Neurones located in cervical segments (C6/C7) of the spinal cord were investigated electrophysiologically in cats deeply anaesthetized with α-chloralose. Extracellular recordings of antidromic action potentials were performed in order to establish whether long descending propriospinal neurones projecting to sacral segments could have collateral axonal branches ascending to supraspinal centres. The effects of stimulation of the lateral reticular nucleus (LRN) and the inferior cerebellar peduncle (restiform body, RB), as well as the thirteenth thoracic (Th13) and sacral (S1/S2) segments of the spinal cord were tested in 93 cells. Two main groups of cells were identified: 54 % of the total sample were classified as purely propriospinal and 46 % as bidirectional neurones. Various patterns of projections, as well as the ipsi-, contra- or bilateral courses of axons in the lateral funiculi of the spinal cord, enabled several types of neurones to be distinguished within the above groups. Comparison between particular types showed no significant difference with respect to location in the grey matter (predominantly Rexed's laminae VII-VIII) and the conduction velocities of descending axons. However, the mean axonal conduction velocities of branches ascending to LRN and/or RB were significantly lower in comparison to those measured for spinal collaterals. The hypothetical function of the neurones examined is discussed. Since the same information can be conveyed simultaneously by these branching neurones to lower spinal segments and supraspinal centres, an integrative role in the system of motor control is suggested.

This study seeks to confirm the progressive changes in cardiac output and heart rate previously reported with 8 h exposures to constant hypoxia, and to examine the role of sympathetic mechanisms in generating these changes. Responses of ten subjects to four 8 h protocols were compared: (1) air breathing with placebo; (2) isocapnic hypoxia (end-tidal PO2 = 50 mmHg) with placebo; (3) isocapnic hypoxia with beta-blockade; and (4) air breathing with beta -blockade. Regular measurements of heart rate and cardiac output (using ultrasonography and N2O rebreathing techniques) were made with subjects seated in the upright position. The sensitivity of heart rate to rapid variations in hypoxia (GHR) and heart rate in the absence of hypoxia were measured at times 0, 4 and 8 h. No significant progressive effect of hypoxia on cardiac output was detected. There was a gradual rise in heart rate with hypoxia of 11 ± 2 beats min-1 in the placebo protocol and of 10 ± 2 beats min-1 in the beta-blockade protocol over 8 h, compared to the air breathing protocols. The rise in heart rate was progressive (P < 0.001) and accompanied by progressive increases in both GHR (P < 0.001) and heart rate measured in the absence of hypoxia (P < 0.05). No significant effect of beta-blockade was detected on any of these progressive changes. We conclude that sympathetic mechanisms that act via beta -receptors play little role in the progressive changes in heart rate observed over 8 h of moderate hypoxia. Experimental Physiology (2000) 85.5, 557-565.

It is not resolved whether the anionic channel involved in volume regulation after cell swelling comprises one or more subunits. Moreover, it remains to be determined which of the different proteins cloned so far, for which an involvement in cell volume regulation has been postulated, is the ideal candidate. In this review, we consider the role of the ICln protein, cloned from MDCK cells, in cell volume regulation.

This study was performed to determine whether the absorption of liquid from the lungs of postnatal sheep is dependent on pulmonary perfusion pressure, blood gases or blood flow. Relationships between perfusion pressure, rate of lung liquid absorption and perfusate PO2, PCO2 and pH were examined by linear regression analysis from in situ perfused lungs from sheep aged 6 weeks to 6 months. The airspaces of the lungs were filled with liquid containing an impermeant tracer, to allow measurement of the rate of liquid absorption. There was no significant relationship between the rate of lung liquid absorption and pulmonary blood flow (n = 36, r = -0.01, P > 0.1), pulmonary perfusion pressure (n = 36, r = 0.28, P > 0.05) or perfusate PO2, PCO2 or pH. No significant relationships were found between pulmonary blood flow and perfusate PO2, PCO2 or pH. There was no evidence to suggest that the absorption of liquid from the lungs of postnatal sheep is dependent on pulmonary blood flow, blood gases or perfusion pressure, within the limits studied, indicating that lung liquid absorption is dependent on the pulmonary epithelium and not on the pulmonary vasculature. The findings that lung liquid absorption continues in hypoxic environments and despite severe reductions in blood flow may be relevant to the field of transplant surgery. Experimental Physiology (2001) 86.3, 391-398.

