bluuebunny/crossref_metadata_2025 huggingface dataset:
Perimetric [Ca2+]i rise and exocytosis detected by ultraviolet laser scanning confocal microscopy in rat peritoneal mast cells
Authors: Habara Y , Kanno T
Published: December 2014

It has been reported that in dialysed mast cells an increase in mean [Ca2+]i is neither necessary nor sufficient for secretion; however, it is possible that juxtamembranal [Ca2+]i may exceed the mean [Ca2+]i before exocytosis. The present study was carried out to analyse spatial and temporal dynamics of [Ca2+]i and concomitant exocytosis in intact rat peritoneal mast cells using UV laser scanning confocal microscopy. Stimulation with Compound 48/80 (16 microM) increased mean [Ca2+]i, causing an initial rapid elevation from 40 to 200 nM, which lasted for 6 s and was followed by a delayed secondary increase to 600 nM. Exocytotic images were seen in the cell perimeter 16 s after the stimulation. Perimetric and nuclear [Ca2+]i increased to several mumoles per litre, while that in the intermediate region remained low. In a Ca(2+)−deficient environment, Compound 48/80 still increased perimetric [Ca2+]i to micromolar values and induced exocytosis. This study clearly indicates, for the first time, that a perimetric increase in [Ca2+]i to micromolar levels precedes exocytosis, in intact, as opposed to permeabilized cells.
The effect of distension of the stomach on peripheral blood flow in anaesthetized pigs
Authors: Vacca G , Mary DA , Battaglia A , Grossini E , Molinari C
Published: December 2014

The present study was undertaken in anaesthetized pigs to determine the primary reflex effects of gastric distension on the peripheral circulation. Changes in blood flow in the splenic, superior mesenteric, left renal and left external iliac arteries were assessed using electromagnetic flowmeters during distension of a balloon in the stomach, performed at constant aortic blood pressure and heart rate, with 0.6 l of Ringer solution (mean gastric transmural pressure of about 12 mmHg). In fourteen pigs, a decrease in splenic, renal and iliac flows and variable changes in mesenteric flow were obtained. A decrease in mesenteric flow and more marked decreases in the other flows occurred in response to the distension after the administration of propranolol or butoxamine. In five pigs, the vasoconstrictive responses were graded by step increments in gastric distending volume from 0.4 to 0.8 l. The above responses were abolished by the administration of phentolamine (eight pigs) and by bilateral cervical vagotomy (six pigs). The results showed that innocuous distension of the stomach in anaesthetized pigs reflexly caused vasoconstriction in the splenic, renal and iliac vascular beds; vasoconstriction also occurred in the mesenteric vascular bed but only after beta‐blockade. These reflex responses were mediated by sympathetic mechanisms which involved both alpha and beta vascular adrenoceptors and their afferent limb was in the vagal nerves.
The effect of exposure to increased hydrostatic pressure up to 46 ATA on respiratory sinus arrhythmia in humans
Authors: Byrne DE , Bowser‐Riley F , Cornish MJ , Kidd C
Published: December 2014

Respiratory sinus arrhythmia (RSA) has been used as a non‐invasive estimate of vagal tone to determine whether the reduction in resting heart rate commonly seen when humans are exposed to increases in the ambient pressure is associated with changes in vagal autonomic control. Two sets of divers (n = 4) were examined during two simulated saturation dives, one to 46 atmospheres absolute (ATA) and another to 37 ATA. A significant reduction in resting heart rate was seen in both dives upon exposure to high pressure compared with controls at 1 ATA. The reduction in heart rate seen at increased pressure was consistent regardless of respiratory rate, indicating that moderate changes in respiratory rate do not affect heart rate under these conditions. No changes in the overall magnitude of RSA were observed over a range of respiratory rates during either dive compared with control values at 1 ATA. During both dives, RSA was significantly larger (P < 0.05) at low respiratory rates compared with higher rates for both 1 ATA controls and at high pressure. The presence of a hyperbaric bradycardia strongly suggests that vagal tone is altered during hyperbaric exposure; however, the lack of change in the magnitude of RSA at high pressure brings into question the viability of using RSA as an accurate indicator of vagal tone and this lack of correspondence deserves further investigation.
Role of the actin cytoskeleton in the regulation of the cystic fibrosis transmembrane conductance regulator
Authors: Cantiello HF
Published: December 2014

The anion‐selective channel CFTR (cystic fibrosis transmembrane conductance regulator), whose dysfunction is responsible for the onset of cystic fibrosis, is regulated by cAMP through the activation of protein kinase A (PKA). The nature of this activation process is unknown. In the present study, patch‐clamp techniques were applied to both mouse mammary adenocarcinoma cells expressing human epithelial CFTR (CFTR cells) and cultured neonatal rat ventricular myocytes (NRVM), to determine whether CFTR is modulated by the actin cytoskeleton, and whether the actin cytoskeleton may be implicated in the cAMP‐stimulated activation of the channel protein. Acute changes in the actin cytoskeleton by addition of cytochalasin D (CD) activated whole‐cell currents in CFTR cells and NRVM. Addition of actin to excised, inside‐out patches also activated CFTR. A functional characterization of CFTR in either cell type included cAMP‐induced, linear whole‐cell and single‐channel currents in symmetrical Cl‐, permeability to ATP, and inhibition by either diphenylamine‐carboxylate (DPC) or a monoclonal antibody raised against CFTR. Incubation of CFTR cells and NRVM with CD for over 6 h prevented CFTR activation either by the cAMP pathway under whole‐cell conditions or by PKA under excised inside‐out conditions. Thus a complete derangement of the actin cytoskeleton prevents the cAMP‐dependent activation of CFTR. CFTR activation, however, was restored by subsequent addition of actin. In summary, changes in actin filament organization modulate CFTR channel activity by a mechanism entailing a direct interaction between actin filaments and CFTR.
Ca2+ signalling in cultured smooth muscle cells from human bladder
Authors: Chambers P , Neal DE , Gillespie JI
Published: December 2014

