In mammals, leptin (Lep) binding proteins (LepBPs) derived from Lep receptor (LepR) gene or protein bind most of the circulating Lep, but to date, information on LepBPs in nonmammalian vertebrate classes is lacking. This study details the characterization of multiple LepBPs in rainbow trout (Oncorhynchus mykiss), an early poikilothermic vertebrate, and presents the complete coding sequences for 3 of them. Size-exclusion chromatography and cross-linking assay identified plasma proteins bound to Lep ranging from 70 to 100 kDa. LepBPs were isolated from plasma by affinity chromatography, and their binding specificity was assessed by a competitive binding assay. A RIA for LepBPs indicates that plasma LepBP levels decline after fasting for 3 weeks. Immunoblotting of LepBPs using antibodies against different LepR epitopes shows that the LepBPs are indeed LepR isoforms. The alternatively spliced LepR transcripts (LepRS1-3) that include only the extracellular segment transcribe the 90-kDa LepBP1, the 80-kDa LepBP2, and the 70-kDa LepBP3, respectively. LepRS1 generally has lower expression than the long-form LepR in most tissues. LepRS2 is primarily expressed in adipose tissue, whereas LepRS3 is expressed abundantly in brain and spleen, and moderately in liver and gills. The mRNA levels of hypothalamic LepRS1 and hepatic LepRS3 increase after 2 weeks of fasting, but decrease after 3 weeks. This study demonstrates a mechanism in fish for the generation of LepBPs that differs from that seen in mammals and indicates that the physiologic action of Lep in these poikilothermic vertebrates can be modulated, both centrally and peripherally, by the differentiated, tissue-specific expression of multiple LepBPs.

The pancreatic hormone amylin acts in the central nervous system (CNS) to decrease food intake and body weight. We hypothesized that amylin action in the CNS promotes energy expenditure by increasing the activity of the sympathetic nervous system. In mice, ip administration of amylin significantly increased c-Fos immunoreactivity in hypothalamic and brainstem nuclei. In addition, mice treated with intracerebroventricular (icv) amylin (0.1 and 0.2 nmol) exhibited a dose-related decrease in food intake and body weight, measured 4 and 24 hours after treatment. The icv injection of amylin also increased body temperature in mice. Using direct multifiber sympathetic nerve recording, we found that icv amylin elicited a significant and dose-dependent increase in sympathetic nerve activity (SNA) subserving thermogenic brown adipose tissue (BAT). Of note, icv injection of amylin also evoked a significant and dose-related increase in lumbar and renal SNA. Importantly, icv pretreatment with the amylin receptor antagonist AC187 (20 nmol) abolished the BAT SNA response induced by icv amylin, indicating that the sympathetic effects of amylin are receptor-mediated. Conversely, icv amylin-induced BAT SNA response was enhanced in mice overexpressing the amylin receptor subunit, RAMP1, in the CNS. Our data demonstrate that CNS action of amylin regulates sympathetic nerve outflow to peripheral tissues involved in energy balance and cardiovascular function.

Chronic stress is a risk factor for several neuropsychiatric diseases such as depression and psychosis. In response to stress glucocorticoids (GCs) are secreted that bind to the glucocorticoid receptor (GR), a ligand-activated transcription factor that regulates the transcription of gene networks in the brain necessary for coping with stress, recovery and adaptation. Chronic stress particularly affects the dentate gyrus (DG) subregion of the hippocampus, causing several functional and morphological changes with consequences for learning and memory, that are likely adaptive, but at the same time make DG neurons more vulnerable to subsequent challenges.The aim of this study was to investigate the transcriptional response of DG neurons to a GC-challenge in male rats previously exposed to chronic restraint stress (CRS). An intriguing finding of the current study was that having a history of CRS had profound consequences for the subsequent response to acute GC-challenge, differentially affecting the expression of several hundreds of genes in the DG compared to challenged non-stressed control animals. This enduring effect of prior stress exposure suggests that epigenetic processes may be involved. In line with this, CRS indeed affected the expression of several genes involved in chromatin structure and epigenetic processes, including Asf1, Ash1l, Hist1h3f and Tp63. The data presented here indicate that CRS alters the transcriptional response to a subsequent GC-injection. We propose that this altered transcriptional potential forms part of the molecular mechanism underlying the enhanced vulnerability for stress-related disorders like depression caused by chronic stress.