Diabetes mellitus and alcohol (ethanol) intake are two positively correlated major risk factors for cardiovascular abnormalities. However, the interaction of the two on cardiac function is largely unknown. The purpose of the present study was to examine the impact of genetically predisposed diabetes on acute ethanol exposure-induced cardiac contractile depression at the myocyte level. Ventricular myocytes from spontaneously biobreeding diabetes-prone (BBDP) rats and their diabetes-resistant littermates (BBDR) were stimulated to contract at 0.5 Hz. Contractile properties analysed include: peak shortening amplitude (PS), time-to-PS (TPS), time-to-90 % relengthening (TR90) and maximal velocities of shortening/relengthening (± dL/dt). BBDP rats displayed hyperglycaemia, reduced body weight gain and increased cardiac, hepatic and renal size. Myocytes isolated from BBDP rat hearts exhibited prolonged TPS and TR90 associated with normal PS and ± dL/dt, compared with myocytes from the BBDR group. Acute ethanol exposure (80-640 mg dl-1) caused a concentration-dependent inhibition of PS in both BBDR and BBDP myocytes. However, the degree of inhibition of PS was significantly reduced in BBDP myocytes compared to that of BBDR myocytes. The maximal inhibition was 52.9 % and 28.4 % in BBDR and BBDP groups, respectively. Ethanol significantly depressed ± dL/dt in both BBDR and BBDP myocytes. In addition, ethanol did not affect TPS or TR90 in either the BBDR or BBDP group. Collectively, these results suggest that the ethanol-induced depression in cardiac myocyte contraction may be 'shadowed' by genetically predisposed diabetes. Experimental Physiology (2002) 87.3, 293-298.

Acetate deriving from microbial fermentation may occur at considerable concentrations in the distal small intestine, where it appears to be absorbed by two different mechanisms: acetate-HCO3- exchange and non-ionic diffusion. Whether acetate affects absorption of other nutrients at this intestinal site is not known. Therefore the influence of acetate (30 mmol l-1) on oligopeptide absorption was studied using an in vitro mucosal uptake technique allowing measurement of substrate uptake across the brush border membrane (BBM). Acetate significantly inhibited mucosal uptake of 14C-labelled glycylsarcosine (Gly-Sar) at pH 6 and pH 7 in the presence of sodium. No inhibition occurred in the absence of sodium. Both acetate and the absence of sodium decreased Vmax of mucosal Gly-Sar uptake without substantially affecting the apparent Km value. Since it is well established that Vmax of peptide transport across the intestinal BBM depends on the size of the transmembrane H+ gradient as a driving force the present findings are in accordance with the assumption that acetate inhibits peptide absorption by attenuating the H+ gradient across the BBM, which depends on the presence of sodium.

We discovered in 1987 that the shorter form of apolipoprotein B (B48) synthesized in the intestine is due to the action, previously unrecognized in mammalian cells, of an mRNA-editing process, and more recently we demonstrated that this was due to a specific enzyme (APOBEC-1) with cytidine deaminase activity. We show here, by sequence alignment, molecular modelling and mutagenesis, that APOBEC-1 is a cytidine deaminase, responsible for editing apoB mRNA, and that is related in crystal structure to the cytidine deaminase of Escherichia coli (ECCDA). The two enzymes are both homodimers with composite active sites formed with loops from each monomer. In the sequence of APOBEC-1, three gaps compared with ECCDA match the size and contour of the minimal RNA substrate. We propose a model in which the asymmetric binding of one active site to the substrate cytidine which is positioned by the downstream binding of the product uridine and that this helps to target the other active site for deamination.

