An isolated perfused human placental lobule model for multiple indicator dilution studies

We describe a method for multiple indicator dilution studies in the isolated perfused human placental lobule developed to investigate the relationships between changes in pressure and flow and solute clearance. A peripheral lobule of a human placenta is perfused with a tissue culture-based medium and the perfusate oxygen tension, arterial and venous pressures, pH and perfusion temperature continuously monitored by a computerized system. Flow rates are readily changed. Bolus injections of vascular, extracellular and water space markers, and study compounds can be made into either maternal or fetal circulations, and precisely timed outflow fractions can be collected with computer-controlled fraction collectors, allowing simultaneous determination of concentration-time profiles of each marker. (C) 1997 Elsevier Science Inc.

Retinal and lenticular ultrastructure in the aestivating salamanderfish, Lepidogalaxias salamandroides (Galaxiidae, Teleostei) with special reference to a new type of photoreceptor mosaic

The salamanderfish, Lepidogalaxias salamandroides (Galaxiidae, Teleostei) is endemic to southwestern Australia and inhabits shallow, freshwater pools which evaporate during the hot summer months. Burrowing into the substrate in response to falling water levels allows these fish to aestivate for extended periods of time while encapsulated in a mucous cocoon even when the pools contain no water. Only a few minutes after a major rainfall, these fish emerge into relatively clear water which subsequently becomes laden with tannin, turning the water black and reducing the pH to approximately 4.3. As part of a large study of the visual adaptations of this unique species, the retinal and lenticular morphology of the aestivating salamanderfish is examined at the level of the light and electron microscopes. The inner retina is highly vascularised by a complex system of vitreal blood vessels, while the outer retina receives a blood supply by diffusion from a choriocapillaris. This increased retinal blood supply may be an adaptation for reducing the oxygen tension during critical periods of aestivation. Large numbers of Muller cells traverse the thickness of the retina from the inner to the outer limiting membranes. The ganglion cells are arranged in two ill-defined layers, separated from a thick inner nuclear layer containing two layers of horizontal cells by a soma-free inner plexiform layer. The photoreceptors can be divided into three types typical of many early actinopterygian representatives; equal double cones, small single cones and large rods (2:1:1). These photoreceptors are arranged into a unique regular square mosaic comprising a large rod bordered by four equal double cones with a small single cone located at the corner of each repeating unit. The double cones may optimise perception of mobile prey which it tracks by flexion of its head and neck and the large rods may increase sensitivity in the dark tannin-rich waters in which it lives. Each single cone also possesses a dense collection of polysomes and glycogen (a paraboloid) beneath its ellipsoid, the first such finding in teleosts. The retinal pigment epithelium possesses melanosomes, pha,oocytes and a large number of mitochondria. The anatomy of the retina and the photoreceptor mosaic is discussed in relation to the primitive phylogeny of this species and its unique life history.

PrrC from Rhodobacter sphaeroides, a homologue of eukaryotic Sco proteins, is a copper-binding protein and may have a thiol-disulfide oxidoreductase activity

PrrC from Rhodobacter sphaeroides provides the signal input to a two-component signal transduction system that senses changes in oxygen tension and regulates expression of genes involved in photosynthesis (Eraso, J.M. and Kaplan, S. (2000) Biochemistry, 39, 2052-2062; Oh, J.-I. and Kaplan, S. (2000) EMBO J. 19, 42374247). It is also a homologue of eukaryotic Sco proteins and each has a C-x-x-x-C-P sequence. In mitochondrial Sco proteins these cysteines appear to be essential for the biogenesis Of the Cu-A centre of respiratory cytochrome oxidase. Overexpression and purification of a water-soluble and monomeric form of PrrC has provided sufficient material for a chemical and spectroscopic study of the properties of the four cysteine residues of PrrC, and its ability to bind divalent cations, including copper. PrrC expressed in the cytoplasm of Escherichia coli binds Ni2+ tightly and the data are consistent with a mononuclear metal site. Following removal of Ni2+ and formation of renatured metal-free rPrrC (apo-PrrC), Cu2+ could be loaded into the reduced form of PrrC to generate a protein with a distinctive UV-visible spectrum, having absorbance with a lambda(max) of 360 nm. The copper:PrrC ratio is consistent with the presence of a mononuclear metal centre. The cysteines of metal-free PrrC oxidise in the presence of air to form two intramolecular disulfide bonds, with one pair being extremely reactive. The cysteine thiols with extreme O-2 sensitivity are involved in copper binding in reduced PrrC since the same copper-loaded protein could not be generated using oxidised PrrC. Thus, it appears that PrrC, and probably Sco proteins in general, could have both a thiol-disulfide oxidoreductase function and a copper-binding role. (C) 2002 Published by Elsevier Science B.V. on behalf of the Federation of European Biochemical Societies.

