Dual-energy X-ray absorptiometry scans accurately predict differing body fat content in live sheep.

BACKGROUND: There is considerable interest in implementing mobile scanning technology for on-farm body composition analysis on live animals. These experiments evaluated the use of dual energy X-ray absorptiometry (DXA) as an accurate method of total body fat measurement in live sheep. RESULTS: In Exp. 1, visceral and whole body fat analysis was undertaken in sheep with body condition scores (BCS) in the range 2 to 3.25 (scale 1: thin to 5: fat). The relationship of BCS was moderately correlated with visceral fat depot mass (r = 0.59, P < 0.01, n = 24) and whole body fat (r = 0.70, P < 0.001, n = 24). In Exp. 2, sheep with BCS in the range 2.25 to 3.75 were blood sampled to analyse circulating leptin concentrations, and were DXA scanned immediately post mortem for total body fat. Plasma leptin concentrations had low correlations with BCS (r = 0.50, P < 0.05, n = 17) and DXA body fat (r = 0.42, P < 0.05, n = 17), and no correlation with chemical body fat (r = 0.17, P > 0.05, n = 9). There was a moderate correlation between DXA body fat and BCS (r = 0.70, P < 0.01, n = 17), and DXA body fat was highly correlated with chemical body fat (r = 0.81, P < 0.001, n = 9). In Exp. 3, a series of five DXA scans, at 8-week intervals, was performed on growing sheep over a 32-week period. The average BCS ranged from 2.39 ± 0.07 (S.E.M.) to 3.05 ± 0.11 and the DXA body fat (%) ranged from 16.8 ± 0.8 to 24.2 ± 1.2. There was a moderate correlation between DXA body fat and BCS over the 32 weeks (r = 0.61, P < 0.001, n = 24). CONCLUSIONS: Overall, these experiments indicated that there was good agreement between BCS, DXA and chemical analysis for measuring total body fat in sheep, and that DXA scanning is a valid method for longitudinal measurement of total body fat in live sheep.

Direct conscious telemetry recordings demonstrate increased renal sympathetic nerve activity in rats with chronic kidney disease.

Chronic kidney disease (CKD) is associated with sympathetic hyperactivity and impaired blood pressure control reflex responses, yet direct evidence demonstrating these features of autonomic dysfunction in conscious animals is still lacking. Here we measured renal sympathetic nerve activity (RSNA) and mean arterial pressure (MAP) using telemetry-based recordings in a rat model of CKD, the Lewis Polycystic Kidney (LPK) rat, and assessed responses to chemoreflex activation and acute stress. Male LPK and Lewis control animals (total n = 16) were instrumented for telemetric recording of RSNA and MAP. At 12-13 weeks-of-age, resting RSNA and MAP, sympathetic and haemodynamic responses to both peripheral (hypoxia: 10% O2) and central chemoreflex (hypercapnia: 7% CO2) activation and acute stress (open-field exposure), were measured. As indicators of renal function, urinary protein (UPro) and creatinine (UCr) levels were assessed. LPK rats had higher resting RSNA (1.2 ± 0.1 vs. 0.6 ± 0.1 µV, p < 0.05) and MAP (151 ± 8 vs. 97 ± 2 mmHg, p < 0.05) compared to Lewis. MAP was negatively correlated with UCr (r = -0.80, p = 0.002) and positively correlated with RSNA (r = 0.66, p = 0.014), with multiple linear regression modeling indicating the strongest correlation was with Ucr. RSNA and MAP responses to activation of the central chemoreflex and open-field stress were reduced in the LPK relative to the Lewis (all p < 0.05). This is the first description of dual conscious telemetry recording of RSNA and MAP in a genetic rodent model of CKD. Elevated RSNA is likely a key contributor to the marked hypertension in this model, while attenuated RSNA and MAP responses to central chemoreflex activation and acute stress in the LPK indicate possible deficits in the neural processing of autonomic outflows evoked by these sympathoexcitatory pathways.

Cardiovascular autonomic dysfunction in a novel rodent model of polycystic kidney disease.

Autonomic dysfunction, hypertension and cardiovascular morbidity in end stage renal disease are critically linked, however there are limited models available to investigate this relationship and develop clinical interventions. This study aimed to define the relationship between hypertension and autonomic function in a new rodent model of polycystic kidney disease (PKD). Using measures of heart rate and systolic blood pressure variability (HRV, SBPV), and time domain analysis of cardiac and sympathetic baroreflex function, we compared the Lewis PKD model (LPK) to a Lewis control. Systolic BP and SBPV were significantly higher in LPK vs. Lewis (168+/-7 vs. 131+/-8mm Hg, P

Photoperiod influences the central effects of ghrelin on food intake, GH and LH secretion in sheep.