It has been suggested that the isolation and culture of human detrusor smooth muscle cells may provide useful insights into the physiology of the intact detrusor muscle. In the present paper, data are presented from cultured human bladder smooth muscle cells isolated from small, routinely available biopsies. Since the initiation of contractions involves a rise in intracellular Ca2+, this study has focused on the mechanisms involved in the rise of Ca2+ in cultured cells. Exposure of cells to bathing solutions with elevated K+ concentrations resulted in an increase in Ca2+ consistent with the presence of voltage‐activated Ca2+ channels. Agonists, including carbachol, histamine and ATP, also activated repetitive transient increases in Ca2+ in the presence and absence of external Ca2+. Spontaneous Ca2+ transients were recorded in 31% of cells isolated from normal bladders. Such spontaneous and agonist‐induced oscillations were not abolished in depolarized cells, suggesting that the mechanisms underlying the oscillations are not dependent on the cyclical operation of voltage‐operated Ca2+ channels. However, the spontaneous activity was inhibited by the Ca2+ blocker verapamil, pointing to the presence of Ca2+ channels. The operation of an IP3‐sensitive Ca2+ release mechanism was examined using saponin‐permeabilized cells, which demonstrated that IP3 increased the rate of 45Ca2+ efflux. The conclusion from this study is that many of the mechanisms described in the intact tissue are operational in cultured cells.
The deactivation kinetics of the delayed rectifier components IKr and IKs in guinea‐pig isolated ventricular myocytes
Authors: Heath BM , Terrar DA
Published: December 2014

The deactivation kinetics of the delayed rectifier potassium current (IK) were studied in guinea‐pig isolated ventricular myocytes at 35‐36 degrees C using 5 microM E4031 to selectively block the rapidly activating component (IKr) and 300 microM thiopentone to selectively suppress the slowly activating component (IKs). IK was activated by depolarization of 400 or 1000 ms duration to +40 mV, and tail currents were analysed on repolarization to potentials between ‐30 and ‐60 mV. Before exposure to drugs, the deactivation of total IK was fitted by two exponential functions at all potentials and, at ‐40 mV, the time constants were 188 +/− 12 and 2510 +/− 185 ms (n = 25); increasing the pulse duration from 400 to 1000 ms (expected to increase IKs rather than IKr under these conditions) caused a significant increase in the amplitude of only the fast component of deactivation at ‐40 mV (from 372 +/‐ 32 to 479 +/− 46 pA). The decay of IKr was biexponential at all potentials when detected as the E4031‐sensitive current and at ‐30 to ‐50 mV when detected as the thiopentone‐resistant current and was accelerated as the membrane potential was made more negative. The amplitudes of the two components of decay of IKr (as either E4031‐sensitive or thiopentone‐resistant current) were similar and neither were significantly increased when pulse duration was increased from 400 to 1000 ms. The decay of IKs detected as the E4031‐resistant current was biexponential at ‐30 mV and, in some cells, at ‐40 mV, but decayed with predominately a single, fast component of deactivation at ‐50 and ‐60 mV, which was accelerated at the more negative potentials. A slow component of decay of IKs was detected in more cells when thiopentone was used to isolate IKs, although it was of much smaller amplitude than the fast component of decay. The presence of a slow component of decay of the thiopentone‐sensitive IKs may result from a slight block of IKr by thiopentone. The amplitude of only the fast decay phase of IKs (detected using either drug) was increased when pulse duration was lengthened from 400 to 1000 ms. It seems likely that the relative importance of IKr and IKs in rate‐dependent shortening of action potential duration may need to be re‐evaluated in view of the relatively rapid deactivation of IKs, together with the prominence of the slow component of deactivation of IKr compared with that of IKs.
In vivo microperfusion of the ductuli efferentes testis of the rat: flow dependence of fluid reabsorption
Authors: Clulow J , Hansen LA , Jones RC
Published: December 2014

