Evidence for daily rhythms of the expression of proopiomelanocortin, interleukin-1-beta and interleukin-6 in adenopituitaries of male long-evans rats: effect of adjuvant arthritis.

OBJECTIVE: Several interleukins (ILs) including IL-1 beta and IL-6 are produced in the anterior pituitary (AP) where they probably participate in the local regulation of hormone production. Immune challenge brings about the dysregulation of immune-endocrine interaction and enhanced the expression of pituitary IL-1 beta and IL-6. Little is known about regulation of their production, and therefore the purpose of the present work was to describe the relationship between circulating corticosterone and the mRNA expression of proopiomelanocortin (POMC), IL-1 beta and IL-6 in the AP during a 24-hour cycle in normal rats and rats with acute adjuvant arthritis (AA). METHODS: Groups of intact male Long-Evans rats and rats 23 days after induction of AA kept on a 12-hour light/dark cycle (light on at 6:00 a.m.) were killed at 4-hour intervals starting at 2:00 p.m. Trunk blood was used for corticosterone determination by radioimmunoassay. Adenopituitaries were extracted for total RNA and the message of interest was quantitated by real-time PCR using specific primers and TaqMan probes. Parameters of rhythms were evaluated by cosinor analysis. RESULTS: In normal rats, serum corticosterone showed a circadian rhythm with the peak at 6:00 p.m. and the nadir in the morning hours (p < 0.001). POMC mRNA in AP also showed a circadian rhythm (p < 0.05) which was inversely related to corticosterone levels. IL-1beta and IL-6 expression in normal rats showed clear-cut daily rhythms (p < 0.001) with the nadirs in the dark period, in contrast to the corticosterone peak in plasma. In arthritic rats, rhythmic corticosterone secretion was suppressed with a plateau pattern of the rhythm. The mean POMC expression was higher than in controls, and the rhythm failed to be significant. IL-1 beta expression was suppressed by AA (p < 0.001) but the rhythm was still present (p < 0.05). The rhythmic pattern of IL-6 expression was similar to that of controls, but with higher mesor values (p < 0.05). CONCLUSION: These results suggest a regulatory relationship between circulating corticosterone and the expression of POMC, IL-1 beta and IL-6 in AP of normal rats. Arthritis induced a higher expression of POMC and IL-6 in the AP and a suppression of IL-1 beta mRNA during the 24-hour cycle which suggests the involvement of different regulatory mechanisms compared to normal conditions.

Primary human CD34+ hematopoietic stem and progenitor cells express functionally active receptors of neuromediators.

Recently, overlapping molecular phenotypes of hematopoietic and neuropoietic cells were described in mice. Here, we examined primary human CD34(+) hematopoietic stem and progenitor cells applying specialized cDNA arrays, real-time reverse-transcriptase-polymerase chain reaction (RT-PCR), and fluorescent-activated cell sorter (FACS) analysis focusing on genes involved in neurobiologic functions. We found expression of vesicle fusion and motility genes, ligand- and voltage-gated ion channels, receptor kinases and phosphatases, and, most interestingly, mRNA as well as protein expression of G protein-coupled receptors of neuromediators (corticotropin-releasing hormone 1 [CRH 1] and CRH 2 receptors, orexin/hypocretin 1 and 2 receptors, GABAB receptor, adenosine A(2)B receptor, opioid kappa 1 and mu 1 receptors, and 5-HT 1F receptor). As shown by 2-color immunofluorescence, the protein expression of these receptors was higher in the more immature CD38(dim) than in the CD38(bright) subset within the CD34(+) population, and completely absent in fully differentiated blood cells, suggesting that those receptors play a role in developmentally early CD34(+) stem and progenitor cells. The intracellular concentration of cyclic adenosine monophosphate (cAMP) in CD34(+) cells was diminished significantly upon stimulation of either CRH or orexin receptors, indicating that those are functionally active and coupled to inhibitory G proteins in human hematopoietic cells. In conclusion, these findings suggest a molecular interrelation of neuronal and hematopoietic signaling mechanisms in humans.

Corticotropin-releasing hormone system in human adipose tissue.

Mounting evidence exists for a role of the CRH system in energy balance, including a direct influence on human adipocytes, the regulation of adipose 11 beta-hydroxysteroid dehydrogenase type 1 activity, and cortisol formation. We characterized the expression of CRH receptors 1 and 2 and CRH-like peptides stresscopin and urocortin in human adipose tissue in comparison with other peripheral tissues, adrenal, and heart. The effect of CRH on CRH receptor and CRH-like peptide expression was analyzed in isolated human adipocytes using quantitative TaqMan PCR. CRH receptors were detectable in fat tissue at mRNA and protein levels. CRH-R2 expression in fat was comparable with its expression in the heart, the organ with the highest CRH-R2 expression known. CRH-R1:CRH-R2 ratio varied according to fat-depot type; whereas CRH-R1 expression was higher in sc fat than in visceral fat, the opposite was true for CRH-R2. Adipose tissue also expressed urocortin and stresscopin, the predominant ligands of peripheral CRH-R2. CRH down-regulated CRH-R1 and CRH-R2 mRNA expression in isolated adipocytes. These data, together with the recently published observation that CRH regulates adipocyte metabolism by down-regulating 11 beta-hydroxysteroid dehydrogenase, indicate that a CRH system exists within human adipose tissue. This system could be implicated in energy homeostasis and in mediating the anorexic effects of CRH at adipose level.