Subjects sat on a 'Balans' chair supported by the shaft of a large torque generator. The lower but not the upper part of the body was free to make to/fro movements maintained by feeding back a modified velocity signal to the generator. The stiffness could be increased by the use of a position signal. Using the equations for a torsion pendulum the good linearity observed in relationship to the applied forces allowed stiffness, inertia, damping, critical damping and the damping factor to be determined in absolute terms. The method enables parameters relevant to back function and problems to be evaluated. Eleven adults acted as subjects. Experimental Physiology (2003) 88.3, 305-308.

Lung liquid (LL) is secreted into the fetal lung lumen, but it must be rapidly absorbed at birth to allow air breathing. In vitro studies have implicated oxygen as a possible factor causing the switch from secretion to absorption of lung liquid at birth. We developed a technique of oxygenating the fetal lung using liquid ventilation with haemoglobin (Hb) solutions in chronically catheterized fetal lambs (129-140 days gestation; term, 147 days). In some experiments 2,3-diphosphoglycerate (DPG) was added to increase oxygen delivery. LL secretion rate (Jv) was measured using an indicator dilution method. Eighteen fetuses were divided into four groups and ventilated with liquid under the following conditions: (i) Hb with oxygen, (ii) Hb without oxygen, (iii) Hb with DPG and oxygen and (iv) Hb with DPG without oxygen. There was a significant rise (2á6 mmHg, P < 0á02) in fetal arterial PO2 in group iii, but in none of the other groups. In the first 3 h of liquid ventilation there was no difference in Jv between the groups. In group i, during hours 4-6 of liquid ventilation, there was a significant rise in secretion rate from 2á25 ± 0á88 to 3á74 ± 0á85 ml h-1 kg-1 (P < 0á001). In group iii, when comparing Jv in the first 3 h of liquid ventilation with that in the following 3 h period of liquid ventilation, a strong trend towards reduction in secretion was observed, falling from 3á03 ± 0á65 to 0á74 ± 0á92 ml h-1 kg-1 (three of the four experiments showed a significant decrease in Jv in hours 4-6). These experiments indicate that oxygen delivered to the fetus using liquid ventilation with haemoglobin solutions leads to increased LL secretion when oxygen delivery is small, and suggest there is a decrease in secretion with greater oxygen delivery to the lung.

We investigated the hypothesis that exogenous stimulation with thyrotrophin-releasing hormone (TRH) immediately prior to umbilical cord clamping can improve thermoregulatory adaptation after birth in near-term lambs delivered by Caesarean section. Lambs received an umbilical vein injection of saline ± TRH (8 µg) prior to cord clamping. The rate of change in colonic temperature and oxygen consumption after birth were not influenced by TRH, but TRH-treated lambs exhibited a greater incidence of shivering compared with controls over the first hour of neonatal life. Two and a half hours after birth, TRH-treated lambs possessed brown adipose tissue (BAT) with a higher thermogenic activity (i.e. GDP binding to mitochondrial protein), but their BAT had a reduced DNA content and they had less hepatic glycogen than control lambs. TRH administration had no effect on iodothyronine 5' deiodinase activity in BAT and liver, or on plasma concentrations of total triiodothyronine, thyroxine, cortisol or free fatty acids. Three TRH-treated but no control lambs, failed to establish continuous breathing, so tissues from these treated lambs together with time-matched controls were sampled 25 min after birth. These 'non-surviving' TRH-treated lambs had very high plasma catecholamine concentrations, but their lung weights were similar to controls. 'Surviving' TRH-treated lambs possessed lungs with less DNA than non-surviving TRH-treated lambs. It is concluded that umbilical vein injection of TRH prior to umbilical cord clamping increases the recruitment of both shivering and non-shivering thermogenesis after birth.