Individual ducts from the initial zone of the efferent ducts of the rat were microperfused in vivo using a double cannulation procedure, which allowed the recovery of perfused fluids for analysis and determination of the rate of fluid reabsorption from the perfused duct. The ducts were perfused at rates from 0.025 to 0.4 microliters min‐1 with either Krebs‐Ringer bicarbonate (KRB) solution or the native rete testis fluid (nRTF) that perfuses the ducts in situ. Reabsorption of KRB solution increased linearly with a perfusion rate of between 0.025 and 0.1 microliter min‐1 (from 17.4 +/− 1.5 to 34.3 +/− 3.2 nl (10 mm duct)‐1 min‐1), then increased no further. Reabsorption of nRTF increased linearly between 0.025 and 0.2 microliters min‐1 (from 17.7 +/− 1.5 to 61.4 +/− 13.5 nl (10 mm duct)‐1 min‐1) and then declined. The reabsorption rate from nRTF perfusates was significantly higher than from KRB perfusates. As a proportion of the luminal perfusate, reabsorption declined from 73.0 +/− 6.0 to 7.4 +/− 3.0% (10 mm duct)‐1 for KRB solution and from 73.1 +/− 6.0 to 4.1 +/− 1.3% (10 mm duct)‐1 for nRTF. There was no significant change in the concentration of either Na+ or Cl‐ in KRB solution or nRTF during perfusion through the efferent ducts, indicating that the reabsorption of these ions was isomolar. However, the reabsorption of K+ from nRTF occurred at a greater rate than that of water, and the initial [K+] declined from 17.2 +/− 0.4 mM in nRTF perfusates to 5.7 +/− 0.5 mM in collectates (perfusion rate, 0.1 microliter min‐1) to achieve equilibrium with blood plasma (4.7 +/− 0.4 mM). The osmotic pressure of both KRB and nRTF perfusates equilibrated with blood plasma, indicating a high permeability of the epithelium to water. The results of this study provide further evidence that fluid reabsorption in the efferent ducts is isosmotic, or close to isosmotic, and have shown that, as in the homologous proximal kidney tubule, reabsorption is dependent on luminal flow rate. In contrast to the proximal tubule, however, reabsorption in the efferent ducts is not maintained as a constant proportion of the perfusion load. It is concluded that microperfusion in vivo provides a useful technique for studying fluid reabsorption in the efferent ducts of the rat.
Influence of rearing temperature on lung development following methimazole treatment of postnatal lambs
Authors: Symonds ME , Andrews DC , Buss DS , Clarke L , Lomax MA
Published: December 2014

This study examines the effect of ambient temperature on lung development over the first month of postnatal life in lambs treated with a drug which inhibits thyroid hormone synthesis (methimazole, 50 mg day‐1 (kg body wt‐1)). Twin lambs were hand‐reared at a fixed level of nutrition in either a warm (WR, 25 degrees C) or cool (CR, 10‐15 degrees C) ambient temperature. Plasma thyroid hormone concentrations were significantly (P < 0.05) lower in WR lambs, but there were no differences between WR and CR groups in thyroid gland weight or hepatic iodothyronine 5'‐deiodinase activity at 8 or 29 days of age. Lung weights and the ratio of lung weight to O2 consumption were similar in WR and CR groups at 8 days but both were significantly lower (P < 0.01) in CR lambs at 29 days. Lung weight at 29 days was significantly (P = 0.002) correlated with plasma triiodothyronine concentration in CR but not WR lambs, and laryngeal braking of expiratory airflow was observed in three of seven CR lambs but not in WR lambs. Four WR, but no CR, lambs exhibited a rapid deterioration in health or died unexpectedly between 14 and 21 days of age. These WR lambs were characterized by having small lungs and slower growth rates. It is concluded that in WR lambs chronically treated with methimazole, both lung development and the ability of an individual to adapt effectively to methimazole treatment are compromised. In CR lambs, although lung growth may be reduced this does not appear to compromise O2 consumption or health.
Unaltered oxygen uptake kinetics at exercise onset with lower‐body positive pressure in humans
Authors: Williamson JW , Raven PB , Whipp BJ
Published: December 2014

The purpose of this study was to determine the influence of a reduced skeletal muscle blood flow on oxygen uptake (VO2) kinetics at the onset of cycle ergometer exercise. Seven healthy subjects performed rest‐to‐exercise transitions with a lower‐body positive pressure (LBPP) of 45 Torr. Two work rates were selected for each subject: a moderate intensity (VO2, approximately 1.9 l min‐1; delta[lactate], approximately 1 mequiv l‐1) below the estimated lactate threshold and a heavy intensity (VO2, approximately 2.6 l min‐1; delta[lactate], approximately 3 mequiv l‐1) above this threshold. Pulmonary gas exchange variables and ventilatory (VE) responses were computed breath‐by‐breath from mass spectrometer and turbine volume meter signals, respectively, and mean response times (MRT) calculated. Samples of 'arterialized' venous blood were used for the determination of [lactate], pH and [K+]. While the application of 45 Torr LBPP had no effects on VO2 kinetics during moderate exercise (MRT: 33.5 +/− 1.2 s at 45 Torr vs. 32.8 +/− 1.3 s at 0 Torr; P > 0.05) or on [lactate], pH or [K+], breathing frequency (f) was increased (P < 0.05) and tidal volume (VT) reduced (P < 0.05). The addition of LBPP during heavy exercise did not alter VO2 kinetics (MRT: 35.2 +/− 1.5 s at 45 Torr vs. 34.8 +/− 1.5 s at 0 Torr; P > 0.05), or [lactate], pH or [K+]. Although both the VE (via an increased f) and CO2 output (VCO2) were significantly greater with LBPP by approximately 30 l min‐1 and approximately 500 ml min‐1, respectively, end‐tidal CO2 partial pressure was decreasing, suggesting an additional ventilatory stimulus. These data can be interpreted to suggest that oxygen delivery is not critically dependent upon blood flow to the working muscle at exercise onset, while LBPP‐induced increases in VE during suprathreshold exercise may be related to an accumulation of metabolites at the working muscle or the effects of pressure per se.
5‐Hydroxytryptamine induces forestomach hypomotility in sheep through 5‐HT4 receptors
Authors: Plaza MA , Arruebo MP , Murillo MD
Published: December 2014

