The effect of the ghrelin-receptor agonist capromorelin on glucose metabolism in healthy cats.

Somatostatin secretion from islet delta cells is important in maintaining low glycemic variability (GV) by providing negative feedback to beta cells and inhibiting insulin secretion. Capromorelin is a ghrelin-receptor agonist that activates the growth hormone secretagogue receptor on delta cells. We hypothesized that in cats, capromorelin administration will result in decreased GV at the expense of reduced insulin secretion and glucose tolerance. Seven healthy cats were treated with capromorelin from days 1-30. After the first day, fasting blood glucose increased (+13 ± 3 mg/dL, P < 0.0001), insulin decreased (+128 ± 122 ng/dL, P = 0.03), and glucagon was unchanged. Blood glucose was increased throughout an intravenous glucose tolerance test on day 1 with blunting of first-phase insulin response ([FPIR] 4,931 ± 2,597 ng/L/15 min) compared with day -3 (17,437 ± 8,302 ng/L/15 min, P = 0.004). On day 30, FPIR was still blunted (9,993 ± 4,285 ng/L/15 min, P = 0.045), but glucose tolerance returned to baseline. Mean interstitial glucose was increased (+19 ± 6 mg/dL, P = 0.03) on days 2-4 but returned to baseline by days 27-29 (P = 0.3). On days 2-4, GV was increased (SD = 9.7 ± 3.2) compared with baseline (SD = 5.0 ± 1.1, P = 0.02) and returned to baseline on days 27-29 (SD = 6.1 ± 1.1, P = 0.16). In summary, capromorelin caused a decline in insulin secretion and glycemic control and an increase in glucose variability early in the course of treatment, but these effects diminished toward the end of 30 d of treatment.

Glucocorticoids differentially regulate the expression of CRFR1 and CRFR2α in MIN6 insulinoma cells and rodent islets.

Urocortin 3 (Ucn 3), member of the corticotropin-releasing factor (CRF) family of peptide hormones, is released from ß-cells to potentiate insulin secretion. Ucn 3 activates the CRF type-2 receptor (CRFR2) but does not activate the type-1 receptor (CRFR1), which was recently demonstrated on ß-cells. While the direct actions of Ucn 3 on insulin secretion suggest the presence of cognate receptors within the islet microenvironment, this has not been established. Here we demonstrate that CRFR2α is expressed by MIN6 insulinoma cells and by primary mouse and human islets, with no detectable expression of CRFR2ß. Furthermore, stimulation of MIN6 cells or primary mouse islets in vitro or in vivo with glucocorticoids (GCs) robustly and dose-dependently increases the expression of CRFR2α, while simultaneously inhibiting the expression of CRFR1 and incretin receptors. Luciferase reporters driven by the mouse CRFR1 or CRFR2α promoter in MIN6 cells confirm these differential effects of GCs. In contrast, GCs inhibit CRFR2α promoter activity in HEK293 cells and inhibit the expression of CRFR2ß in A7r5 rat aortic smooth muscle cells and differentiated C2C12 myotubes. These findings suggest that the GC-mediated increase of CRFR2α depends on the cellular context of the islet and deviates from the GC-mediated suppression of CRFR1 and incretin receptors. Furthermore, GC-induced increases in CRFR2α expression coincide with increased Ucn 3-dependent activation of cAMP and MAPK pathways. We postulate that differential effect of GCs on the expression of CRFR1 and CRFR2α in the endocrine pancreas represent a mechanism to shift sensitivity from CRFR1 to CRFR2 ligands.

The first appearance of Rodlet cells in carp (Cyprinus carpio L.) ontogeny and their possible roles during stress and parasite infection.

The origin and function of rodlet cells (RCs) are still a matter of discussion. Whereas the exogenous hypothesis considers them parasites, the endogenous hypothesis regards them as a genuine fish cell population with a secretory and/or leukocyte function. In order to shed more light on these questions we focused on the location and appearance of RCs during carp (Cyprinus carpio) ontogeny. Typical RCs were seen at 5days post fertilisation (dpf) between kidney and intestine, at 6dpf in the intestine and at 8dpf in both anterior and posterior kidney and in the abdominal cavity among the mesothelial cells. The RC number increased with age and after 14dpf they were also present in gills. The early appearance of the RCs during carp ontogeny support the endogenous hypothesis stating that RCs are genuine constituents of fish tissue and suggest that they are 'immune cells'. The fact that the RCs of the gills secrete their content into the surrounding water, combined with the strategic location around blood vessels in kidney and within intestinal epithelium, would also support an important role in host defense. To investigate whether RC numbers in gills and kidney are related to typical fluctuations in the physiology during stress and infection we counted their number in gills and kidney after parasite infection and stress. In the gills the number of RCs increased after infection but did not change after stress while in the kidney their number increased after stress and no significant changes were observed after infection.

