrenal (HPA) responses to different stress paradigms (ether stress, electric shock and restraint). Rats were subjected to the stressful stimuli after intracerebroventricular administration of BNP (32.5 ng-6.5 microg) and plasma corticosterone was used as an indicator of the HPA activation. BNP did not modify the basal secretion, but inhibited the stress-induced rise in plasma corticosterone in a dose-dependent manner. BNP proved most effective in decreasing the corticosterone response to ether stress and attenuated the electric shock and restraint-induced HPA activation to a lesser extent. These results confirm the view that BNP takes part in the regulation of the HPA system.     

Endocrine-mediated parallel changes in hepatic glucocorticoid and prolactin receptors

The objective of this study was to determine whether in female rat liver any relationship existed between prolactin and glucocorticoid receptors after hormonal manipulation. Bromocryptine (CB-154) treatment of adult SD female rats (80-100 days old) for 48 h decreased prolactin binding to hepatic membranes 49% and dexamethasone binding in hepatic cytosol 40% below control values. Administration of rat prolactin along with bromocriptine prevented these changes. In another study, prolactin binding to hepatic membranes increased 53% and dexamethasone binding in hepatic cytosol increased 113% above sham-control values, 3 days after adrenalectomy. On the other hand, hydrocortisone treatment of sham-operated rats reduced prolactin binding by 57% and dexamethasone binding by 76%. Scatchard analyses of the prolactin or dexamethasone binding data indicated that these manipulations changed the number of prolactin or dexamethasone binding sites rather than their apparent affinity constants. In vitro treatment of rat whole liver homogenate with various doses (10(-9) - 10(-5) M) of dexamethasone and corticosterone for 15 min at 22 degrees C resulted in a dose-dependent decrease in prolactin binding activity. However, direct addition of dexamethasone to a hepatic 15 000 X g to 100 000 X g membrane preparation exhibited no significant effects on prolactin binding. In conclusion, these studies show that (a) there is a parallel in vivo modulation of rat liver prolactin and glucocorticoid receptors under various experimental conditions and (b) in vitro exposure of whole liver homogenate to glucocorticoids inhibits the prolactin binding activity.     

脊椎动物脑发育中糖皮质激素受体功能的研究进展

糖皮质激素受体（glucocorticoid reccptor,GR）广泛分布在脊椎动物中枢神经系统的多个组织区域中,而且结构及功能保守。在与激素结合的状态下,受体能够特异性地与靶基因的启动子结合影响基因的表达,或通过激活G蛋白偶联的信号途径引起神经递质的释放。外界环境刺激和外源糖皮质激素暴露都能改变GR在脑中的表达,并对神经的发育及功能产生影响,同时也对学习、记忆以及情感等高级神经活动和行为起到重要的作用。该文对脊椎动物糖皮质激素受体的结构和在脑中的分布,以及对神经发育和功能的影响及其中的分子机制的最新研究进展进行综述。     

Developmental changes in the level of glucocorticoid receptors in chick-embryo tissues

The concentrations of glucocorticoid receptors were assayed in various chick embryo tissues by a cytosol charcoal-dextran method using [3H]dexamethasone as ligand. The highest levels of receptors were found in muscle on developmental day 15-16. The order of maximum binding of dexamethasone in various tissues was muscle greater than heart greater than skin greater than tendon greater than kidney greater than cartilage greater than liver. However, marked variation in the level of receptors was found even in the same tissue during development of the chick-embryo. The highest levels of receptors were generally found on days 15-17. For example, in tendon the difference between the highest and lowest level of receptors was about 6-7 fold. Thus the results show that specific glucocorticoid receptors may be found in various tissues of developing chick-embryos.