Differences in mitochondrial DNA inheritance and function align with body conformation in genetically lean and fat sheep.

Body weight and adiposity are determined by the balance between energy intake, energy expenditure, and nutrient deposition. We have identified differences in appetite-regulating peptides in sheep selectively bred to be either lean or fat, wherein gene expression for orexin and melanin-concentrating hormone are elevated in the lean group. Despite this, the underlying mechanisms leading to differences in body composition in the lean and fat lines remains unknown. We measured postprandial temperature in adipose tissue and muscle to ascertain whether a difference in thermogenesis is associated with the difference in body composition in genetically lean (n = 8) and fat (n = 12) ewes. Body weight was higher (P < 0.01) but percent fat mass was lower (P < 0.001) in the lean group. The percent lean mass was similar in lean and fat groups. Animals received intracerebroventricular cannulae and temperature probes implanted into the retroperitoneal fat and the hind-limb skeletal muscle (vastus lateralis). Animals were meal fed (1100-1600 h) to entrain postprandial thermogenesis. Food intake was similar between lean and fat animals. Postprandial thermogenesis was greater (P < 0.05) in the retroperitoneal adipose tissue of lean animals but not in skeletal muscle. Intracerebroventricular infusion of leptin reduced (P< 0.05) food intake by an equal extent in both groups. Postprandial expression of UCP1 mRNA was greater (P < 0.05) in retroperitoneal fat of lean animals, with similar UCP3 expression in skeletal muscle. Mitochondrial genome sequencing indicated haplotypic clustering in lean and fat animals within both the encoding and nonencoding regions. This demonstrates that differences in body composition may be underpinned by differences in thermogenesis, specifically within adipose tissue. Furthermore, thermogenic differences may be associated with specific mitochondrial DNA haplotypes, suggesting a strong genetic component inherited through the maternal lineage.

High cortisol response to adrenocorticotrophic hormone identifies ewes with reduced melanocortin signalling and increased propensity to obesity.

We have identified female sheep that have either high (HR) or low (LR) cortisol responses to adrenocorticotrophin. On a high-energy diet, HR have greater propensity to weight gain and obesity, although the underlying mechanisms remain to be determined. Hypothalamic appetite-regulating peptides (ARP) exert reciprocal effects on food intake and energy expenditure. We aimed to quantify the expression and function of ARP in LR and HR ewes (n = 4 per group). Gene expression for neuropeptide Y (NPY), agouti-related peptide (AgRP) pro-opiomelanocortin (POMC), melanin-concentrating hormone (MCH), orexin and the melanocortin receptors (MC3R and MC4R) was measured by in situ hybridisation. Expression of NPY, AgRP and POMC was similar in HR and LR, although expression of orexin, MCH, MC3R and MC4R was higher (P < 0.05) in LR. Intracerebroventricular infusions of a low dose (50 µg/h) of NPY, α-melanocyte-stimulating hormone (αMSH), orexin and MCH were performed between 10.00 h and 16.00 h in meal-fed ewes (n = 6-7 per group). Skeletal muscle and retroperitoneal (RP) fat temperatures were recorded using dataloggers. Post-prandial thermogenesis in muscle was higher (P < 0.05) in LR. There was little effect of ARP infusion on muscle or fat temperature in either group. Infusion of these doses of NPY, MCH or orexin did not stimulate food intake in meal-fed ewes, although αMSH reduced (P < 0.01) food intake in LR only. Using 24-h ARP infusions with ad lib. feeding, NPY increased (P < 0.001) food intake in both groups but αMSH was only effective in LR (P < 0.05). In summary, we show that HR are resistant to the satiety effects of αMSH and this coincides with a reduced expression of both the MC3R and MC4R in the paraventricular nucleus of the hypothalamus. We conclude that an increased propensity to obesity in HR female sheep is associated with reduced melanocortin signalling.

Global trends in wind speed and wave height.

Studies of climate change typically consider measurements or predictions of temperature over extended periods of time. Climate, however, is much more than temperature. Over the oceans, changes in wind speed and the surface gravity waves generated by such winds play an important role. We used a 23-year database of calibrated and validated satellite altimeter measurements to investigate global changes in oceanic wind speed and wave height over this period. We find a general global trend of increasing values of wind speed and, to a lesser degree, wave height, over this period. The rate of increase is greater for extreme events as compared to the mean condition.

Development of the technical capabilities needed to build and position a prepolarization coil for a magnetic resonance imaging magnet.