Identification and in vivo characterization of PpaA, a regulator of photosystem formation in Rhodobacter sphaeroides

A regulatory protein, PpaA, involved in photosystem formation in the anoxygenic phototrophic proteobacterium Rhodobacter sphaeroides has been identified and characterized in vivo. Based on the phenotypes of cells expressing the ppaA gene in extra copy and on the phenotype of the ppaA null mutant, it was concluded that PpaA activates photopigment production and puc operon expression under aerobic conditions. This is in contrast to the function of the PpaA homologue from Rhodobacter capsulatus, AerR, which acts as a repressor under aerobic conditions [Dong, C., Elsen, S., Swem, L. R. & Bauer, C. E. (2002). J Bacteriol 184, 2805-2814]. The expression of the ppaA gene increases several-fold in response to a decrease in oxygen tension, suggesting that the PpaA protein is active under conditions of low or no oxygen. However, no discernible phenotype of a ppaA null mutant was observed under anaerobic conditions tested thus far. The photosystem gene repressor PpsR mediates repression of ppaA gene expression under aerobic conditions. Sequence analysis of PpaA homologues from several anoxygenic phototrophic bacteria revealed a putative corrinoid-binding domain. It is suggested that PpaA binds a corrinoid cofactor and the availability or structure of this cofactor affects PpaA activity.

Hypoxia induces chondrocyte-specific gene expression in mesenchymal cells in association with transcriptional activation of Sox9

Endochondral bone is formed during an avascular period in an environment of low oxygen. Under these conditions, pluripotential mesenchymal stromal cells preferentially differentiate into chondrocytes and form cartilage. In this study, we investigated the hypothesis that oxygen tension modulates bone mesenchymal cell fate by altering the expression of genes that function to promote chondrogenesis. Microarray of RNA samples from ST2 cells revealed significant changes in 728 array elements (P < 0.01) in response to hypoxia. Real-time PCR on these RNA samples, and separate samples from C3H10T1/2 cells, revealed hypoxia-induced changes in the expression of additional genes known to be expressed by chondrocytes including Sox9 and its downstream targets aggrecan and Col2a. These changes were accompanied by the accumulation of mucopolysacharide as detected by alcian blue staining. To investigate the mechanisms responsible for upregulation of Sox9 by hypoxia, we determined the effect of hypoxia on HIF-1 alpha levels and Sox9 promoter activity in ST2 cells. Hypoxia increased nuclear accumulation of HIF-1 alpha and activated the Sox9 promoter. The ability of hypoxia to transactivate the Sox9 promoter was virtually abolished by deletion of HIF-1 alpha consensus sites within the proximal promoter. These findings suggest that hypoxia promotes the differentiation of mesenchymal cells along a chondrocyte pathway in part by activating Sox-9 via a HIF-1 alpha-dependent mechanism. (c) 2005 Elsevier Inc. All rights reserved.

Diel 'tuning' of coral metabolism: physiological responses to light cues

Hermatypic-zooxanthellate corals track the diel patterns of the main environmental parameters temperature, UV and visible light - by acclimation processes that include biochemical responses. The diel course of solar radiation is followed by photosynthesis rates and thereby elicits simultaneous changes in tissue oxygen tension due to the shift in photosynthesis/respiration balance. The recurrent patterns of sunlight are reflected in fluorescence yields, photosynthetic pigment content and activity of the two protective enzymes superoxide dismutase (SOD) and catalase (CAT), enzymes that are among the universal defenses against free radical damage in living tissue. All of these were investigated in three scleractinian corals: Favia favus, Plerogyra sinuosa and Goniopora lobata. The activity of SOD and CAT in the animal host followed the course of solar radiation, increased with the rates of photosynthetic oxygen production and was correlated with a decrease in the maximum quantum yield of photochemistry in Photosystem H (PSII) (Delta F'/F-m'). SOD and CAT activity in the symbiotic algae also exhibited a light intensity correlated pattern, albeit a less pronounced one. The observed rise of the free-radical-scavenger enzymes, with a time scale of minutes to several hours, is an important protective mechanism for the existence and remarkable success of the unique cnidarian-dinoflagellate associations, in which photosynthetic oxygen production takes place within animal cells. This represents a facet of the precarious act of balancing the photosynthetic production of oxygen by the algal symbionts with their destructive action on all living cells, especially those of the animal host.

