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Endocrinology Vol. 148, No. 6 2716-2722
Copyright © 2007 by The Endocrine Society

Hippocampal 11ß-Hydroxysteroid Dehydrogenase Type 1 Messenger Ribonucleic Acid Expression Has a Diurnal Variability that Is Lost in the Obese Zucker Rat

Jonas Burén, Sven-Anders Bergström, Edmund Loh, Ingegerd Söderström, Tommy Olsson and Cecilia Mattsson

Department of Public Health and Clinical Medicine, Umeå University Hospital, SE-901 85 Umeå, Sweden

Address all correspondence and requests for reprints to: Jonas Burén, Ph.D., Department of Public Health and Clinical Medicine, Building 6M, 4th floor, Umeå University Hospital, SE-901 85 Umeå, Sweden. E-mail: jonas.buren{at}medicin.umu.se.


   Abstract
Top
 Abstract
Introduction
 Materials and Methods
 Results
 Discussion
 References
 
Circulating levels of glucocorticoids show a circadian rhythm. Obesity is associated with a flattening of the diurnal rhythm; plasma cortisol levels are slightly increased during the trough, although they are normal or low in the morning. Studies in humans and in leptin-resistant Zucker rats suggest that tissue-specific alterations in glucocorticoid exposure might play a key role for development of obesity and obesity-associated dysregulation of the hypothalamic-pituitary-adrenal axis. We hypothesized that there is a circadian rhythm in prereceptor metabolism of glucocorticoids exerted by 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) in brain and/or peripheral tissues (liver, fat, and muscle) that might be abrogated in obesity. The present study demonstrates a circadian rhythm in 11ß-HSD1 mRNA expression (35–45% increase at morning vs. evening, P < 0.05) in dentate gyrus granular layer and CA1 subregions of the hippocampus in lean Zucker rats that was lost in the obese rats. Sprague Dawley rats also revealed a diurnal rhythm in hippocampal 11ß-HSD1 mRNA expression. There was no circadian variation in 11ß-HSD enzyme activity in peripheral tissues, although obese Zucker rats had a decreased enzyme activity in liver and epididymal fat (by ~40%, P < 0.05) compared with lean rats. In Sprague Dawley rats, 11ß-HSD activity in adipose tissue was higher in retroperitoneal and epididymal vs. sc fat (P < 0.001). In summary, obese Zucker rats lack a circadian rhythm of 11ß-HSD1 gene expression in the hippocampus, which may contribute to increased activity of the hypothalamic-pituitary-adrenal axis and altered diurnal variation of circulating corticosterone levels.


   Introduction
Top
 Abstract
 Introduction
Materials and Methods
 Results
 Discussion
 References
 
MANY FEATURES OF Cushing’s syndrome, e.g. high circulating cortisol levels due to ACTH- or cortisol-producing tumors, are found in simple obesity. However, circulating glucocorticoid (GC) levels have not been shown to be constantly elevated. This conundrum may be attributed to a subtle increase in basal levels of circulating cortisol and/or increased tissue-specific GC exposure (1, 2). Prereceptor cortisol concentrations are regulated by the intracellular enzyme 11ß-hydroxysteroid dehydrogenase type 1 (11ß-HSD1) regenerating inactive cortisone (or 11-deoxycorticosterone in rodents) to active cortisol (or corticosterone). It has been repeatedly shown that obese humans have increased enzyme activity and gene expression of 11ß-HSD1 in sc abdominal fat, whereas hepatic 11ß-reduction of cortisone is reduced (3, 4, 5, 6). Monogenic animal models of obesity, such as the leptin-deficient Lepob mice and the leptin-resistant Zucker rat, show similar tissue-specific alterations in 11ß-HSD1 activity (7, 8). The physiological relevance of 11ß-HSD1 dysregulation has been demonstrated by transgenic mice overexpressing 11ß-HSD1 in adipose tissue. These mice develop central obesity, insulin resistance, hyperlipidemia, and hypertension (9, 10). In contrast, 11ß-HSD1 knockout mice seem protected from these metabolic derangements when challenged by high-fat feeding (11, 12, 13).

Circulating levels of GCs follow a diurnal pattern in humans with a peak in early morning and low levels during the evening and night. This cyclic secretion is controlled by the hypothalamus-adrenal-pituitary (HPA) axis, driven by the hypothalamic suprachiasmatic nucleus, and regulated in a negative feedback loop by the binding of circulating GC to glucocorticoid and mineralocorticoid receptors (GR and MR, respectively) (14). Human obesity seems to be associated with a flattening of the diurnal rhythm; trough plasma cortisol levels are slightly increased (15, 16), whereas peak levels are normal or low (15, 17, 18). In the obese Zucker rat, trough levels of corticosterone are clearly elevated, whereas peak levels do not differ between the lean and obese phenotype (19, 20, 21). Both obese humans and Zucker rats have an increased excretion rate of GCs, indicating an increased activity of the HPA axis (7, 22, 23, 24, 25). This may be due to an increased cortisol clearance (26, 27, 28) and/or a partial central resistance to GC feedback (29).