An experiment to show that a magnetic resonance imaging (MRI) magnet could be assembled around a patient, and used as part of a prepolarization system in which substantial transient forces are applied to parts of it, is described. The paper describes the circumstances that develop as a result of the application of the large transient fields used in this type of study, and outlines the reason for the tolerances that are permissible on the alignment of the system components. It then describes a test rig used to evaluate how the various problems might be overcome, and reports on the performance achieved with this rig. On the basis of this work, it appears that a system could be developed that would allow the application of these methods in clinical MRI.

Mechanisms responsible for parturition; the use of experimental models.

In this review, our knowledge, gleaned from a range of species, of what determines gestation length, how fetal maturation and birth are synchronized and how the uterotonic mechanisms are activated at birth are discussed. Accumulated data indicate that fetal glucocorticoids are involved in, but do not necessarily play a causative role in, the initiation of parturition in eutherian mammals generally. Present observations are consistent with a complex, positive regulatory interaction between estrogens, prostaglandins and oxytocin and are consistent with a role for prostaglandins as the final, common effector in myometrial activation. We are, however, left with the possibility that the initial mechanism for the timing of birth is encoded in the fetal genome and is closely linked to, and activated when, certain prerequisite developmental events have occurred in the fetus. Our understanding of these events in the sheep have led to its extensive use as an experimental model for the study of human clinical correlates of fetal maturation and development and the control of the initiation of parturition.

Changes in the expression of prostaglandin E and F synthases at induced and spontaneous labour onset in the sheep.

The differential production of prostaglandin (PG) F(2 alpha) and PGE(2) within the uterine compartment may play a role in controlling myometrial contraction. We hypothesized that the enzymes downstream of PG endoperoxide synthase-2 (PGHS-2) determine the ratio of PGF(2 alpha) and PGE(2) in the utero-ovarian vein plasma and the time of normal and preterm labour onset. The aim of this study was to simultaneously determine the expression of PGF and PGE synthases (PGFS and PGES) in gestational tissues at spontaneous and induced-preterm labour in sheep. Myometrial, endometrial and placental tissue were obtained from ewes in dexamethasone-induced preterm labour, age-matched control ewes, and ewes in spontaneous term labour for analysis of mRNA expression by real-time PCR. PGFS mRNA expression was significantly increased following dexamethasone-induced and spontaneous labour onset in placentome (P<0.01) but was unchanged in the myometrium and endometrium. In contrast, PGES mRNA expression remained unchanged or decreased. PGHS-2 mRNA expression was increased in all tissues examined in both dexamethasone-induced and spontaneous labour (P<0.001). Plasma PGE(2) and PGF(2 alpha) concentrations rose in both dexamethasone-induced and spontaneous labour with the ratio of PGF(2 alpha):PGE(2) increased with labour onset (P<0.05). These results are consistent with the hypothesis that the increased expression, of PGFS is responsible for the increased PGF(2 alpha):PGE(2) ratio and this, together with increased PGHS-2 expression, accounts for myometrial activity at labour onset. The findings point to PGFS expression as a key factor in regulating the uterotonic process in the sheep.

Combined xylazine and ketamine as an analgesic regimen in sheep.

OBJECTIVE: To determine whether low dose xylazine with ketamine reduces the concentrations of cortisol and prolactin in sheep postoperatively and to characterise the effects of the drugs on behaviour during recovery. DESIGN: Analysis of variance was used to compare the effects of anaesthesia, surgery and combined ketamine/xylazine treatment on the plasma cortisol and prolactin concentrations and on behavioural variables in pregnant ewes subjected to abdominal surgery. PROCEDURE: Twelve ewes were randomly assigned to receive either ketamine/xylazine or placebo in association with anaesthesia and surgery. Both groups of ewes underwent anaesthesia alone followed a week later by anaesthesia with laparotomy and hysterotomy. Plasma cortisol and prolactin concentrations were assayed during these procedures and for 5 days afterwards. Behavioural observations were made remotely during recovery from anaesthesia and anaesthesia plus surgery. RESULTS: The concentrations of cortisol in the plasma of pregnant ewes undergoing surgery were increased by preoperative handling and the onset of thiopentone/halothane anaesthesia, with a further increase during surgery (P = 0.033). Cortisol concentrations decreased over the first four postoperative hours (P = 0.029) and were normal by 24 h. The drug treatment did not affect the immediate responses of ewes to anaesthesia or surgery, although treated ewes had lower cortisol concentrations than saline-treated controls over the first five postoperative days (P = 0.018). Prolactin concentrations increased in response to anaesthesia (P = 0.047), but were not affected by surgery or the drug treatment. Drug-treated ewes had prolonged sleeping time after surgery (P = 0.002), but they took no longer to stand than saline-treated controls and required fewer attempts to stand successfully (P = 0.025). CONCLUSION: At the doses used, ketamine and xylazine did not mitigate the immediate endocrine consequences of surgery but the behavioural data provide a basis for further investigations that may lead to improvements in analgesic treatments.