The effects evoked by 5‐hydroxytryptamine (5‐HT; serotonin) on forestomach myoelectric activity were investigated in conscious sheep. Myoelectric signals were recorded with electrodes chronically implanted in the reticulum, rumen (dorsal sac) and omasal body, and were analysed by a computer‐based method. The 5‐HT receptors and the neuronal pathways involved in these actions were studied. The intravenous (i.v.) infusion of 5‐HT (8 micrograms kg‐1 min‐1 for 5 min) evoked an inhibition of activity of the whole forestomach. Methiothepin, injected i.v. at 0.1 mg kg‐1, inhibited rumen secondary contractions and omasum activity. However, forestomach activity remained unchanged after the administration of 0.2 mg kg‐1 of ketanserin, ondansetron, tropisetron, GR‐113808, phentolamine, propranolol, domperidone and naloxone. Atropine (0.2 mg kg‐1), hexamethonium (2 mg kg‐1) or haloperidol (0.1 mg kg‐1) abolished rumen secondary cycles and inhibited omasum activity. In addition, atropine also suppressed primary cycles. GR‐113808 blocked all 5‐HT‐induced effects. Furthermore, atropine or hexamethonium prevented the 5‐HT‐evoked inhibition of reticulorumen primary cycles. In contrast, the remaining antagonists did not alter the 5‐HT‐evoked forestomach hypomotility. In conclusion, 5‐HT induces inhibition of forestomach myoelectric activity through 5‐HT4 receptors, these actions being mediated by cholinergic neural pathways involving muscarinic and nicotinic receptors. However, adrenergic, dopaminergic or opiate pathways are not implicated.
Blood and milk prolactin and the rate of milk synthesis in women
Authors: Cox DB , Owens RA , Hartmann PE
Published: December 2014

In women, the concentration of prolactin in the plasma increases in response to nipple stimulation. This response has led to the assumption that prolactin influences the rate of milk synthesis. To investigate this hypothesis we have measured 24 h milk production, the short‐term (between breastfeeds) rates of milk synthesis and the concentration of prolactin in the blood and breastmilk, from 1 to 6 months of lactation in eleven women. Over the long term, the 24 h milk production remained constant (means +/− S.E.M.): 708 +/− 54.7 g/24 h (n = 11) and 742 +/− 79.4 g/24 h (n = 9) at 1 and 6 months, respectively. The average short‐term rate of milk synthesis (calculated from the increase in breast volume between breastfeeds; means +/− S.E.M.) did not change: 23 +/− 3.5 ml/h (n = 23) and 23 +/− 3.4 ml/h (n = 21) at 1 and 6 months, respectively. However, significant variation in the short‐term rate of milk synthesis (from < 5.8 to 90 ml/h) was found both between breasts, measured concurrently (coefficient of variation, c.v. = 72%), and within the same breast, measured over consecutive breastfeeds (c.v. = 85%). The basal and suckling‐stimulated concentrations of prolactin in the plasma (means +/− S.E.M.) declined from 1 to 6 months (basal, from 119 +/− 93 to 59 +/− 29 micrograms/1; peak, from 286 +/− 109 to 91 +/− 44 micrograms/l). In contrast, the concentration of prolactin in milk was much lower than in plasma, and decreased only slightly from 1 to 6 months of lactation (fore‐milk, from 26.4 +/− 10 to 23.3 +/− 9.8 micrograms/l; hind‐milk, from 18.9 +/− 5.1 to 13.2 +/− 6.3 micrograms/l). The concentration of prolactin in the milk was related to the degree of fullness of the breast, such that the concentration was highest when the breast was full. We found no relationship between the concentration of prolactin in the plasma and the rate of milk synthesis in either the short or long term. However, the relationship between the concentration of prolactin in milk and the degree of fullness of the breast suggests that the internalization of prolactin, after binding to its receptor, may be restricted when the alveolus is distended with milk.
Nitric oxide and peroxynitrite released by ultraviolet B‐irradiated human endothelial cells are possibly involved in skin erythema and inflammation
Authors: Deliconstantinos G , Villiotou V , Stavrides JC
Published: December 2014

In this study we attempted to demonstrate whether endothelial cell nitric oxide synthase (eNOS) and xanthine oxidase (XO) could be activated to release nitric oxide (NO) and peroxynitrite (ONOO‐) following exposure to ultraviolet B (UVB) radiation and to define whether this light‐induced response could be involved in the pathogenesis of sunburn erythema and inflammation. Treatment of human endothelial cells with UVB (290‐320 nm) radiation (up to 100 mJ/cm2) resulted in an increase of both NO and ONOO‐ release that was inhibited by NG‐monomethyl‐L‐arginine (L‐NMMA). Treatment of cell cytosol with various doses of UVB radiation (up to 20 mJ/cm2) resulted in a threefold increase of XO activity that was inhibited (approximately 90% by oxypurinol. In reconstitution experiments, when purified eNOS was added to purified XO, an almost fourfold increase in ONOO‐ production at 20 mj/cm2 UVB radiation was observed. UVB radiation (100 mg/cm2) decreased cell membrane fluidity, indicating changes in the physicochemical characteristics of the membranes. In in vivo experiments, when human volunteers were subjected to UVB light, a protection factor (PF) of 3.90 +/− 0.85 was calculated when an emulsified cream formulation containing nitro‐L‐arginine (L‐NA; 2%) and L‐NMMA (2%) was applied to their skin. The present studies indicate that UVB radiation acts as a potent stimulator of eNOS and XO in human endothelial cells. The cytotoxic effects of NO and ONOO‐ may be the main factors in the integrated response of the skin leading to vasodilatation, the first key event of erythema production and the inflammation process.
Activation of an anion conductance and beta‐cell depolarization during hypotonically induced insulin release
Authors: Best L , Miley HE , Yates AP
Published: December 2014