Hippocampal GR expression shows a circadian pattern (30), and a diurnal rhythm in glucocorticoid sensitivity is present in whole blood cells (31). Notably, the fatty Zucker rat has a reduced 11ß-HSD1 and MR, but not GR, expression in the hippocampus that at least partly may explain the elevated trough plasma corticosterone concentration (32). Yet, it is not clear why peak levels of corticosterone do not differ between the phenotypes. Our hypothesis was that 11ß-HSD1 enzyme expression and/or activity has a diurnal variability in brain and/or peripheral tissues (liver, fat, and muscle) that is disrupted in obesity and contributes to the metabolic changes and the altered diurnal rhythm of circulating GC levels.


   Materials and Methods
Top
 Abstract
 Introduction
 Materials and Methods
Results
 Discussion
 References
 
Animals and experimental design
The experimental protocol was approved by the Ethics Committee for Animal Research at Umeå University. Eight lean (228 ± 8 g) and eight obese (334 ± 23 g) male 7- to 8-wk-old Zucker rats (Charles River, Sulzfeld, Germany) and 18 male 7- to 8-wk-old Sprague Dawley (S-D) (239 ± 15 g) rats (B&K Universal Lab, Sollentuna, Sweden) were housed in groups of three to four in standard laboratory cages until 5 d before the start of the experiment when they were singly housed. The animals had free access to water and food (standard rat chow) with lights on at 0600 h and off at 1800 h. Four lean and four obese Zucker rats and nine S-D rats were decapitated at 0800–0900 h (morning groups) and 2000–2100 h (evening groups), respectively. All rats were killed within 30 sec of initiation of contact. Trunk blood was collected in EDTA tubes, and plasma samples were stored at –80 C until corticosterone assay. Liver (right lobe), fat (retroperitoneal, epididymal, and sc depots) and skeletal muscle (quadriceps) were dissected and snap-frozen in liquid nitrogen. The brains were removed, immediately frozen on dry ice, and stored at –80 C. Coronal cryostat sections (10 µm) at the level of the dorsal hippocampus (33) were thaw-mounted onto Superfrost Plus microscope slides (Menzel-Gläzer, Braunschweig, Germany). Slides were stored at –80 C.

Corticosterone assay
Plasma corticosterone was measured using a RIA kit (Rat corticosterone 125I assay system; Amersham Pharmacia Biotech, Uppsala, Sweden) according to company instructions. The sensitivity of the assay was 0.06 µg/dl, and the intra- and interassay coefficients of variation for corticosterone were 5 and 4–6%, respectively.

In situ hybridization
In situ hybridization was performed essentially as previously described (34). Antisense and sense cRNA probes for 11ß-HSD1 were generated by in vitro transcription in the presence of [35S]UTP (Amersham Biosciences, Uppsala, Sweden) using nucleotides of approximately 600 base pairs of rat 11ß-HSD1 cDNA, subcloned into Bluescript vector, as templates. The plasmid was linearized with StyI and then transcribed using T3 RNA polymerase.

Slides were fixed (4% paraformaldehyde in 0.1 mM phosphate buffer), acetylated, and dehydrated in graded ethanol before hybridization. The prehybridization mixture containing the nucleic acid was denatured, and then the hybridization was performed using 200 µl hybridization solution [50% formamide, 0.6 M NaCl, 20 mM Tris-HCl (pH 7.5), 1x Denhardt’s solution, 1 mM EDTA, 0.2 mg/ml salmon sperm DNA, 10% dextran sulfate, 0.5 mg/ml yeast tRNA, and 0.1 M dithiothreitol] onto each slide and incubated for 2 h at 50 C. All in situ hybridizations were then performed using 200 µl hybridization solution per slide containing 1.0 x 106 cpm of denatured riboprobe. Slides were hybridized overnight at 50 C in humidified sealed boxes.

After hybridization, slides were washed with 1x standard saline citrate (SSC)/0.01% SDS at 60 C before incubation with RNase A (30 µg/ml) for 1 h at 37 C followed by 1x SSC/0.01% SDS at 60 C and finally washed with 0.1x SSC/0.01% SDS at 60 C. Slides were then dehydrated in graded ethanol (in 0.3 M NH4Ac) and air dried before exposure to Biomax MR-1 (Amersham) for 1 wk at room temperature and developed in Agfa Curix 60 (Mortsel, Belgium). Slides were then dipped in NTB-2 emulsion (Eastman Kodak Co., Rochester, NY) and exposed for 4 wk at 4 C and thereafter developed and counterstained with 1% pyronin Y.

Quantification of in situ hybridization
The mRNA expression was quantified by counting silver grains over individual neurons under bright-field conditions using a computerized image analysis system (AnalySIS Pro; Soft Imaging System Gmbh, Münster, Germany). Within the hippocampus, the dentate gyrus (granular and molecular layer) and CA1 subregions were studied; also, the overlying cortex (layer 4/5) was investigated. Background counted over areas of white matter was subtracted. All measurements were performed blind to the experimental conditions.