We investigated the influence of exogenous thyroxine (T4) administration in conjunction with level of feeding on adipose tissue and liver growth in postnatal lambs. Pairs of lambs were fed either 100 g (i.e. low fed) or 200 g (i.e. high fed) of milk powder per litre of reconstituted milk replacer over the first month of life. Half of the pairs of lambs were fed a bolus dose of T4 (15 mg (kg body weight)-1) daily until 8 days of age. Perirenal adipose tissue and hepatic tissue were sampled at either 8 or 35 days of age. High fed lambs grew faster, possessed more adipose tissue and had larger livers than low fed lambs at 8 and 35 days of age. T4 administration resulted in a lower thermogenic activity (i.e. GDP binding) in adipose tissue at 8 days of age in low, but not high fed lambs. There was no difference between groups in colonic temperature or oxygen consumption. Between 17 and 35 days of age high fed lambs previously treated with T4 had lower daily milk consumption than untreated siblings, but still attained the same growth rate. Plasma insulin-like growth factor-I concentrations were greater in high than low fed lambs, a relationship that was not influenced by T4 treatment. In adipose tissue, iodothyronine 5′ deiodinase activity was not influenced by T4 administration and was greater in high than low fed lambs. Hepatic iodothyronine 5′ deiodinase activity was not influenced by T4 administration in low fed lambs, but was reduced by T4 in high fed lambs. In conclusion, T4 administration over the first 8 days of life can accelerate the rate of decline in thermogenic activity of uncoupling protein-1. This effect is not observed when the level of feeding is increased. Following withdrawal of T4 treatment, high fed T4-treated lambs were able to maintain the same growth rate as untreated lambs despite having a lower food intake.

In nine 2- to 3-month-old hyperketonaemic piglets the kinetics of glucose and D-[beta]-hydroxybutyrate (D-BHB) metabolism were studied during hypo- and normocalcaemia in paired experiments. Hyperketonaemia (1á3 and 2á5 mmol D-BHB (l plasma)-1) was generated by a stepwise increase of DL-BHB infusion. Hypocalcaemia spontaneously developed in five piglets due to an inherited calcitriol deficiency and was induced in four control piglets by a continuous infusion of Na2-EDTA. The method of single isotopic marker injections of glucose and D-BHB was used to calculate replacement rates, rate constants and half-lives of glucose and D-BHB in plasma. When DL-BHB was infused at the same rate into normo- and hypocalcaemic piglets, hypocalcaemia reduced the rate constant of glucose by 20-30 % and the replacement rate of glucose by 34 %. In the presence of hyperketonaemia, hypocalcaemia increased the rate of replacement of D-BHB by 6-40 %. The replacement rate represents the sum of endogenous production and the rate of DL-BHB infusion. This observation shows that the endogenous production of D-BHB was higher during hypocalcaemia than during normocalcaemia.

The vasoregulatory role of nitric oxide (NO) and angiotensin II type 1 (AT1) receptors in the circulation of the submandibular gland (SMG) of rats was studied. The glandular blood flow was determined by means of laser Doppler flowmetry and rubidium isotope technique. The data obtained by these two methods correlated well (r = 0.77; P < 0.01). The AT1 receptor antagonist candesartan (0.5 mg kg-1, I.V.) reduced the vascular resistance in the SMG by 37 % (P < 0.05). By contrast, the NO synthase blocker L-NAME (15 mg kg-1, I.V.) significantly increased vascular resistance in the SMG both in candesartan-treated (P < 0.001) and non-treated (P < 0.001) animals. The increase in resistance was greater (P < 0.05) after previous blockade of AT1 receptors. These findings suggest that the AT1 receptors have an important role in the vasoregulation of the SMG in the rat. As a result of AT1 blockade, NO-dependent tone of glandular vessels may be enhanced significantly. Experimental Physiology (2002) 87.3, 327-333.