Endurance exercise differentially stimulates heart and axial muscle development in zebrafish (Danio rerio).

Mechanical load is an important factor in the differentiation of cells and tissues. To investigate the effects of increased mechanical load on development of muscle and bone, zebrafish were subjected to endurance swim training for 6 h/day for 10 wk starting at 14 days after fertilization. During the first 3 wk of training, trained fish showed transiently increased growth compared with untrained (control) fish. Increased expression of proliferating cell nuclear antigen suggests that this growth is realized in part through increased cell proliferation. Red and white axial muscle fiber diameter was not affected. Total cross-sectional area of red fibers, however, was increased. An improvement in aerobic muscle performance was supported by an increase in myoglobin expression. At the end of 10 wk of training, heart and axial muscle showed increased expression of the muscle growth factor myogenin and proliferating cell nuclear antigen, but there were major differences between cardiac and axial muscle. In axial muscle, expression of the "slow" types of myosin and troponin C was increased, together with expression of erythropoietin and myoglobin, which enhance oxygen transport, indicating a shift toward a slow aerobic phenotype. In contrast, the heart muscle shifts to a faster phenotype but does not become more aerobic. This suggests that endurance training differentially affects heart and axial muscle.

Corticotropin-releasing hormone-receptor 1 (CRH-R1) and CRH-binding protein (CRH-BP) are expressed in the gills and skin of common carp Cyprinus carpio L. and respond to acute stress and infection.

We established that corticotropin-releasing hormone (CRH), CRH-binding protein (CRH-BP) and CRH-receptor 1 (CRH-R1) are expressed in the gills and skin of common carp Cyprinus carpio, an early vertebrate. Immunoreactive CRH was detected in macrophage-like cells in gills and skin, in fibroblasts in the skin and in endothelial cells in the gills. The involvement of the CRH system in gills and skin was investigated in response to infection and in an acute restraint stress paradigm. Carp were infected with the protozoan leech-transmitted blood flagellate Trypanoplasma borreli and subjected to acute restraint stress by netting for 24 h. The expression of CRH-BP and CRH-R1 genes in the gills and in the skin is downregulated after both infection and restraint. Thus the peripheral CRH system reacts to infection and stress. The gills and skin separate the internal from the external environment and are permanently exposed to stress and pathogens. Because of their pivotal role in maintaining the homeostatic equilibrium, these organs must act locally to respond to diverse stresses. Clearly, the CRH system is involved in the response of the integument to diverse stresses at the vulnerable interface of the internal and external milieu.

Structural characterisation of a cyprinid (Cyprinus carpio L.) CRH, CRH-BP and CRH-R1, and the role of these proteins in the acute stress response.

We elucidated the structure of the principle factors regulating the initiation of the acute stress response in common carp: corticotrophin-releasing hormone (CRH), CRH-receptor 1 (CRH-R1) and CRH-binding protein (CRH-BP). Phylogenetic analyses reveal that these proteins are evolutionarily well conserved in vertebrates. CRH and CRH-BP expression are not co-localised in the same hypothalamic perikarya. On the contrary, CRH-BP expression is limited to the perimeter of the nucleus preopticus (NPO), but is abundant in other regions, including an area directly rostral from, and in close proximity to, the NPO. Despite the lack of co-expression, the nerve fibres projecting onto both the rostral pars distalis (rPD) as well as the large fibre bundles projecting onto the pars intermedia (PI) contain CRH as well as CRH-BP, suggesting that both ACTH release from the rPD as well as the release of PI melanotrope content is regulated via CRH and CRH-BP. Finally, we show via real-time quantitative PCR that expression of hypothalamic CRH and CRH-BP following a 24 h restraint significantly increases, whereas PD CRH-R1 expression decreases; this reflects desensitisation of the PD for hypothalamic CRH output. We conclude that these factors are actively involved in the regulation of acute stress responses in the teleost fish.