11ß-HSD enzyme activity
11ß-HSD activity was measured in the dehydrogenase direction, which is more stable in vitro than the reductase direction (35). 11ß-HSD activity measurements were performed essentially as previously described (7). Tissue samples were homogenized in ice-cold homogenization buffer [10% glycerol 300 mM NaCl, 1 mM EDTA, 50 mM Tris (pH 7.7)] and dithiothreitol (Sigma, Steinheim, Germany) and centrifuged at 4 C. Samples of liver (0.01 mg protein/ml), fat (0.5 mg protein/ml), and muscle (1.5 mg protein/ml) were incubated in duplicate at 37 C containing an excess (2 mM) of the 11ß-HSD-specific cofactor nicotinamide adenine dinucleotide phosphate (Sigma-Aldrich, Steinheim, Germany) and 50–100 nM [1,2,6,7-3H]corticosterone (Amersham Biosciences, Little Chalfont, UK). After 30 (liver) or 60 (fat and muscle) minutes of incubation, samples were put on ice and steroids were extracted with ethyl acetate, dried, and dissolved in ethanol. Protein concentration and incubation times were optimized such that activity was in the linear range for product formation. Steroids were separated by thin-layer chromatography (mobile phase, 92% chloroform and 8% ethanol, 20 x 20 cm silica gel 60 F254; Merck, Darmstadt, Germany), exposed to a phosphorimager tritium screen, and quantified by a phosphor image scanner (Typhoon 9400; Amersham Biosciences Biotech Inc., Piscataway, NJ). 11ß-HSD activity was expressed as percent conversion after correction for unspecific conversion without homogenate (usually ~2–3%).

RNA extraction
Total RNA was extracted from liver, muscle, and adipose tissue using the RNeasy Lipid Tissue Mini Kit (QIAGEN, Hilden, Germany). The RNA concentration was determined by spectrophotometer (ND-1000 spectrophotometer; NanoDrop Technologies, Inc., Wilmington, DE). The integrity of the RNA was verified by ethidium bromide staining of rRNA bands separated on a denaturing 1% agarose gel.

Real-time RT-PCR
Two micrograms of RNA from each liver, muscle, and adipose tissue sample were reverse transcribed, using TaqMan RT reagents (High Capacity cDNA Archive Kit, Applied Biosystems, Foster City, CA) and RNase inhibitor (Applied Biosystems) at a final concentration of 1.0 U/ml. Subsequently, specific mRNAs were quantified (relative quantification standard curve method), using TaqMan gene expression assays (Applied Biosystems) for 11ß-HSD1 (Rn00567167_m1) as well as 18S rRNA (Hs99999901_s1) and Taqman Universal PCR Master Mix (Applied Biosystems) according to the manufacturer’s instructions. The samples were run in triplicate on an ABI Prism 7000 sequence detection system (Applied Biosystems). Expression of the target gene was normalized to the 18S endogenous control.

Statistics
All data are presented as means ± SD. Differences between groups (lean Zucker morning, lean Zucker evening, obese Zucker morning, obese Zucker evening, S-D morning, and S-D evening) were assessed in a univariate ANOVA. Bonferroni post hoc tests were done to correct for multiple analyses within each tissue or brain subregion. Subsequent P values for each morning/evening comparison are reported. For in situ measurements, a sample size of four in each group was estimated to have approximately 80% power to detect a 35% difference between morning and evening 11ß-HSD1 mRNA expression with a significance level of P = 0.05. Associations between local glucocorticoid metabolism and plasma corticosterone were analyzed by Spearman’s rank correlation coefficients. P < 0.05 was considered statistically significant.


   Results
Top
 Abstract
 Introduction
 Materials and Methods
 Results
Discussion
 References
 
Diurnal rhythm of circulating corticosterone
S-D and lean Zucker rats had the expected diurnal rhythm with trough morning and peak evening plasma corticosterone levels. Obese Zucker rats had elevated trough plasma corticosterone levels compared with lean animals, without a difference in peak corticosterone levels (Fig. 1Go).


Figure 1
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  		FIG. 1. Plasma corticosterone levels at morning (AM) and evening (PM) in lean and obese Zucker and S-D rats. Rats were decapitated at 0800–0900 h (morning groups) and 2000–2100 h (evening groups), respectively. Trunk blood was collected, and plasma corticosterone was assayed. n = 4 for lean and obese Zucker and n = 9 for S-D rats for each time point. Data are expressed as mean ± SD.

 
11ß-HSD1 mRNA expression in hippocampus and cortex
11ß-HSD1 was clearly expressed in the hippocampal regions and layer 4/5 of the cortex (Fig. 2Go). In the dentate gyrus granular layer and CA1 subregion of the hippocampus, lean Zucker rats had a significantly increased morning vs. evening 11ß-HSD1 mRNA expression (Fig. 3Go). This diurnal variation was lost in obese Zucker rats. Hippocampal 11ß-HSD1 mRNA expression in S-D rats (run in a separate in situ hybridization) also showed a diurnal rhythmicity similar to lean Zucker rats (Fig. 4Go). No significant diurnal variation or strain difference in 11ß-HSD1 expression was found in the cerebral cortex.


Figure 2
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  		FIG. 2. 11ß-HSD1 mRNA expression of a S-D rat brain section. Notice the high expression in the hippocampal subregions dentate gyrus (DG) and CA1 and layer 4/5 of the cortex. DG molecular and granular layer (after dipping in NTB-2 emulsion and counterstaining with pyronin Y; see Materials and Methods) is shown in higher magnification in the lower left corner. Scale bar, 50 µm.

 

Figure 3
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  		FIG. 3. 11ß-HSD1 mRNA quantification in hippocampal subregions by in situ hybridization. Shown are counts per neuron for morning (AM) and evening (PM) of lean and obese Zucker rats, respectively, in dentate gyrus (DG) granular and molecular layer and CA1. Data are expressed as mean ± SD of four animals in each group.