Neuroendocrine or behavioral effects of acute or chronic emotional stress in Wistar Kyoto (WKY) and spontaneously hypertensive (SHR) rats.

OBJECTIVE: Spontaneously hypertensive rats (SHR) selected from Wistar Kyoto (WKY) strain represent an animal model of human essential hypertension. This strain of rats is known by excessive neuroendocrine and cardiovascular responses under stress. The aim of the present study was: 1. To compare the reactivity of hypothalamic-pituitary-adrenocortical axis (HPA) to acute mild stress of handling between SHR and WKY rats, 2. to compare the behavioral activity of both strains under basal conditions and during chronic unpredictable emotional stress. METHODS: Seven to eight weeks old male SHR and WKY rats bred in the Physiological Institute, Academy of Sciences of the Czech Republic (Prague) were used. Acute stress was induced by 2-minute handling of the animals in their cage. Blood plasma was analyzed for ACTH and corticosterone (CORT) by specific radioimmunoassay. Chronic unpredictable stress lasted 20 days and consisted of random exposures to following interventions: Light on or off for 24 h, overcrowding i.e. pooling the rats from two cages into one (size 24 x 39 x 23 cm) for 24 h, isolation by placing a single rat into one cage for 24 h, new hierarchy by mixing 4 rats from two different cages for 24 h, limited access to food or water for 1 hour in one day between 3 and 6 p.m., inescapable foot shock (20 shocks, duration 5 s, intensity 10 mA, intershock interval 30 s), tilting the cages for 24 h. The sequence of individual stress exposures was the same in all rats. On day 6, 10 and 20, behavioral activity was measured using the elevated plus-maze in non-stressed control and stressed rats. The results were evaluated by non parametrical Kruskal-Wallis test followed by Mann-Whitney U-test. RESULTS: The two-minute handling resulted in a significantly higher activation of HPA in the SHR than in the WKY rats (plasma ACTH: 350 +/- 65 pg/ml for SHR vs. 97 +/- 17 pg/ml for WKY p<0.01; plasma corticosterone: 2.8 +/- 1.4 mg/100 ml for SHR vs. 0.7 +/- 0.06 mg/100 ml for WKY p<0.05). In WKY rats no activation of HPA was observed. Elevated plus-maze anxiety test showed inverse behavioral pattern between SHR and WKY rats. In the first test of anxiety the number of open arm entries (OAE) as well as total mobility expressed as total arm entries of the SHR was lower than of the WKY rats (p<0.01) without any difference between stressed and non-stressed animals in either strain. It was gradually increasing in stressed and non-stressed SHR in subsequent sessions markedly exceeding the activity of WKY rats (p<0.01). Stressed WKY rats showed less OAE and total mobility than their controls (p<0.01). CONCLUSIONS: Our results show enhanced neuroendocrine response to acute handling and enhanced anxiety in acute novelty stress in SHR comparing to WKY rats which suggests a common mechanisms for neuroendocrine and behavioral changes. These results further underline the lack of anxiety related behavior of SHR under chronic emotional stress.

Daily profiles of plasma prolactin (PRL), growth hormone (GH), insulin-like growth factor-1 (IGF-1), luteinizing hormone (LH), testosterone, and melatonin, and of pituitary PRL mRNA and GH mRNA in male Long Evans rats in acute phase of adjuvant arthritis.

We studied the effects of adjuvant arthritis (AA) on the endocrine circadian rhythms of plasma prolactin (PRL), growth hormone (GH), insulin-like growth factor-1 (IGF-1), luteinizing hormone (LH), testosterone, and melatonin and of pituitary PRL and GH mRNA in male Long Evans rats. Groups of control and AA rats (studied 23 days after AA induction) that were housed under a 12/12 h light/dark cycle (light on at 06:00 h) were killed at 4 h intervals starting at 14:00 h. Cosinor analysis revealed a significant 12 h rhythm in PRL and PRL mRNA (p < 0.001) in controls with peaks at 14:00 h and 02:00 h, respectively. The peak at 02:00 h was abolished in the AA group resulting in a significant 24 h rhythm in parallel with that of PRL (p < 0.05) and PRL mRNA (p < 0.0001). Growth hormone showed no rhythm, but a significant rhythm of GH mRNA was present in both groups (p < 0.0001). Insulin-like growth factor-1 showed a 24 h rhythm in control but not in AA rats. The mean values of GH, GH mRNA, and IGF-1 were significantly reduced in AA. Luteinizing hormone displayed a significant 24 h rhythm (p < 0.01) peaking in the dark period in the control but not AA group. Testosterone showed in phase temporal changes of LH levels with AA abolishing the 02:00 h peak. Melatonin exhibited a significant 24 h rhythm in control (p < 0.001) and AA (p < 0.01) rats with maximum levels during the dark phase; the mesor value was higher in the AA males. These results demonstrate that AA interferes with the rhythms of all the studied hormones except the non-24 h (arrhythmic) GH secretion pattern and the rhythm in melatonin. The persistence of a distinct melatonin rhythm in AA suggests the observed disturbances of hormonal rhythms in this condition do not occur at the level of the pineal gland.