Ghrelin is a circulating peptide, primarily secreted by the gut, that has reported actions within the hypothalamo-pituitary axis to stimulate food intake, inhibit GnRH/LH secretion and stimulate GH secretion in monogastric species. Here, we examine responses to centrally administered ghrelin in a seasonal ruminant. Estradiol-implanted castrated male sheep with indwelling intracerebroventricular (i.c.v.) cannulae were kept with unrestricted food for 16 weeks in long day photoperiod (LD, 16 h light/day) then 16 weeks in short days (SD, 8 h light/day). In week 16 of each photoperiod they were given a control (saline) i.c.v. injection on day 1 and ghrelin i.c.v. injection on day 2. Mean circulating endogenous plasma ghrelin concentrations showed no diurnal pattern and were similar between the photoperiods. Central ghrelin injection increased voluntary food intake 2-fold in the first hour after administration in LD but not in SD, decreased LH pulse frequency and amplitude in SD but not in LD, and stimulated GH release in both photoperiods, although there was a 1.5-fold larger response in LD. Therefore, central injection of ghrelin to sheep acutely stimulated food intake in LD, suppressed reproductive neuroendocrine output in SD, and stimulated GH secretion irrespective of photoperiod, although more pronounced in LD. These data indicate that photoperiod can influence hypothalamic appetite and reproductive neuroendocrine responses to ghrelin in seasonal species.

Nutritional influences on reproductive neuroendocrine output: insulin, leptin, and orexigenic neuropeptide signaling in the ovine hypothalamus.

This study investigated how changing nutritional status may alter reproductive neuroendocrine (LH) output via circulating leptin and insulin signaling through orexigenic hypothalamic pathways. Thin sheep were given an increasing nutritional plane (INP), sheep with intermediate adiposity a static nutritional plane (SNP), and fat sheep a decreasing nutritional plane (DNP) for 6 wk. Mean group adiposities converged by wk 6, LH output increased in INP, remained unchanged in SNP, and decreased in DNP sheep. Plasma and cerebrospinal fluid (CSF) insulin and plasma leptin concentrations increased in INP but did not change in the SNP and DNP groups. In INP sheep, LH output correlated positively with adiposity and plasma and CSF insulin concentrations and negatively with orexigenic neuropeptide Y gene expression in the hypothalamic arcuate nucleus (ARC). In DNP sheep, LH output correlated positively with adiposity, CSF leptin concentrations, and ARC proopiomelanocortin gene expression and negatively with leptin receptor (OB-Rb) and agouti-related peptide gene expression in the ARC. These data are consistent with the feedback response to an increasing nutritional plane being mediated by increasing circulating insulin entering the brain and stimulating LH via inhibition of hypothalamic neuropeptide Y and the response to a decreasing nutritional plane being mediated by altered hypothalamic leptin signaling brought about by increased OB-Rb expression and decreased melanocortin signaling. Because end point adiposity was similar yet LH output was different, the hypothalamus apparently retains a nutritional memory, based on changes in orexigenic neuropeptide expression, that influences contemporary neuroendocrine responses.

An immunohistochemical study of the localization and developmental expression of ghrelin and its functional receptor in the ovine placenta.

BACKGROUND: Ghrelin is an orexigenic hormone principally produced by the stomach, but also by numerous peripheral tissues including the placenta. Ghrelin acts via growth hormone secretagogue receptors (GHSR-1a) to alter food intake, fat utilization, and cellular proliferation, and has been suggested to play a role in the developmental growth of the fetoplacental unit. The placental expression of ghrelin and its role in ruminant species is not known. We tested the hypotheses that ghrelin and its functional receptor, GHSR-1a, are present in tissues of the ovine placenta, and that their expression is linked to the stage of development. METHODS: Antibodies raised against ghrelin and GHSR-1a were used in standard immunohistochemical protocols on placental tissues collected from pregnant ewes (n = 6 per gestational time point) at days 50, 80, 100, 128 and 135 of gestation (term approximately day 145). Immunostaining for ghrelin and GHSR-1a was quantified using computer-aided image analysis. Image analysis data were subjected to one-way ANOVA, with differences in immunostaining between time-points determined by Fisher's least significant difference. RESULTS: Positive immunostaining for ghrelin was detected in ovine placentae at all gestational time points, with staining localized to the maternal epithelium, caruncle and trophectoderm. There was a significant effect of gestational age (p < 0.001) on the placental expression of ghrelin, with maximal levels at gestational day 80. GHSR-1a immunostaining was detected in the fetal trophectoderm at all time points. In contrast to the gestational pattern of ghrelin expression, there was no effect of gestational age on placental GHSR-1a immunoexpression. CONCLUSION: Ghrelin and GHSR-1a are both present in the ovine placenta, and ghrelin displays a developmentally-related pattern of expression. Therefore, these data strongly suggest that the ghrelin system may have a role in feto-placental development in sheep.