The effects of the muscarinic agonist oxotremorine-m (Oxo-m) on 45Ca and 86Rb fluxes, insulin secretion, cytoplasmic Ca2+ concentration ([Ca2+]i) and membrane potential in pancreatic B-cells were studied. Oxo-m (40-200 μM) increased the [Ca2+]i by about 250 nM, irrespective of the glucose concentration present in the medium (2·8-22 mM). This effect was reduced by 50 % upon the addition of EGTA. Oxo-m (50 μM) increased the 45Ca efflux from islets perifused in the absence or presence of [Ca2+]o, although under the former condition this efflux was transient. The difference between effluxes measured in the absence and presence of [Ca2+]o represents the sustained second component, which presumably reflects Ca2+ influx. In both the absence and presence of 11·2 mM glucose, Oxo-m (50 μM) transiently increased 86Rb efflux. In the presence of glucose, Oxo-m provoked a transient polarization of the B-cell membrane associated with an increase in the K+ permeability values. K+ permeability returned to basal values (no Oxo-m) after 1-2 min. These results indicate that the initial phase of Oxo-m-induced insulin secretion depends partially on intracellular Ca2+ release, and that the sustained enhancement of release depends on Ca2+ influx. The participation of a calcium release-activated current (ICRAC) is proposed to explain the sustained small changes in membrane potential.

The effects on renal functions of infusing glucagon (100 ng kg-1 min-1) into the portal vein system were studied in young anaesthetized sheep kept on a diet with normal protein and digestible energy contents. The clearance measurements of the left kidney functions showed a decreased urine flow rate (from 0·36 ± 0·04 to 0·18 ± 0·03 ml min-1; P < 0·01) within the first 10 min of glucagon infusion, falling to 0·07 ± 0·02 ml min-1 (P < 0·01) during the next 10 min. The glomerular filtration rate also decreased from 17·56 ± 2·92 ml min-1 to its lowest value, 3·34 ± 1·02 ml min-1, (P < 0·01) after 20 min of glucagon infusion. Decreases in urea excretion (from 53·1 ± 6·46 to 3·82 ± 1·27 μmol min-1; P < 0·01) and osmotic clearance (from 0·67 ± 0·13 to 0·09 ± 0·02 ml min-1; P < 0·01) occurred during the first 20 min. The plasma level of glucose increased from 4·01 ± 0·53 to 8·33 ± 1·12 mmol l-1 (P < 0·05) 20 min after the start of glucagon infusion. Twenty minutes after the start of glucagon infusion, the values of all the parameters measured began to recover, although they did not reach the control level, even after 1 h. Blood pressure was stable during the whole experiment. The results show that the effects of intraportally infused glucagon on kidney functions in sheep are the opposite of those obtained from similar experiments in laboratory animals with simple stomachs or in humans.

We evaluated the hypothesis that a 'central oxygen detector' in the brainstem is necessary for depressions of ventilatory activity to be manifested in the newborn. Decerebrate piglets, ventilated with 100 % O2, were studied following neuromuscular blockade. The vagi and carotid sinus nerves were sectioned bilaterally in order to remove the influence of the peripheral chemoreceptors. Activity of the phrenic nerve was recorded as the index of the central respiratory rhythm. This activity declined and, in some preparations, ceased upon ventilation with air or a hypoxic gas, at either normocapnia or hypercapnia. The degree of depression in hypercapnic hypoxia was greatest in the youngest piglets. Following a medial section of the brainstem, extending to the caudal pons, the depression was reduced. In some preparations, a similar reduction followed the placement of radiofrequency lesions in the caudal ventromedial pons. We conclude that a region of the caudal mesencephalon or pons is necessary for the manifestation of depressions of ventilatory activity in the newborn pig. Experimental Physiology (2003) 88.4, 509-515.

The existence of abdominal arterial baroreceptors has long been controversial. Previously difficulties have been encountered in localizing a stimulus to abdominal arteries without affecting reflexogenic areas elsewhere. In these experiments, using anaesthetized dogs, the abdomen was vascularly isolated at the level of the diaphragm, perfused through the aorta, and drained from the inferior vena cava to a reservoir. Changes in abdominal arterial pressure were effected by changing the perfusion pump speed. During this procedure the flow back to the animal from the venous outflow reservoir was held constant. Increases and decreases in abdominal arterial pressure resulted, respectively, in decreases and increases in perfusion pressure to a vascularly isolated hind-limb and in some dogs also a forelimb. Responses were significantly larger when carotid sinus pressure was high (120-180 mmHg) than when it was low (60 mmHg). Responses were still obtained after cutting vagus, phrenic and splanchnic nerves, but were abolished by spinal cord lesion at T12. These experiments provide evidence for the existence of abdominal arterial baroreceptors. The afferent pathway for the reflex vasodilatation appears to run in the spinal cord.