The perforated patch technique was used to study the effects of hypotonic extracellular solutions on membrane potential and whole‐cell currents in intact rat pancreatic beta‐cells. A 30% reduction in osmolarity resulted in activation of an outwardly rectifying CI(‐)‐selective conductance in rat beta‐cells. This conductance was inhibited by the anion channel blocker 4,4'‐diisothiocyanatostilbene‐2,2'‐disulphonic acid (DIDS). Exposure to a hypotonic medium also led to a transient stimulation of electrical activity accompanied by cell swelling and a gradual return towards control volume. These effects were also associated with the generation of an inward current at a holding potential of ‐70mV, and a stimulation of insulin release from intact perifused islets. All of the above effects were inhibited by DIDS. It is suggested that the stimulation of insulin release by hypotonic solutions results from activation of a volume‐sensitive anion conductance generating an inward current and leading to a subsequent depolarization of the beta‐cell.
Unilateral cooling in the region of locus coeruleus blocks the fall in respiratory output during hypoxia in anaesthetized neonatal sheep
Authors: Moore PJ , Ackland GL , Hanson MA
Published: December 2014

In nineteen alpha‐chloralose‐anaesthetized 3‐ to 8‐day‐old lambs, the effect of cooling focally in the region of the locus coeruleus on respiratory output, as measured from phrenic nerve activity, was examined during normoxia and isocapnic hypoxia, and during infusion of adenosine or saline. Eleven lambs showed a ‘biphasic’ respiratory response to hypoxia, with an initial increase in phrenic nerve activity (phase 1) being followed by a decline to prehypoxic levels (phase 2). Cooling to 26‐34 degrees C unilaterally in the locus coeruleus reversed the fall in respiratory output. When the probe temperature was returned to body temperature (39 degrees C) respiratory output fell again to phase 2 levels. Cooling had no effect during normoxia. In these lambs, adenosine infusion (0.3 mg kg‐1 min‐1 intra‐arterially) during normoxia also decreased respiratory output. During adenosine infusion, cooling again increased respiratory output. Saline infusion did not affect respiratory output during normoxia with or without cooling. The other lambs did not show a biphasic respiratory response, i.e. increased respiratory output was maintained during isocapnic hypoxia. In these lambs adenosine had no effect on respiratory output. Cooling/rewarming the probe did not affect respiratory output during normoxia or hypoxia, or during adenosine or saline infusion. These results show that focal cooling in the dorsal pons at the locus coeruleus removes the inhibition of respiratory output seen in the neonatal biphasic respiratory response to hypoxia, but does not affect breathing in normoxia. This suggests that an important component of the biphasic respiratory response is mediated by the activation of neurones inhibitory to respiratory output, and involves either axons of passage or cell bodies lying in the locus coeruleus. Furthermore, the inhibitory effects of adenosine on breathing in the neonate appear to be mediated via this site.
Effect of diets varying in nitrogen or phosphorus content on indicators of bone growth in lambs
Authors: Scott D , Loveridge N , Nicodemo L , Buchan W , Milne J , Duncan A , Nicol P , Robins SP
Published: December 2014

Growing lambs were fed diets low in nitrogen and phosphorus (LNLP), low in nitrogen and high in phosphorus (LNHP), high in nitrogen and low in phosphorus (HNLP) or high in nitrogen and phosphorus (HNHP) and the effects on bone growth and on blood and urinary bone marker levels or excretion rates were monitored. Plasma calcium concentrations were higher, and phosphorus concentrations lower, in lambs fed the low phosphorus diets but there were no differences in plasma 1,25‐dihydroxyvitamin D3 (1,25(OH)2D3) concentrations. Lambs fed both low phosphorus diets (LNLP and HNLP) had lower plasma osteocalcin and higher bone‐specific alkaline phosphatase concentrations than those fed the high phosphorus diets. Urinary pyridinoline and deoxypyridinoline excretion were also affected by treatment, with their rates of excretion being highest in lambs fed the diet low in both nitrogen and phosphorus (LNLP). Lambs fed the low phosphorus diets were lighter in weight at slaughter and had lighter bones that were less well mineralized than those fed the high phosphorus diets. Reducing the nitrogen content of the diet appeared to have little effect on bone composition. These results suggest that bone markers that have proved useful in the diagnosis and treatment of bone disease are sensitive to variation in nutrient supply and may prove useful in early detection of nutrient deficiencies that affect bone growth.
The control of blood flow to the placenta
Authors: Poston L
Published: December 2014

The maintenance of adequate blood flow to the placenta is essential for the successful outcome of pregnancy. The placental vascular bed is often regarded as a low‐resistance circulation in which blood flow is determined by the fetal cardiac output, but in pregnancies associated with growth retardation, and for reasons unknown, vascular resistance increases and blood flow is compromised. This review concentrates on recent advances in our understanding of the factors that may influence fetoplacental blood flow, with particular emphasis upon the circulation of the human placenta. The placenta has no neuronal input and vascular resistance is entirely determined by humoral and structural factors. In the absence of catecholamine sensitivity, constrictor prostanoids and endothelin may be important constrictor agonists, whereas nitric oxide is now considered to contribute to tonic reduction in vascular resistance. Little is known of the relative contributions of these vasoactive agents in the elevation of fetoplacental vascular resistance, but recent evidence points to an important role for altered vascular structure in growth‐retarded pregnancies.
Dysfunction of the endothelial nitric oxide signalling pathway in diabetes and hyperglycaemia
Authors: Sobrevia L , Mann GE
Published: December 2014