The role of the pituitary gland and ACTH in the regulation of mRNAs encoding proteins essential for adrenal steroidogenesis in the late-gestation ovine fetus.

To further understand the relative roles of the pituitary gland and ACTH in the regulation of mRNAs encoding proteins that are essential for adrenal development, we investigated the effects of, first, an ACTH infusion and labour in intact fetuses and, secondly, the effect of an ACTH infusion to fetuses with and without a pituitary gland, on the relative abundance of the mRNA encoding for the ACTH receptor (MC2R), steroidogenic factor 1 (SF-1), cholesterol side-chain cleavage enzyme (P450(scc)), 3beta-hydroxysteroid dehydrogenase (3betaHSD) and 17alpha-hydroxylase (P450(C17)) in the fetal adrenal gland. ACTH(1-24) infusion (14.7 pmol/kg per h) to intact fetuses was without effect on the abundance of mRNA encoding MC2R and SF-1, irrespective of whether the infusion was given for 18 (115-132 days of gestation) or 32 days (115 days to term (147 days of gestation)). Hypophysectomy (HX) did not alter the expression of MC2R mRNA; however, the abundance of SF-1 mRNA fell by approximately 50% following the removal of the pituitary gland. ACTH(1-24) infusion to HX fetuses failed to restore levels of SF-1 mRNA to that seen in intact animals. P450(scc) and 3betaHSD mRNAs were increased by ACTH(1-24) infusion for 18 days in intact animals, although no effects of the infusion were seen on P450(C17) mRNA levels. For all three of these mRNAs, there was a significant increase in their abundance between 132 days of gestation and term in intact fetuses. By term, ACTH(1-24) infusion was without any additional effect on their abundance. HX decreased the expression of P450(scc), 3betaHSD and P450(C17) mRNAs, while ACTH(1-24) infusion to HX fetuses increased the expression of these mRNAs to levels seen in intact animals. There were significant correlations between the abundance of the mRNA for P450(scc), 3betaHSD and P450(C17), but not MC2R and SF-1, and premortem plasma cortisol concentrations. These results emphasise the importance of the pituitary gland and ACTH in the regulation of the enzymes involved in adrenal steroidogenesis. Factors in addition to ACTH may also play some role, as the infusion was not always effective in increasing the abundance of the mRNAs. Surprisingly, the mRNA for MC2R and SF-1 did not appear to be regulated by ACTH in the late-gestation ovine fetus, though a pituitary-dependent factor may be involved in the regulation of SF-1 mRNA abundance.

Microstructured magnetic materials for RF flux guides in magnetic resonance imaging.

Magnetic resonance imaging and spectroscopy systems use coils, either singly or as arrays, to intercept radio-frequency (RF) magnetic flux from regions of interest, often deep within the body. Here, we show that a new magnetic material offers novel possibilities for guiding RF flux to the receiver coil, permitting a clear image to be obtained where none might otherwise be detectable. The new material contains microstructure designed according to concepts taken from the field of photonic band gap materials. In the RF range, it has a magnetic permeability that can be produced to specification while exhibiting negligible direct-current magnetism. The latter property is vital to avoid perturbing the static and audio-frequency magnetic fields needed to obtain image and spectral data. The concept offers a new paradigm for the manipulation of RF flux in all nuclear magnetic resonance systems.

Catheter tracking for MR fluoroscopy: design of a transmit/receive coil for use with a nasogastric tube.

A coil tuned to 21.3 MHz was incorporated into a nasogastric tube and used as a marker of tube position during magnetic resonance (MR) imaging in a 0.5-T scanner. Catheter tracking was investigated with the coil used in both transmit/receive and in receive-only modes. Data acquired from this coil were overlaid on images obtained using the body coil of the scanner. Visualization of the full length of the catheter with local high signal at the tip was achieved with a temporal resolution of approximately 1 second. J. Magn. Reson. Imaging 2001;13:127-130.

Use of multicoil arrays for separation of signal from multiple slices simultaneously excited.