This study was designed to assess the haem-peptide interactions which occur during progressive haemoglobin hydrolysis by digestive enzymes and their relationship with haem iron digestive absorption. The behaviour of different haemoglobin hydrolysates was studied using the Ussing chamber model. Hydrolysates were produced from enzyme digestion of bovine haemoglobin at pH 3 by pepsin and at pH 10 by subtilisin. Samples with increasing degrees of hydrolysis (0-15 %) were studied. Biochemical assays (pyridine haemochromogen method and UV absorption spectra) were used to follow haem solubility and haem-peptide interactions in samples. Increasing the hydrolysis level of haemoglobin was associated with an enhanced iron uptake; the highest uptake rate was reached between 8 and 11 % of globin hydrolysis, whichever enzyme was used. The mechanisms rendering iron soluble and available differ between the two enzymes. The comparison between biochemical and absorption data suggests that the formation of soluble peptide-haem complexes was not sufficient to enhance haem iron absorption, since globin-bound iron is poorly absorbed; an efficient absorption occurred only when haem was loosely bound to low molecular weight peptides.

Knowledge of pleural liquid pressure (Pliq) and composition is crucial for studies concerning intrapleural fluid dynamics, and pleural fluid turnover. We measured Pliq at intercostal and costal levels in anaesthetized spontaneously breathing rabbits using a minimally invasive method that assures a long-lasting hydraulic continuity between the pleural liquid and the recording system. Polyethylene tubes were glued either to the exposed endothoracic fascia or inserted into a rib to provide a sealed connection to the recording system. After inducing a pneumothorax with nitrous oxide (N2O) via an intrapleural cannula, a hole ([similar]0·7 mm2) was pierced in the parietal pleura through the tube lumen. The tubes were then connected to pressure transducers and the whole system was filled with heparinized saline to the level of the parietal pleura; finally the pneumothorax was removed after N2O washout and Pliq recordings were performed. A different kind of tube was used to obtain microsamples of pleural fluid (2·5-3 µl) during spontaneous breathing; colloid osmotic pressure of the microsamples ([Pi]liq) was measured with an osmometer, and averaged 9·3 ± 1·5 cmH2O (n = 70 samples). When pooled and plotted against lung height end-expiratory intercostal and costal Pliq data scattered along a single regression line with a slope of -0·83 and -0·90 cmH2O cm-1 in supine and prone animals, respectively. End-inspiratory costal Pliq was significantly more subatmospheric than intercostal in the ventral region of the chest (P < 0·05), and less subatmospheric in the dorsal region, regardless of posture. The techniques presented here could be helpful in gaining a greater insight into the physiology and pathophysiology of the pleural space in terms of pleural fluid dynamics and turnover.

The inward transport of two purines, adenosine and hypoxanthine, at 37¡C by horse erythrocytes was compared. No mediated transport of adenosine was detected in horse erythrocytes, nor was saturable, high-affinity binding of the potent facilitated-diffusion inhibitor nitrobenzylthioinosine demonstrable in horse erythrocyte membranes. In contrast, erythrocytes from most horses possessed a saturable sodium-dependent hypoxanthine transporter (apparent Km, 100 ± 28 µM; Vmax, 0·20 ± 0·08 mmol (l cells)-1 h-1; means ± S.E.M., n = 5). Guanine inhibited hypoxanthine influx (apparent Ki, 24 ± 6 µM), but adenine and xanthine had no effect. Unlike human erythrocytes, no sodium-independent hypoxanthine transporter was detected in horse erythrocytes. There are, however, a small number of animals ([similar]15 %) whose erythrocytes fail to transport hypoxanthine. This variation appears to be under genetic control, but the precise nature of the control is unknown.