Temporal relationship between renal cyst development, hypertension and cardiac hypertrophy in a new rat model of autosomal recessive polycystic kidney disease.

BACKGROUND/METHODS: We have examined the hypothesis that cyst formation is key in the pathogenesis of cardiovascular disease in a Lewis polycystic kidney (LPK) model of autosomal-recessive polycystic kidney disease (ARPKD), by determining the relationship between cyst development and indices of renal function and cardiovascular disease. RESULTS: In the LPK (n = 35), cysts appear at week 3 (1.1 +/- 0.1 mm) increasing to week 24 (2.8 +/- 2 mm). Immunostaining for nephron-specific segments indicate cysts develop predominantly from the collecting duct. Cyst formation preceded hypertension (160 +/- 22 vs. Lewis control 105 +/- 20 mm Hg systolic blood pressure (BP), n = 12) at week 6, elevated creatinine (109 +/- 63 vs. 59 +/- 6 micromol/l, n = 16) and cardiac mass (0.7 vs. 0.4% bodyweight, n = 15) at week 12, and left ventricular hypertrophy (2,898 +/- 207 vs. 1,808 +/- 192 mum, n = 14) at week 24 (all p < or = 0.05). Plasma-renin activity and angiotensin II were reduced in 10- to 12-week LPK (2.2 +/- 2.9 vs. Lewis 11.9 +/- 4.9 ng/ml/h, and 25.0 +/- 19.1 vs. 94.9 +/- 64.4 pg/ml, respectively, n = 26, p < or = 0.05). Ganglionic blockade (hexamethonium 3.3 mg/kg) significantly reduced mean BP in the LPK (52 vs. Lewis 4%, n = 9, p < or = 0.05). CONCLUSION: Cyst formation is a key event in the genesis of hypertension while the sympathetic nervous system is important in the maintenance of hypertension in this model of ARPKD.

Immunohistochemical evidence for an endocrine/paracrine role for ghrelin in the reproductive tissues of sheep.

BACKGROUND: The gut hormone, ghrelin, is involved in the neuroendocrine and metabolic responses to hunger. In monogastric species, circulating ghrelin levels show clear meal-related and body weight-related changes. The pattern of secretion and its role in ruminant species is less clear. Ghrelin acts via growth hormone secretagogue receptors (GHSR-1a) to alter food intake, fat utilization, and cellular proliferation. There is also evidence that ghrelin is involved in reproductive function. In the present study we used immunohistochemistry to investigate the presence of ghrelin and GHSR-1a in sheep reproductive tissues. In addition, we examined whether ghrelin and GHSR-1a protein expression is developmentally regulated in the adult and fetal ovine testis, and whether there is an association with markers of cellular proliferation, i.e. stem cell factor (SCF) and proliferating cell nuclear antigen (PCNA). METHODS: Antibodies raised against ghrelin and its functional receptor, GHSR-type 1a, were used in standard immunohistochemical protocols on various reproductive tissues collected from adult and fetal sheep. GHSR-1a mRNA presence was also confirmed by in situ hybridisation. SCF and PCNA immunoexpression was investigated in fetal testicular samples. Adult and fetal testicular immunostaining for ghrelin, GHSR-1a, SCF and PCNA was analysed using computer-aided image analysis. Image analysis data were subjected to one-way ANOVA, with differences in immunostaining between time-points determined by Fisher's least significant difference. RESULTS: In adult sheep tissue, ghrelin and GHSR-1a immunostaining was detected in the stomach (abomasum), anterior pituitary gland, testis, ovary, and hypothalamic and hindbrain regions of the brain. In the adult testis, there was a significant effect of season (photoperiod) on the level of immunostaining for ghrelin (p < 0.01) and GHSR-1a (p < 0.05). In the fetal sheep testis, there was a significant effect of gestational age on the level of immunostaining for ghrelin (p < 0.001), GHSR-1a (p < 0.05), SCF (p < 0.05) and PCNA (p < 0.01). CONCLUSION: Evidence is presented for the presence of ghrelin and its receptor in various reproductive tissues of the adult and fetal sheep. In addition, the data indicate that testicular expression of ghrelin and its receptor is physiologically regulated in the adult and developmentally regulated in the fetus. Therefore, the ghrelin ligand/receptor system may have a role (endocrine and/or paracrine) in the development (cellular proliferation) and function of the reproductive axis of the sheep.