1. Vasodilatation induced by adenosine is mediated through the activation of A2‐purinoceptors in endothelial and smooth muscle cells. 2. Adenosine induces a rapid and transient membrane hyperpolarization in endothelial cells. 3. Acute exposure to adenosine or A2‐purinoceptor agonists activates the L‐arginine‐NO signalling pathway in human endothelial cells in vitro.
Synchronization of human finger movements: delays and sex differences with isotonic ‘antiphase’ motion
Authors: Walsh EG
Published: December 2014

The interval between the operation of two Morse keys, or the moments at which fingers touched or broke contact with electrical conductors, has been measured, the subjects making the movements as synchronously as possible. The data obtained deal with timings when two fingers flexed or extended together (‘in phase’) and when one finger extended whilst the other flexed (‘antiphase’). For antiphase movements of the index fingers with the conductors, flexion usually occurred before extension so there was ‘overlap’. Statistically, this period was greater in women and unusually high values were found in one subject with mild cerebral palsy. Overlap was not found using the keys. Using conductors, the timing of antiphase movements for the index and middle fingers of the right and left hand showed overlap again. No significant relationships were found with handedness. When the conductors were placed above the fingers, and the contact was thus with the dorsum, extension took place before flexion: both fingers were extended for a while, with overlap occurring again. The significance of these findings for the understanding of motor control is discussed.
Task‐dependent effect of limb immobilization on the fatigability of the elbow flexor muscles in humans
Authors: Yue GH , Bilodeau M , Hardy PA , Enoka RM
Published: December 2014

Because short‐term limb immobilization produces selective adaptations in the neuromuscular system that probably interact with the task‐dependent expression of muscle fatigue, the purpose of this study was to determine the effects of limb immobilization on the ability of human subjects to sustain isometric contractions at low and moderate submaximal forces. Four weeks of elbow joint immobilization caused a substantial decrease in the daily activity of biceps brachii during immobilization, a significant reduction in the cross‐sectional area and volume of the elbow flexor muscles as measured by magnetic resonance imaging, and a decline in the maximum voluntary contraction (MVC) activation and force of the elbow flexor muscles. Immobilization had a task‐dependent effect on muscle fatigue with a substantially increased endurance time (reduced fatigability) at a low force (20% MVC) and no statistical effect at a moderate force (65% MVC). Despite atrophy of the elbow flexor muscles due to the immobilization, the twitch force elicited in biceps brachii by electrical stimulation was greater after immobilization. The selective improvement of fatigue resistance for the low‐force contraction and the absence of a change in the time course of the twitch suggests that the immobilization‐induced adaptations included an improved efficacy of some excitation‐contraction processes and underscored the major role of these mechanisms in determining the endurance time for low‐force, long‐duration contractions.
Ca2+ mobilization in cultured human bladder smooth muscle cells in response to hypotonic stimuli
Authors: Chambers P , Neal DE , Gillespie JI
Published: December 2014

The effects of hypotonic solutions on intracellular Ca2+ were determined on primary cultures of human detrusor smooth muscle cells. In the presence of external Ca2+, changing from a solution containing 290 mosmol l‐1 to one containing 240 mosmol l‐1 (80% normal tonicity) activated a rise in intracellular Ca2+. If the exposure to 240 mosmol l‐1 was followed by exposure to 150 mosmol l‐1 (50% normal tonicity) a further increase in Ca2+ was noted. Exposure to hypotonic solutions which contained nominally zero Ca2+ also resulted in a rise in intracellular Ca2+, suggesting that mechanical perturbation of the surface membrane can mobilize Ca2+ directly from intracellular stores. Stimulation of cells with a brief exposure to agonist (histamine, 5 microM) in the nominal absence of external Ca2+ produced a release of Ca2+ from intracellular stores. Under these conditions the response to a subsequent hypotonic stimulation was small or absent. These observations suggest that hypotonic stress induces a mobilization of Ca2+ from the same store as that activated by agonist. Detailed analysis of data from individual bladders revealed that there were significant differences between cells cultured from normal bladders and those from bladders with detrusor instability. On exposure to 240 mosmol l‐1 cells from unstable bladders gave larger responses than cells from normal bladders. However, exposure of cells from unstable bladders to a subsequent stimulus of 150 mosmol l‐1 resulted in a smaller rise in Ca2+ than was observed with normal cells. Overall, these data suggest that isolated detrusor smooth muscle cells are sensitive to membrane stretch by hyposmotic solutions and that the mechanisms involved include the direct mobilization of intracellular Ca2+. The data also suggest that there may be differences in the responsiveness of cells isolated from stable and unstable bladders.
Pancreatic endocrine responses to the peptides VIP and PACAP in the conscious calf
Authors: Edwards AV , Bloom SR , Ghatei MA
Published: December 2014