 

Figure 4
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  		FIG. 4. 11ß-HSD1 mRNA quantification in hippocampal subregions by in situ hybridization. Shown are counts per neuron for morning (AM) and evening (PM) of S-D rats in dentate gyrus (DG) granular and molecular layer and CA1. Data are expressed as mean ± SD of nine animals in each group.

 
11ß-HSD activity and gene expression in peripheral tissues
Data for 11ß-HSD activity in peripheral tissues are shown in Table 1Go. Obese Zucker rats had a significantly decreased liver 11ß-HSD activity in the morning compared with lean Zucker rats. No diurnal variation or strain difference in liver 11ß-HSD activity was seen in lean Zucker and S-D rats. Muscle 11ß-HSD morning and evening activity was similar in lean and obese Zucker rats as well as in S-D rats without strain differences. No diurnal variation was found in adipose tissue 11ß-HSD activity. Obese Zucker rats had significantly lower morning 11ß-HSD activity in the epididymal fat depot vs. lean animals. Moreover, no fat-depot-specific differences in 11ß-HSD activity were found in the two Zucker groups. In comparison, in S-D rats, retroperitoneal fat displayed highest, epididymal intermediate, and sc fat lowest 11ß-HSD activity.


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  		TABLE 1. 11ß-HSD activity (percent conversion) in peripheral tissues

 
Real-time RT-PCR measurements of 11ß-HSD1 mRNA expression in peripheral tissues were performed in the S-D cohort. Liver as well as retroperitoneal, epididymal, and sc adipose tissue depots did not reveal any circadian rhythms (data not shown). Muscle 11ß-HSD1 gene expression was higher in the evening (~150% compared with morning, P < 0.01).

Correlation analyses
The relationship between plasma corticosterone levels and local glucocorticoid metabolism, reflected as 11ß-HSD activity or gene expression, is presented in Table 2Go. There was a negative correlation between circulating corticosterone levels and hippocampal 11ß-HSD1 expression in S-D and lean Zucker rats that was absent in obese Zucker rats. There were no associations between 11ß-HSD activity in peripheral tissues and plasma corticosterone in S-D and lean rats. Among obese Zucker rats, only 11ß-HSD activity in muscle correlated positively with corticosterone levels (r = 0.71; P < 0.05).


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  		TABLE 2. Relationship between plasma corticosterone levels and local glucocorticoid metabolism (11ß-HSD activity or gene expression)

 

   Discussion
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
References
 
The novel finding in this study is a diurnal variability in hippocampal 11ß-HSD1 mRNA expression with peak mRNA expression during the morning period in lean animals, which is lost in the obese Zucker rat. This is linked to the increased basal plasma corticosterone levels in obese Zucker rats with unaltered peak concentrations, which is in agreement with earlier studies (19, 20, 21). The hippocampus, which is central for the HPA axis regulation in rodents, has a dense expression of MR, GR, and 11ß-HSD1 (14, 36, 37). Binding of GC to the low-affinity, high-capacity MR is important to maintain basal plasma GC levels. In contrast, activation of the high-affinity, low-capacity GR dampens high levels of GC (e.g. during stress and the diurnal peak) (14). A decrease in central expression of 11ß-HSD1 will reduce the local concentration of active GC, thus preventing the negative feedback and elevate circulating levels of corticosteroids as demonstrated in 11ß-HSD1 knockout mice (38). Importantly, the decrease in 11ß-HSD1 expression in obese Zucker rats was restricted to the morning period, which at least partly could explain why only basal but not peak levels of plasma corticosterone are altered.

As previously reported (7, 39), obese animals had a decreased enzyme activity in liver compared with lean animals. This may possibly be related to changes in the pattern of food intake because obese Zucker rats have quantitatively larger food intake than lean rats, especially during the dark phase (40). In addition to a decreased hepatic enzyme activity, the loss of diurnal rhythm in hippocampal 11ß-HSD1 mRNA expression in fatty rats only strengthens the hypothesis of a tissue-specific 11ß-HSD1 enzyme dysregulation in obesity. In accordance with enzyme activity data, there was no circadian rhythmicity in 11ß-HSD1 mRNA expression in the different fat depots as well as in liver of S-D rats. In contrast, gene expression in muscle was higher in the evening. The mechanism behind this finding could be related to physical activity because this is shown to induce a tissue-specific change in 11ß-HSD1 enzyme activity (41).

The enzyme activity did not differ comparing the different adipose depots within the Zucker phenotypes. In contrast, S-D rats had higher 11ß-HSD activity in retroperitoneal and epididymal fat vs. the sc fat depot. It has been reported that human omental stromal cells in culture have a higher enzyme activity than sc stromal cells (42), although the conformity of those cells with primary adipocytes remains to be established.