The prevalence of insulin resistance and type 2 diabetes mellitus are rising dramatically and as a consequence there is an urgent need to understand the pathogenesis underpinning these conditions to develop new and more efficacious treatments.We have tested the hypothesis that glucocorticoid-mediated changes in insulin sensitivity may be associated with changes in lipid flux. Furthermore, pre-receptor modulation of glucocorticoid availability by 11β-hydroxysteroid dehydrogenase type 1 (11β-HSD1) may represent a critical regulatory step.Dexamethasone decreased lipogenesis in both murine C2C12 and human LHC-NM2 myotubes. Inactivating p-Ser-79/218 of acetyl-CoA carboxylase 1/2 (ACC1/2) and activating p-Thr-172 of AMP-activated protein kinase (AMPK) were both increased following dexamethasone treatment in C2C12 myotubes. By contrast, dexamethasone increased β-oxidation. Selective 11β-HSD1 inhibition blocked the 11-dehydrocorticosterone (11DHC)-mediated decrease in lipogenic, and increase in lipolytic gene expression. Lipogenic gene expression was decreased, whilst lipolytic and β-oxidative genes expression increased in corticosterone (CORT) and 11DHC treated wild-type mice, and CORT (but not 11DHC) treated 11β-HSD1(-/-) mice. Furthermore, CORT and 11DHC treated wild-type mice, and CORT (but not 11DHC) treated 11β-HSD1(-/-) mice had increased p-Ser-79/218 ACC1/2, p-Thr-172 AMPK and intramyocellular diacylglyderide content.In summary, we have shown that glucocorticoids have potent actions upon intramyocellular lipid homeostasis by decreasing lipid storage, increasing lipid mobilisation and utilisation and increasing diacylglyderide content. It is plausible that dysregulated intramyocellular lipid metabolism may underpin GC-induced insulin resistance of skeletal muscle.

Diabetes mellitus is characterized by insulin secretion from pancreatic β cells that is insufficient to maintain blood glucose homeostasis. Autoimmune destruction of β cells results in type 1 diabetes mellitus, whereas conditions that reduce insulin sensitivity and negatively affect β-cell activities result in type 2 diabetes mellitus. Without proper management, patients with diabetes mellitus develop serious complications that reduce their quality of life and life expectancy. Biomarkers for early detection of the disease and identification of individuals at risk of developing complications would greatly improve the care of these patients. Small non-coding RNAs called microRNAs (miRNAs) control gene expression and participate in many physiopathological processes. Hundreds of miRNAs are actively or passively released in the circulation and can be used to evaluate health status and disease progression. Both type 1 diabetes mellitus and type 2 diabetes mellitus are associated with distinct modifications in the profile of miRNAs in the blood, which are sometimes detectable several years before the disease manifests. Moreover, circulating levels of certain miRNAs seem to be predictive of long-term complications. Technical and scientific obstacles still exist that need to be overcome, but circulating miRNAs might soon become part of the diagnostic arsenal to identify individuals at risk of developing diabetes mellitus and its devastating complications.

The availability of synthetic recombinant human growth hormone (GH) in potentially unlimited quantities since the 1980s has improved understanding of the many nonstatural effects of GH on metabolism, body composition, physical and psychological function, as well as the consequences of GH deficiency in adult life. Adult GH deficiency is now recognized as a distinct if nonspecific syndrome with considerable adverse health consequences. GH replacement therapy in lower doses than those used in children can reverse many of these abnormalities and restore functional capacities toward or even to normal; if dosed appropriately, GH therapy has few adverse effects. Although some doubts remain about possible long-term risks of childhood GH therapy, most registries of adult GH replacement therapy, albeit limited in study size and duration, have not shown an increased incidence of cancers or of cardiovascular morbidity or mortality.

Emerging data indicate that growth hormone (GH) therapy could have a role in improving cognitive function. GH replacement therapy in experimental animals and human patients counteracts the dysfunction of many behaviours related to the central nervous system (CNS). Various behaviours, such as cognitive behaviours related to learning and memory, are known to be induced by GH; the hormone might interact with specific receptors located in areas of the CNS that are associated with the functional anatomy of these behaviours. GH is believed to affect excitatory circuits involved in synaptic plasticity, which alters cognitive capacity. GH also has a protective effect on the CNS, as indicated by its beneficial effects in patients with spinal cord injury. Data collected from animal models indicates that GH might also stimulate neurogenesis. This Review discusses the mechanisms underlying the interactions between GH and the CNS, and the data emerging from animal and human studies on the relationship between GH and cognitive function. In this article, particular emphasis is given to the role of GH as a treatment for patients with cognitive impairment resulting from deficiency of the hormone.