Increased acquisition efficiency has been achieved by exciting several slices simultaneously. The mixed data were unfolded to produce separate slices using the spatial encoding information inherent in a multicoil receiver system. Each coil yields a linear combination of signals from all excited slices weighted by the sensitivity of each coil. A matrix inversion provides a solution to unfold these images.

Immunohistochemical localization of prostaglandin G/H synthase 1 and 2 in sheep placenta after glucocorticoid-induced and spontaneous labour.

Enhanced prostaglandin production and release by the placenta is an essential element in the normal transition to labour in many animal species. In sheep, expression of prostaglandin G/H synthase (PGHS) is the central enzyme regulating this process. In this study immunohistochemistry was used to examine the distribution of cells expressing PGHS-1 and PGHS-2 in ovine placenta in association with spontaneous parturition (n = 6) and glucocorticoid-induced labour (n = 5). Labour was induced in ewes after the intrafetal injection of betamethasone on day 131 of gestation. Animals administered an intrafetal injection of isotonic saline (n = 5) acted as non-labour controls. In placentomes collected from all groups, immunoreactive PGHS-1 was present in the mononuclear trophoblast cells of the fetal placenta. Cells in the maternal mesenchyme and epithelial syncytium were weakly immunopositive for this enzyme. PGHS-1 immunoreactivity was also demonstrated in the endothelial cells of the chorionic vessels. The PGHS-2 isozyme was localized exclusively to the trophoblast epithelial cells. Immunoreactive PGHS-2 was not detectable in the maternal epithelial syncytium or the stroma of the cotyledons. The binucleate cells of the fetal placenta were consistently immunonegative for both PGHS isozymes. These results indicate that the cellular localization of PGHS-1 and PGHS-2 in ovine placenta does not change during the last 15 days of pregnancy. Co-localization of these isozymes indicates that the source of arachidonic acid and the site of prostanoid formation are the same. Quantitation of the percentage area of positive staining for PGHS-1 and PGHS-2 using image analysis software demonstrated a significant increase in PGHS-2 in the fetal trophoblast after glucocorticoid-induced labour and spontaneous parturition. This finding indicates that increased formation of the PGHS-2 isozyme is responsible for the large increase in prostaglandin production by the ovine placenta at term labour.

SAR and temperature changes in the leg due to an RF decoupling coil at frequencies between 64 and 213 MHz.

Specific absorption rate (SAR) due to a butterfly surface coil in a realistic model of the leg was calculated for frequencies 64 < or = v < or = 213 MHz. The resulting temperature distribution and temperature changes (deltaT) were found using the bioheat transfer equation. To compare results at different frequencies, the minimum B-field within the coil's footprint in the plane parallel to the coil but displaced 50 mm from it was kept constant. To achieve the same minimum B-field as that associated with operation at 64 MHz that was compliant with safety guidelines (peak SAR in 1 cm3 of tissue of 2.3 W/kg), it was predicted that SAR would exceed recommended levels when v > or = 149 MHz. The corresponding maximum deltaT at 64, 128, and 213 MHz were 0.3, 1.3, and 5.0 degrees C, respectively.

Notes on current safety issues in MRI.

The present unsatisfactory state of the worldwide regulatory system for whole body magnetic resonance is explored. The priorities of a number of the most important regulators are outlined, and the differences between factors affecting the safety of patients on the one hand, and of those operating equipment on the other, are discussed. At the end, a strategy is outlined for obtaining the data needed to establish new, more firmly based, limits of operation. A prime factor in getting this data will be collaboration between the regulators, users of equipment, and manufacturers. A measure of urgency is desirable as the longer the present situation is allowed to persist the greater the risk that the stresses already present in it will become intolerable.

Localisation and temporal changes in prostaglandin G/H synthase-1 and -2 content in ovine intrauterine tissues in relation to glucocorticoid-induced and spontaneous labour.