Putative regulators of 11ß-HSD1 include glucocorticoids, sex steroids, growth factors, insulin, thyroid hormones, and cytokines (43). Of interest for the present study, factors shown to have a positive influence on the 11ß-HSD1 transcription in vitro include TNF-{alpha}, IL-1ß, and leptin, whereas ACTH and CRH have a down-regulatory effect (43, 44, 45). Serum and/or central levels of several of these factors have been reported altered in the obese Zucker rat. Hypothalamic IL-1ß, but not TNF-{alpha}, mRNA expression is approximately 30% less in obese compared with lean rats (46), and the immunoreactivity of ACTH is increased in the anterior lobe of the pituitary in the fatty rat (47), whereas central expression of CRH has been reported to be increased (48) or not to differ (49, 50) compared with lean animals. The obese Zucker rat has a defective leptin receptor with increased circulating leptin levels. The attenuated enzyme expression of 11ß-HSD1 may consequently also result from a lack of leptin stimulation of the central 11ß-HSD1 expression. Importantly, these hormones and inflammatory mediators exhibit a diurnal variability in the normal state, but to our knowledge it is not yet studied whether the circadian rhythm of these factors is changed in animal models of obesity. Insulin has also been shown to stimulate 11ß-HSD1 mRNA expression in adipose tissue from insulin-resistant subjects (51). In rats fed on a high-energy diet, hippocampal insulin binding is decreased (52). It is possible to speculate that this is also applicable in obese Zucker rats and another factor contributing to a decreased expression of 11ß-HSD1.

Circulating levels of active GCs have been suggested to regulate the hippocampal 11ß-HSD1 expression and activity in a temporal manner. Chronic stress during 1 month decreased 11ß-HSD1 activity in tree shrews (53), whereas administration of a supraphysiological dose of dexamethasone to adrenalectomized rats for 10 d increased 11ß-HSD1 expression (54). Recently, Pelletier and colleagues (55) reported that hippocampal 11ß-HSD1 mRNA levels in mice increase after 7 d of adrenalectomy. The changes were reversible after corticosterone substitution for 24 h. In line with these findings, we found that there was a negative association between hippocampal 11ß-HSD1 mRNA expression and peak plasma corticosterone levels in S-D and lean Zucker rats.

These results may have clear implications for obesity in humans. 11ß-HSD1 is thus expressed in the human brain, including the hippocampus (56). Notably, overweight males have decreased concentrations of IL-6, another potential positive regulator of 11ß-HSD1 expression, in the cerebrospinal fluid despite increased serum levels (57). The present data suggest that regulation of 11ß-HSD activity in the central nervous system may influence the circadian rhythm of serum cortisol in human obesity. In peripheral tissues, the effect of GCs is mediated by activation of GR. A tonic activation of GR occurs when serum cortisol levels exceed approximately 83 nmol/liter, and thus, a relatively subtle increase in basal cortisol levels (as in obese humans) may contribute to the metabolic changes seen in the metabolic syndrome (1).

Some points need to be considered when interpreting our results. First, hippocampal 11ß-HSD1 enzyme activity was not estimated, which should be taken into consideration. The results from this kind of analysis is difficult to interpret because all subregions of the hippocampus would be homogenized together during the preparation, and important effects found comparing different subregions might easily be lost. It has, however, repeatedly been shown that 11ß-HSD1 gene expression corresponds to the enzyme activity in brain (36, 54) as well as in adipose tissue (6). In addition, to avoid the stressful situation that is created when removing the rats one by one from the cages before decapitation, the rats were housed singly for 5 d. Separating the animals might induce a temporary change of the HPA axis activity and modify the basal corticosterone secretion. Basal plasma corticosterone levels in both S-D and lean Zucker rats were, however, low and in accordance with what was earlier reported (7, 19, 20, 21, 58), reflecting an unstressed status. Obese Zucker rats had elevated basal plasma corticosterone levels, in agreement with previous reports (19, 20, 21). We therefore conclude that this was not likely a confounder for our results. Finally, because hippocampal GR and MR expression show a circadian pattern (30), it would be interesting to study corticosteroid receptors in animal models of obesity.

In summary, we have shown that obese Zucker rats lack a circadian rhythm of 11ß-HSD1 gene expression in the hippocampus, which may contribute to increased activity of the HPA axis and altered diurnal variation of circulating corticosterone levels in this rodent model of obesity.


   Acknowledgments
 
The excellent technical assistance of Jim Sandström, Ann-Kristin Norrman, Astrid Höglund, Ingrid Renholm, Malin Alvehus, and Therese Andersson are acknowledged.


   Footnotes
 
First Published Online March 1, 2007

Abbreviations: GC, Glucocorticoid; GR, glucocorticoid receptor; HPA, hypothalamic-pituitary-adrenal; HSD, hydroxysteroid dehydrogenase; MR, mineralocorticoid receptor; S-D, Sprague Dawley; SSC, standard saline citrate.

This work was supported by the Medical Faculty of Umeå University, the Swedish Society for Medical Research, the Swedish Society of Medicine, the Bergwalls Foundation, and the Lars Hiertas Minne and the Sigurd and Elsa Golje Foundations.

Disclosure Statement: The authors have nothing to declare.

Received July 5, 2006.

Accepted for publication February 20, 2007.