Intrauterine growth restriction (IUGR) is an important fetal developmental problem resulting from two broad causes: maternal under-nutrition and/or decreased fetal nutrient delivery to the fetus via placental insufficiency. IUGR is often accompanied by up-regulation of the hypothalamo-pituitary-adrenal axis (HPAA). Sheep studies show fetal HPAA autonomy in late gestation. We hypothesized that IUGR, resulting from poor fetal nutrient delivery, up-regulates the fetal baboon HPAA in late gestation driven by hypothalamo-pituitary glucocorticoid receptor (GR) insensitivity and decreased fetal leptin in peripheral plasma. Maternal baboons were fed as ad lib controls (CTR) or nutrient restricted to produce IUGR (fed 70% CTR diet) from 0.16 - 0.9 gestation. Peripheral ACTH, cortisol and leptin were measured by immunoassays. Corticotropin-releasing hormone (CRH), arginine vasopressin (AVP), glucocorticoid receptor (GR), leptin receptor (ObRb) and proopiomelanocortin (POMC) peptide expression were determined immunohistochemically (IHC). IUGR fetal peripheral cortisol and ACTH, but not leptin, were increased (p< 0.05). IUGR increased CRH peptide expression in the fetal hypothalamic paraventricular nucleus (PVN) and median eminence (p< 0.05), but not AVP. PVN ObRb peptide expression was decreased (p< 0.05) with IUGR, but not GR. ObRb and POMC were robustly expressed in anterior pituitary, but ∼ 1% of cells showed co-localization. We conclude that 1) CRH, but not AVP, is the major releasing hormone driving ACTH and cortisol secretion during primate IUGR, 2) fetal HPAA activation was aided by GR insensitivity and decreased ObRb expression in the PVN and 3) the AP is not a site for ObRb effects on the HPAA.

Islet cell growth and function are affected by ligands from the Epidermal Growth Factor family. We describe here the expression, regional distribution and effect on growth and secretion of insulin of a subset of these, the Neuregulin family. The expression of NRG1 alpha, NRG1 beta, NRG2 alpha, NRG2 beta, NRG3 and NRG4 in rat islets was determined using immunohistochemical and double immunofluorescent staining. We also report the expression of the four receptors and the remaining seven ligands using immunohistochemistry. The NRG1 alpha splice variant was expressed in beta cells and the NRG1 beta variant mainly in alpha cells. NRG3 was also predominantly present in alpha cells. Most of the members of the EGF family of ligands were also expressed, with Epigen being present at the highest levels. The rat islet derived cell line CRI-G1 was used to study the effect of addition of EGF, NRG1 beta, NRG3 and NRG4 on cell growth and insulin secretion. Synthetic refolded NRG3 strongly stimulated the growth of the CRI-G1 cells and NRG4 gave the greatest stimulation of insulin release. Different members of the Neuregulin family are therefore potentially potent stimuli for islet cell growth and insulin release and differ in expression in alpha and beta cells.

Estrogen has been reported to affect pain perception, although the underlying mechanisms remain unclear. In this investigation, pain behavior testing, patch clamp recording and immunohistochemistry were used on rats and transgenic mice to determine which estrogen receptors and the related signaling pathway are involved in the rapid modulation of estrogen on P2X3 receptor-mediated events. The results showed that 17β-estradiol rapidly inhibited pain induced by α,β-methylene ATP (α,β-me-ATP), a P2X1 and P2X3 receptor agonist in ovariectomized rats and normal rats in diestrus. The estrogen receptor-α (ERα) agonist 4,49,499-(4-propyl-[1H]-pyrazole-1,3,5-triyl) trisphenol (PPT) and GPR30 receptor agonist G-1 mimicked the estrogen effect, while the ERβ agonist diarylpropionitrile (DPN) had no effect. In cultured rat dorsal root ganglion (DRG) neurons, PPT and G-1 but not DPN significantly attenuated α,β-meATP-mediated currents, with the dose-response curve of these currents shifted to the right. The inhibitory effect of 17β-estradiol on P2X3 currents was blocked by G-15, a selective antagonist to the GPR30 estrogen receptor. 17β-estradiol lacked this effect in DRG neurons from ERα knockout mice while partly remained in those from ERβ knockout mice. The P2X3 and GPR30 receptors were co-expressed in the rat DRG neurons. Further, the ERK1/2 inhibitor U0126 reversed the inhibitory effect of 17β-estradiol on α,β-me-ATP-induced pain and of PPT or G-1 on P2X3 receptor-mediated currents. The cyclic AMP-protein kinase A (PKA) agonist forskolin, but not the protein kinase C agonist phorbol-12-myristate-13-acetate (PMA) mimicked the estrogen inhibitory effect on P2X3 receptor currents, which was blocked by another ERK1/2 inhibitor, PD98059. These results suggest that estrogen regulates P2X3-mediated peripheral pain by acting on ERα and GPR30 receptors expressed in primary afferent neurons, which probably involves the intracellular cAMP-PKA-ERK1/2 pathway.