Parturition in the ewe is preceded by an increase in the synthesis of prostaglandins (PGs) by gestational tissues. To establish the uterine source of these PGs, placental cotyledons, fetal membranes and maternal uterine tissues were collected from ewes (n=6) at spontaneous parturition. Solubilised tissue extracts were prepared and analysed by Western blots using polyclonal antibodies to PG G/H synthase-1 and -2 (PGHS-1 and PGHS-2). PGHS-1 was expressed by all intrauterine tissues at term labour. Densitometric analysis of Western blot autoradiographs showed that the fetal membranes and maternal cervix contained the largest amounts of PGHS-1. PGHS-1 enzyme content of ovine amnion was significantly greater than that of either chorion or allantois (P<0.05). PGHS-1 protein content of myometrial, endometrial and cotyledonary tissue extracts was minimal. Formation of the PGHS-2 isozyme was confined to placental tissue at term labour. PGHS-2 protein levels in sheep placenta were significantly higher than those of PGHS-1 in all intrauterine tissues examined. This result supports the hypothesis that PGHS-2 is a major contributor to PG formation at term labour. To elucidate the developmental changes in PGHS-1 and PGHS-2 relative to labour onset, an experimental paradigm of glucocorticoid-induced delivery was used. Previous characterisation and validation of this labour model demonstrated that direct, transabdominal, intrafetal injection of the synthetic glucocorticoid betamethasone (5.7 mg in 1 ml aqueous vehicle) on day 131 of gestation induced labour onset in 56.6+/-0.8 h (mean+/-s.e.m.). As the latent period to induced-labour was known, the time course of enzyme formation could be ascertained. Sheep (n=20) were killed by barbiturate injection at various time intervals post-injection (0, 14, 28, 42 and 56 h). Tissue extracts collected at post-mortem examination were prepared and analysed by Western blots. PGHS-2 was induced in ovine cotyledon in a time-dependent fashion following glucocorticoid injection (P<0.05). There was a 12-fold increase in abundance between the time of betamethasone administration (0 h) and established labour (56 h). The PGHS-2 isozyme was not detected in any of the other tissues examined. In contrast, formation of the PGHS-1 isozyme did not change in relation to induced-labour in any of the intrauterine tissues. This finding is consistent with constitutive formation of PGHS-1. Previous studies have demonstrated a rise in PG production in association with glucocorticoid-induced labour and spontaneous delivery. The results of the present study indicate that this rise in PG production is due to increased formation of the PGHS-2 isozyme in ovine cotyledon. PGHS-2 appears to be induced by exogenous glucocorticoid administration and/or the mechanisms controlling ovine parturition. The role of PG formation by the fetal membranes is yet to be elucidated.

Influence of growth hormone on the lung growth response to tracheal obstruction in fetal sheep.

Obstructing the fetal trachea is a potent stimulus for fetal lung growth, but little is known about the factors that regulate this process. Our aim was to determine the role of growth hormone (GH) in regulating the increase in lung growth induced by obstruction of the trachea in fetal sheep. Twenty chronically catheterized fetal sheep, nine of which were hypophysectomized, were divided into four experimental groups: 1) control group (n = 4), 2) a group in which the fetal trachea was obstructed for 3 days (3-day obstructed; n = 6), 3) a 3-day obstructed group in which the pituitary was removed [hypophysectomized (HX)] and the fetus was given maintenance infusions of ACTH, thyroxine, and human GH (hGH; HX hGH 3-day obstructed; n = 5), and 4) a HX 3-day obstructed group in which the fetus was given maintenance infusions of ACTH and thyroxine (n = 5). Tracheal obstruction significantly increased fetal lung liquid volumes from 37.2 +/- 3.2 ml/kg in control fetuses to 75.6 +/- 9.0 ml/kg in 3-day obstructed fetuses, and the presence or absence of GH did not affect this increase. Similarly, the presence or absence of GH did not affect the increase in lung weight or protein content induced by 3 days of tracheal obstruction. However, in the absence of GH, 3 days of tracheal obstruction failed to increase total lung DNA content above unobstructed control values (107.9 +/- 5.3 and 94. 1 +/- 7.0 mg/kg for control and HX 3-day obstructed groups, respectively). In contrast, 3 days of tracheal obstruction increased total lung DNA content to a similar extent in fetuses with an intact pituitary and HX fetuses that received GH replacement (126.0 +/- 4.4 and 126.7 +/- 4.0 mg/kg for 3-day obstructed and HX hGH 3-day obstructed groups, respectively). These data indicate that the absence of GH either abolishes or delays the acceleration in cell division caused by an increase in fetal lung expansion.

Pre-polarized saline: an in vivo feasibility study of a potential contrast agent.

The potential for using pre-polarized liquids as contrast agents in vivo is investigated and the feasibility of the method demonstrated. In this study we show the enhancement obtained following intravenous delivery of pre-polarized saline into the antecubital vein of a volunteer. This form of contrast agent provides signal gain on time scales commensurate with its T(1) and allows repeated doses to be administered, thus making alternate acquisitions of data with and without enhancement practicable.