   References
Top
 Abstract
 Introduction
 Materials and Methods
 Results
 Discussion
 References
 

  1. Dallman MF, Akana SF, Bhatnagar S, Bell ME, Strack AM 2000 Bottomed out: metabolic significance of the circadian trough in glucocorticoid concentrations. Int J Obes Relat Metab Disord 24(Suppl 2):S40–S46
  2. Seckl JR, Walker BR 2004 11ß-Hydroxysteroid dehydrogenase type 1 as a modulator of glucocorticoid action: from metabolism to memory. Trends Endocrinol Metab 15:418–424[CrossRef][Medline]
  3. Rask E, Olsson T, Söderberg S, Andrew R, Livingstone DE, Johnson O, Walker BR 2001 Tissue-specific dysregulation of cortisol metabolism in human obesity. J Clin Endocrinol Metab 86:1418–1421[Abstract/Free Full Text]
  4. Rask E, Walker BR, Söderberg S, Livingstone DE, Eliasson M, Johnson O, Andrew R, Olsson T 2002 Tissue-specific changes in peripheral cortisol metabolism in obese women: increased adipose 11ß-hydroxysteroid dehydrogenase type 1 activity. J Clin Endocrinol Metab 87:3330–3336[Abstract/Free Full Text]
  5. Lindsay RS, Wake DJ, Nair S, Bunt J, Livingstone DE, Permana PA, Tataranni PA, Walker BR 2003 Subcutaneous adipose 11ß-hydroxysteroid dehydrogenase type 1 activity and messenger ribonucleic acid levels are associated with adiposity and insulinemia in Pima Indians and Caucasians. J Clin Endocrinol Metab 88:2738–2744[Abstract/Free Full Text]
  6. Wake DJ, Rask E, Livingstone DE, Soderberg S, Olsson T, Walker BR 2003 Local and systemic impact of transcriptional up-regulation of 11ß-hydroxysteroid dehydrogenase type 1 in adipose tissue in human obesity. J Clin Endocrinol Metab 88:3983–3988[Abstract/Free Full Text]
  7. Livingstone DE, Jones GC, Smith K, Jamieson PM, Andrew R, Kenyon CJ, Walker BR 2000 Understanding the role of glucocorticoids in obesity: tissue-specific alterations of corticosterone metabolism in obese Zucker rats. Endocrinology 141:560–563[Abstract/Free Full Text]
  8. Gomez-Sanchez EP, Ganjam V, Chen YJ, Liu Y, Zhou MY, Toroslu C, Romero DG, Hughson MD, de Rodriguez A, Gomez-Sanchez CE 2003 Regulation of 11ß-hydroxysteroid dehydrogenase enzymes in the rat kidney by estradiol. Am J Physiol Endocrinol Metab 285:E272–E279
  9. Masuzaki H, Paterson J, Shinyama H, Morton NM, Mullins JJ, Seckl JR, Flier JS 2001 A transgenic model of visceral obesity and the metabolic syndrome. Science 294:2166–2170[Abstract/Free Full Text]
  10. Masuzaki H, Yamamoto H, Kenyon CJ, Elmquist JK, Morton NM, Paterson JM, Shinyama H, Sharp MG, Fleming S, Mullins JJ, Seckl JR, Flier JS 2003 Transgenic amplification of glucocorticoid action in adipose tissue causes high blood pressure in mice. J Clin Invest 112:83–90[CrossRef][Medline]
  11. Kotelevtsev Y, Holmes MC, Burchell A, Houston PM, Schmoll D, Jamieson P, Best R, Brown R, Edwards CR, Seckl JR, Mullins JJ 1997 11ß-Hydroxysteroid dehydrogenase type 1 knockout mice show attenuated glucocorticoid-inducible responses and resist hyperglycemia on obesity or stress. Proc Natl Acad Sci USA 94:14924–14929[Abstract/Free Full Text]
  12. Morton NM, Holmes MC, Fievet C, Staels B, Tailleux A, Mullins JJ, Seckl JR 2001 Improved lipid and lipoprotein profile, hepatic insulin sensitivity, and glucose tolerance in 11ß-hydroxysteroid dehydrogenase type 1 null mice. J Biol Chem 276:41293–41300[Abstract/Free Full Text]
  13. Morton NM, Paterson JM, Masuzaki H, Holmes MC, Staels B, Fievet C, Walker BR, Flier JS, Mullins JJ, Seckl JR 2004 Novel adipose tissue-mediated resistance to diet-induced visceral obesity in 11ß-hydroxysteroid dehydrogenase type 1-deficient mice. Diabetes 53:931–938[Abstract/Free Full Text]
  14. De Kloet ER, Vreugdenhil E, Oitzl MS, Joels M 1998 Brain corticosteroid receptor balance in health and disease. Endocr Rev 19:269–301[Abstract/Free Full Text]
  15. Ljung T, Andersson B, Bengtsson BA, Bjorntorp P, Marin P 1996 Inhibition of cortisol secretion by dexamethasone in relation to body fat distribution: a dose-response study. Obes Res 4:277–282[Medline]
  16. Rosmond R, Dallman MF, Bjorntorp P 1998 Stress-related cortisol secretion in men: relationships with abdominal obesity and endocrine, metabolic and hemodynamic abnormalities. J Clin Endocrinol Metab 83:1853–1859[Abstract/Free Full Text]
  17. Walker BR, Soderberg S, Lindahl B, Olsson T 2000 Independent effects of obesity and cortisol in predicting cardiovascular risk factors in men and women. J Intern Med 247:198–204[CrossRef][Medline]