Pancreatic endocrine responses to vasoactive intestinal peptide (VIP), pituitary adenylate cyclase‐activating peptide (PACAP) and acetylcholine have been investigated in conscious calves, while continuously monitoring aortic blood pressure and heart rate, in order to assess the effects of these neuropeptides under normal physiological conditions. VIP was found to modulate the release of pancreatic polypeptide (PP) in response to acetylcholine at a dose which had no significant effect on the release of either pancreatic glucagon or insulin. PACAP stimulated the release of insulin and the effect was potentiated during infusions of exogenous glucose. PACAP, when infused together with acetylcholine, also stimulated the release of pancreatic glucagon in the absence of exogenous glucose but not in its presence. Release of PP was stimulated during infusions of PACAP, whether or not exogenous glucose was given, and this effect was found to be additive with that of acetylcholine.
Electrophysiological characteristics of submucosal neurones in the proximal colon of guinea‐pigs: comparisons with caecum and descending colon
Authors: Cunningham SM , Hirai K , Mihara S , Lees GM
Published: December 2014

A systematic examination has been made of the active and passive electrophysiological properties and synaptic inputs of forty‐four randomly impaled submucosal neurones in the proximal colon of the guinea‐pig to compare these characteristics directly with those of submucosal neurones in the caecum (n = 70) and descending colon (n = 45). Within each of the three electrophysiological classes of submucosal neurones identified (S, S/AH and AH), no statistically significant regional differences were found with respect to the resting membrane potential, membrane time constant or input resistance between neurones of the proximal colon, descending colon and caecum. Of submucosal neurones from the proximal colon, forty‐three of forty‐four (98%) received fast excitatory synaptic potentials (fast EPSPs); thirty‐nine (91%) were S neurones and the others were S/AH neurones; only one of the forty‐four cells (2%) was an AH neurone. An idazoxan‐sensitive slow inhibitory postsynaptic potential (slow IPSP) was induced in thirty of forty‐three S and S/AH neurones (70%) of the proximal colon, compared with sixty‐one of sixty‐six caecal neurones (92%) and twelve of forty‐one neurones (29%) in the descending colon. The mean (+/‐ S.E.M.) amplitude of the slow IPSP in proximal colonic neurones was 17 +/ࢤ 1 mV (range, 6‐30 mV; n = 30), compared with the significantly larger synaptic response (25 +/ࢤ 1 mV; range, 7‐38 mV; n = 66; P < 0.05) recorded in the caecum; the mean slow IPSP amplitude in the descending colon was significantly smaller (12 +/ࢤ 2 mV; range, 5‐27 mV; n = 12; P < 0.05) than that in the caecum. In the proximal colon and caecum, only those neurones with a slow IPSP had a hyperpolarizing response to noradrenaline, whereas about 50% of those neurons of the descending colon that lacked a slow IPSP were hyperpolarized by noradrenaline, acting via alpha 2‐adrenoceptors. Thus, the electrophysiological characteristics of the submucosal neurones of the proximal colon more closely resemble those of the caecum than those of the descending colon, of which many do not have a functional noradrenergic synaptic input. Furthermore, the results confirm that there are fundamental regional differences in the guinea‐pig large intestine with respect to the synaptic organization of submucosal neurones of particular electrophysiological classes.
A quantitative description of dynamic left ventricular geometry in anaesthetized rats using magnetic resonance imaging
Authors: Crowley JJ , Huang CL , Gates AR , Basu A , Shapiro LM , Carpenter TA , Hall LD
Published: December 2014

We report a functional application of magnetic resonance imaging (MRI) for the quantitative description of left ventricular geometry through systole and diastole in normal anaesthetized Wistar rats that might be applicable for the analysis of chronic changes resulting from pathological conditions. Images of cardiac anatomy were acquired through planes both parallel and perpendicular to the principal cardiac axis at times that were synchronized to the R wave of the electrocardiogram. The images of the transverse sections were assembled into three‐dimensional representations of left ventricular geometry at consecutive time points through the cardiac cycle. This confirmed the geometrical coherence of the data sets, that each slice showed circular symmetry, and that the images were correctly aligned with the appropriate anatomical axes. Different models for the three‐dimensional geometry of the left ventricle were then tested against the epi‐ and endocardial surfaces reconstructed from images of the transverse sections of the left ventricle in both systole and diastole using least‐squares minimizations in three dimensions. In agreement with previous reports in the human heart, an elliptical figure of revolution offered an optimal fit to the epicardial and endocardial geometry for the rat heart in diastole. This was in preference to models that used spherical, quartic or parabolic geometries. However, in contrast to contraction in the human heart, all these geometrical representations broke down during systolic ejection in the rat heart. We therefore introduced a more general hybrid model which described left ventricular geometry in terms of the variation of the radii r(z), independently determined for each slice, with its position z along the principal cardiac axis. The resulting function r(z) could then be described by a simple ellipsoid of revolution not only during diastole, but also throughout ventricular ejection. The findings also ruled out alternative geometrical representations. It was then possible additionally to reconstruct the luminal and total left ventricular volumes, wall thicknesses and ejection fractions through the cardiac cycle and to confirm that the predicted total ventricular wall volume was conserved throughout the cardiac cycle. Our hybrid model of cardiac geometry may thus be useful for non‐invasive serial studies of chronic pathological changes that use the rat as a model experimental system.
Programmed cell death in bovine mammary tissue during lactation and involution
Authors: Wilde CJ , Addey CV , Li P , Fernig DG
Published: December 2014