  18. Ljung T, Holm G, Friberg P, Andersson B, Bengtsson BA, Svensson J, Dallman M, McEwen B, Bjorntorp P 2000 The activity of the hypothalamic-pituitary-adrenal axis and the sympathetic nervous system in relation to waist/hip circumference ratio in men. Obes Res 8:487–495[Medline]
  19. Martin RJ, Wangsness PJ, Gahagan JH 1978 Diurnal changes in serum metabolites and hormones in lean and obese Zucker rats. Horm Metab Res 10:187–192[Medline]
  20. Fletcher JM, Haggarty P, Wahle KW, Reeds PJ 1986 Hormonal studies of young lean and obese Zucker rats. Horm Metab Res 18:290–295[Medline]
  21. Walker CD, Scribner KA, Stern JS, Dallman MF 1992 Obese Zucker (fa/fa) rats exhibit normal target sensitivity to corticosterone and increased drive to adrenocorticotropin during the diurnal trough. Endocrinology 131:2629–2637[Abstract]
  22. Cunningham JJ, Calles-Escandon J, Garrido F, Carr DB, Bode HH 1986 Hypercorticosteronuria and diminished pituitary responsiveness to corticotropin-releasing factor in obese Zucker rats. Endocrinology 118:98–101[Abstract]
  23. White BD, Corll CB, Porter JR 1989 The metabolic clearance rate of corticosterone in lean and obese male Zucker rats. Metabolism 38:530–536[CrossRef][Medline]
  24. Fraser R, Ingram MC, Anderson NH, Morrison C, Davies E, Connell JM 1999 Cortisol effects on body mass, blood pressure, and cholesterol in the general population. Hypertension 33:1364–1368[Abstract/Free Full Text]
  25. Stewart PM, Boulton A, Kumar S, Clark PM, Shackleton CH 1999 Cortisol metabolism in human obesity: impaired cortisone->cortisol conversion in subjects with central adiposity. J Clin Endocrinol Metab 84:1022–1027[Abstract/Free Full Text]
  26. Lottenberg SA, Giannella-Neto D, Derendorf H, Rocha M, Bosco A, Carvalho SV, Moretti AE, Lerario AC, Wajchenberg BL 1998 Effect of fat distribution on the pharmacokinetics of cortisol in obesity. Int J Clin Pharmacol Ther 36:501–505[Medline]
  27. Purnell JQ, Brandon DD, Isabelle LM, Loriaux DL, Samuels MH 2004 Association of 24-hour cortisol production rates, cortisol-binding globulin, and plasma-free cortisol levels with body composition, leptin levels, and aging in adult men and women. J Clin Endocrinol Metab 89:281–287[Abstract/Free Full Text]
  28. Strain GW, Zumoff B, Kream J, Strain JJ, Levin J, Fukushima D 1982 Sex difference in the influence of obesity on the 24 hr mean plasma concentration of cortisol. Metabolism 31:209–212[CrossRef][Medline]
  29. Jessop DS, Dallman MF, Fleming D, Lightman SL 2001 Resistance to glucocorticoid feedback in obesity. J Clin Endocrinol Metab 86:4109–4114[Abstract/Free Full Text]
  30. Herman SP, Watson SJ, Chao HM, Coirini H, McEwan BS 1993 Diurnal regulation of glucocorticoid receptor and mineralocorticoid receptor mSNAs in rat hippocampus. Mol Cell Neurosci 4:181–190[CrossRef]
  31. Gratsias Y, Moutsatsou P, Chrysanthopoulou G, Tsagarakis S, Thalassinos N, Sekeris CE 2000 Diurnal changes in glucocorticoid sensitivity in human peripheral blood samples. Steroids 65:851–856[CrossRef][Medline]
  32. Mattsson C, Lai M, Noble J, McKinney E, Yau JL, Seckl JR, Walker BR 2003 Obese Zucker rats have reduced mineralocorticoid receptor and 11ß-hydroxysteroid dehydrogenase type 1 expression in hippocampus-implications for dysregulation of the hypothalamic-pituitary-adrenal axis in obesity. Endocrinology 144:2997–3003[Abstract/Free Full Text]
  33. Paxinos G, Watson C 1998 The rat brain in stereotaxic coordinates. Orlando, FL: Academic Press
  34. Rönnbäck A, Dahlqvist P, Bergström SA, Olsson T 2005 Diurnal effects of enriched environment on immediate early gene expression in the rat brain. Brain Res 1046:137–144[CrossRef][Medline]
  35. Lakshmi V, Monder C 1988 Purification and characterization of the corticosteroid 11ß-dehydrogenase component of the rat liver 11ß-hydroxysteroid dehydrogenase complex. Endocrinology 123:2390–2398[Abstract]
  36. Moisan MP, Seckl JR, Edwards CR 1990 11ß-Hydroxysteroid dehydrogenase bioactivity and messenger RNA expression in rat forebrain: localization in hypothalamus, hippocampus, and cortex. Endocrinology 127:1450–1455[Abstract]
  37. Seckl JR, Olsson T 1995 Glucocorticoid hypersecretion and the age-impaired hippocampus: cause or effect? J Endocrinol 145:201–211[Medline]
  38. Harris HJ, Kotelevtsev Y, Mullins JJ, Seckl JR, Holmes MC 2001 Intracellular regeneration of glucocorticoids by 11ß-hydroxysteroid dehydrogenase (11ß-HSD)-1 plays a key role in regulation of the hypothalamic-pituitary-adrenal axis: analysis of 11ß-HSD-1-deficient mice. Endocrinology 142:114–120[Abstract/Free Full Text]
  39. Livingstone DE, Kenyon CJ, Walker BR 2000 Mechanisms of dysregulation of 11ß-hydroxysteroid dehydrogenase type 1 in obese Zucker rats. J Endocrinol 167:533–539[Abstract]
  40. Alingh Prins A, de Jong-Nagelsmit A, Keijser J, Strubbe JH 1986 Daily rhythms of feeding in the genetically obese and lean Zucker rats. Physiol Behav 38:423–426[CrossRef][Medline]
  41. Coutinho AE, Campbell JE, Fediuc S, Riddell MC 2006 Effect of voluntary exercise on peripheral tissue glucocorticoid receptor content and the expression and activity of 11ß-HSD1 in the Syrian hamster. J Appl Physiol 100:1483–1488[Abstract/Free Full Text]
  42. Bujalska IJ, Kumar S, Stewart PM 1997 Does central obesity reflect "Cushing’s disease of the omentum"? Lancet 349:1210–1213[CrossRef][Medline]
  43. Tomlinson JW, Stewart PM 2001 Cortisol metabolism and the role of 11ß-hydroxysteroid dehydrogenase. Best Pract Res Clin Endocrinol Metab 15:61–78[CrossRef][Medline]
  44. Tomlinson JW, Moore J, Cooper MS, Bujalska I, Shahmanesh M, Burt C, Strain A, Hewison M, Stewart PM 2001 Regulation of expression of 11ß-hydroxysteroid dehydrogenase type 1 in adipose tissue: tissue-specific induction by cytokines. Endocrinology 142:1982–1989[Abstract/Free Full Text]
  45. Friedberg M, Zoumakis E, Hiroi N, Bader T, Chrousos GP, Hochberg Z 2003 Modulation of 11ß-hydroxysteroid dehydrogenase type 1 in mature human subcutaneous adipocytes by hypothalamic messengers. J Clin Endocrinol Metab 88:385–393[Abstract/Free Full Text]
  46. Wisse BE, Ogimoto K, Morton GJ, Wilkinson CW, Frayo RS, Cummings DE, Schwartz MW 2004 Physiological regulation of hypothalamic IL-1ß gene expression by leptin and glucocorticoids: implications for energy homeostasis. Am J Physiol Endocrinol Metab 287:E1107–E1113
  47. Bestetti GE, Abramo F, Guillaume-Gentil C, Rohner-Jeanrenaud F, Jeanrenaud B, Rossi GL 1990 Changes in the hypothalamo-pituitary-adrenal axis of genetically obese fa/fa rats: a structural, immunocytochemical, and morphometrical study. Endocrinology 126:1880–1887[Abstract]
  48. Timofeeva E, Richard D 1997 Functional activation of CRH neurons and expression of the genes encoding CRH and its receptors in food-deprived lean (Fa/?) and obese (fa/fa) Zucker rats. Neuroendocrinology 66:327–340[Medline]
  49. Pesonen U, Huupponen R, Rouru J, Koulu M 1992 Hypothalamic neuropeptide expression after food restriction in Zucker rats: evidence of persistent neuropeptide Y gene activation. Brain Res Mol Brain Res 16:255–260[Medline]
  50. Plotsky PM, Thrivikraman KV, Watts AG, Hauger RL 1992 Hypothalamic-pituitary-adrenal axis function in the Zucker obese rat. Endocrinology 130:1931–1941[Abstract]
  51. Westerbacka J, Corner A, Kannisto K, Kolak M, Makkonen J, Korsheninnikova E, Nyman T, Hamsten A, Fisher RM, Yki-Jarvinen H 2006 Acute in vivo effects of insulin on gene expression in adipose tissue in insulin-resistant and insulin-sensitive subjects. Diabetologia 49:132–140[CrossRef][Medline]
  52. Irani BG, Dunn-Meynell AA, Levin BE 2007 Altered hypothalamic leptin, insulin, and melanocortin binding associated with moderate-fat diet and predisposition to obesity. Endocrinology 148:310–316[Abstract/Free Full Text]
  53. Jamieson PM, Fuchs E, Flugge G, Seckl JR 1997 Attenuation of hippocampal 11ß-hydroxysteroid dehydrogenase type 1 by chronic psychosocial stress in the tree shrew. Stress 2:123–132[Medline]
  54. Low SC, Moisan MP, Noble JM, Edwards CR, Seckl JR 1994 Glucocorticoids regulate hippocampal 11ß-hydroxysteroid dehydrogenase activity and gene expression in vivo in the rat. J Neuroendocrinol 6:285–290[CrossRef][Medline]
  55. Pelletier G, Luu-The V, Li S, Bujold G, Labrie F 2007 Localization and glucocorticoid regulation of 11ß-hydroxysteroid dehydrogenase type 1 mRNA in the male mouse forebrain. Neuroscience 145:110–115[CrossRef][Medline]
  56. Sandeep TC, Yau JL, MacLullich AM, Noble J, Deary IJ, Walker BR, Seckl JR 2004 11ß-Hydroxysteroid dehydrogenase inhibition improves cognitive function in healthy elderly men and type 2 diabetics. Proc Natl Acad Sci USA 101:6734–6739[Abstract/Free Full Text]
  57. Stenlof K, Wernstedt I, Fjallman T, Wallenius V, Wallenius K, Jansson JO 2003 Interleukin-6 levels in the central nervous system are negatively correlated with fat mass in overweight/obese subjects. J Clin Endocrinol Metab 88:4379–4383[Abstract/Free Full Text]
  58. Sapolsky RM 1992 Do glucocorticoid concentrations rise with age in the rat? Neurobiol Aging 13:171–174[CrossRef][Medline]




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