Cessation of milk removal causes mammary tissue involution, which in rodents is characterized by extensive tissue degeneration and loss of the majority of luminal epithelial cells by apoptosis. In contrast, bovine mammary tissue shows little histological evidence of tissue remodelling between lactations. In this study, we combined histology with molecular biology to examine the cellular and molecular changes in bovine mammary tissue on cessation of milking. Oligonucleosomal laddering of genomic DNA extracted from lactating tissue indicated that a proportion of cells were dying by apoptosis. This was confirmed by terminal deoxynucleotide transferase‐mediated deoxyuridine nick end‐labelling of apoptotic cells in lactating tissue sections (TUNEL). One week after cessation of milking, alpha‐lactalbumin and alpha S1‐casein messenger RNA (mRNA) abundance had decreased by 99 and 85%, respectively, whereas lactoferrin mRNA had increased 20‐fold. Drying off was also accompanied by an increase in oligonucleosomal laddering of genomic DNA, and by an increase in the proportion of TUNEL‐positive cells, which were localized preferentially in regions where alveolar structure had deteriorated. Therefore, termination of lactation was associated with partial loss of the mammary cell population and dedifferentiation of the remainder.
A new protocol for the measurement of picomole quantities of magnesium in rat renal tubular fluid
Authors: Kibble JD , Audsley N , Day JP , Green R
Published: December 2014

The analysis of picomolar quantities of magnesium by electrothermal atomic absorption spectrophotometry (EAAS) was studied using a Perkin‐Elmer‐Zeeman 3030 spectrophotometer. The absorbance signal was not heavily dependent on the atomization temperature, but was greatly reduced when ashing temperatures in excess of 1200 degrees C were applied. The magnesium signal was significantly depressed in the presence of excess chloride in the sample matrix. However, use of NH4NO3 as a matrix modifier was sufficient to overcome this artefact. The analytical sensitivity was 0.15 absorbance units pmol‐1 and the detection limit was 0.04 pmol. Using nanolitre constriction pipettes to dispense standards, the mean coefficient of variation was 5%. Measurement of magnesium handling in the rat proximal convoluted tubule revealed a significant correlation between the tubular fluid‐to‐plasma ultrafiltrate (TF/UF) concentration ratio for magnesium and the tubular fluid‐to‐plasma (TF/P) concentration ratio for [3H]inulin (r2 = 0.56, n = 17). This indicated that magnesium is concentrated during its passage along the proximal tubule. In contrast, this was not the case for sodium (r2 = 0.11, n = 16). Mean (TF/UF)Mg (1.16 +/− 0.07, n = 17) for random punctures was significantly greater than that for sodium ((TF/UF)Na = 1.02 +/− 0.02, n = 16). Despite concentration of magnesium in the lumen, significant net reabsorption of magnesium was observed along the length of the tubule (fractional reabsorption, FRMg = 19.4 +/− 3.0%, n = 17). In conclusion, EAAS provides a highly sensitive, reproducible and technically simple method for measuring picomolar quantities of magnesium in renal tubular fluid.
The influence of stimulation frequency on force‐velocity characteristics of in situ rat medial gastrocnemius muscle
Authors: de Haan A
Published: December 2014

Force (and power)‐velocity characteristics were determined at different stimulation frequencies in th situ rat muscle with nerve stimulation at 36 degrees C. In isometric contractions (duration, 150 ms), maximal force is generated at approximately 120 Hz. In contrast, in the high velocity (250 mm s‐1) shortening contractions, frequencies of approximately 400 Hz were needed to obtain maximal dynamic force, while 120 Hz elicited only approximately 26% of the maximum. At the highest velocity measured, power production was significantly different (P < 0.05) among frequencies of 80, 120, 200 and 400 Hz, suggesting that maximal shortening velocity should be assessed using very high stimulation frequencies. However, the results further indicate that lower frequencies may be adequate in exercise studies that investigate fatigue and changes in power output during series of repetitive contractions.
Calcium‐activated transient membrane currents are carried mainly by chloride ions in isolated atrial, ventricular and Purkinje cells of rabbit heart
Authors: Szigeti G , Rusznak Z , Kovacs L , Papp Z
Published: December 2014

Under physiological conditions, calcium‐dependent ([Ca2+]i‐dependent) Cl‐ currents (ICl(Ca)) have been suggested to participate in the repolarizing processes. In this paper, the possible contribution of ICl(Ca) to transient inward currents and, hence to arrhythmias, has been studied in myocytes from the working myocardium and from the conductive system. Single atrial, ventricular and Purkinje cells, isolated enzymatically from rabbit heart, have been studied under whole‐cell voltage‐clamp and were internally perfused with the fluorescent Ca2+ indicator, fura‐2 (100 microM). Ca2+ release from the sarcoplasmic reticulum was either induced by external application of caffeine or occurred spontaneously in Ca(2+)‐overloaded cells. Membrane currents accompanying Ca2+ transients showed linear current‐voltage characteristics between ‐60 and +80 mV as evidenced from fast voltage ramps. When intra‐ and extracellular Cl‐ concentrations were kept symmetrical in the absence of the Na(+)‐Ca2+ exchange mechanism, transient currents had a reversal potential close to 0 mV. Reduction of external Cl‐ concentration shifted this reversal potential towards the new Cl‐ equilibrium potential. Neither the time course of the transient currents nor the shift in their reversal potentials was significantly affected by the presence of Na+. Approximately 90% of this current was blocked by the application of the Cl‐ channel blocker, anthracene‐9‐carboxylic acid (0.5 mM) at +80 mV. It is concluded, that [Ca2+]i‐activated transient membrane currents in atrial, ventricular and Purkinje cells of rabbit heart are mainly due to the activation of a [Ca2+]i‐dependent Cl‐ current.