A model to investigate hepatic extraction of oxygen during anaesthesia in the dog

Measurement of hepatic oxygen extraction was performed on six healthy Greyhound dogs over a two hour period. The Greyhounds were anaesthetised and a right subcostal surgical incision performed. Ultrasonic flow transducers were used to measure flow rate in the hepatic artery and the portal vein. The blood oxygen tensions in arterial blood and in the portal and hepatic veins were also measured. Hepatic oxygen extraction remained stable throughout the study, despite a steady decline in arterial blood pressure. The methodology described in this study provides a direct measure of oxygen uptake by the liver in the dog and could readily be used to investigate hepatic uptake of drugs. (C) 2003 Elsevier Ltd. All rights reserved.

Use of a thermodilution catheter to measure core body temperature, central venous pressure and cardiac output during anaesthesia and surgery in the dog

The use of thermodilution and other methods of monitoring in dogs during surgery and critical care was evaluated. Six Greyhounds were anaesthetised and then instrumented by placing a thermodilution catheter into the pulmonary artery via the jugular vein. A catheter in the dorsal pedal artery also permitted direct measurement of arterial pressures. Core body temperature (degreesC) and central venous pressure (mmHg) were measured, while cardiac output (mL/min/kg) and mean arterial pressure (mmHg) were calculated. A mid-line surgical incision was performed and the physiological parameters were monitored for a total of two hours. All physiological parameters generally declined, although significant increases (P<0.05) were noted for cardiac output following surgical incision. Central venous pressure was maintained at approximately 0mmHg by controlling an infusion of sterile saline. Core body temperature decreased from 37.1+/-0.6degreesC (once instrumented) to 36.6+/-0.60degreesC (at the end of the study), despite warming using heating pads. Physiological parameters indicative of patient viability will generally decline during surgery without intervention. This study describes an approach that can be undertaken in veterinary hospitals to accurately monitor vital signs in surgical and critical care patients.

Geochemical mass-balance and oxygen-isotope constrains on silcrete formation and its paleoclimatic implications in Southern Australia

Two geographically distinct silcrete associations are present in southern Australia, inland and eastern; these were sampled in central South Australia and central Victoria, respectively, At each site, both silicified and immediately adjacent unsilicified parent material were collected. Analytical data from these pairs were used to construct isocons, assuming Zr immobility, and to calculate the volume change and amount of silica introduced during silicification, These results, together with whole-rock oxygen isotope compositions, were used to determine the delta(18)O of th, introduced silica, The results show that the eastern silcretes in central Victoria are probably linked genetically to the associated basalts, weathering of which supplied the introduced silica, This conclusion is based on the close spatial connection between the two, as well as the substantial amount of introduced silica in the silcretes (greater than in the inland silcretes), resulting in volume increases in some eastern silcretes, Oxygen isotopic calculations for the silcretes indicate that the silica precipitated from groundwaters at temperatures slightly higher than present conditions. Silcrete formation apparently occurred during the Miocene and Pliocene (basalts in Victoria younger than Pliocene lack associated silcrete) and may reflect the much wetter climate in southeastern Australia at that time. The inland silcretes of central South Australia can be divided into pedogenic (the most common) and groundwater varieties. The pedogenic silcretes, which show typical soil features like columnar and nodular textures, contain moderate amounts of introduced silica that precipitated by evaporation from saline groundwaters, For the groundwater silcretes, which have massive textures and formed at or close to the water table, insufficient data are available to determine the mode of formation. The inland pedogenic silcretes have probably been farming from the Eocene-Miocene to the present, implying that conditions of seasonally high evaporation have occurred in central Australia during this time period. Thus silcrete formation depends on a complex interplay between climate and silica supply, and it is impossible to generalize that the presence of silcrete is indicative of a particular climate. Likewise, the elemental composition of silcretes, particularly Ti content, is not necessarily of climatic significance, Nevertheless, detailed geochemical and oxygen isotopic studies of a silcrete and its parent material can elucidate the mechanisms of silcrete formation, and if evaporation is indicated as a major factor in silcrete formation, then the climate at the time was likely to have been at least seasonally arid.

A pore-size-dependent equation of state for multilayer adsorption in cylindrical mesopores

The chemical potential of adsorbed film inside cylindrical mesopores is dependent on the attractive interactions between the adsorbed molecules and adsorbent, the curvature of gas/adsorbed phase interface, and surface tension. A state equation of the adsorbed film is proposed to take into account the above factors. Nitrogen adsorption on model adsorbents, MCM-41, which exhibit uniform cylindrical channels, are used to verify the theoretical analysis. The proposed theory is capable of describing the important features of adsorption processes in cylindrical mesopores. According to this theory, at a given relative pressure, the smaller the pore radius is, the thicker the adsorbed film will be. The thickening of adsorbed films in the pores as the vapor pressure increases inevitably causes an increase in the interface curvature, which consequently leads to capillary condensation. Besides, this study confirmed that the interface tension depends substantially on the interface curvature in small mesopores. A quantitative relationship between the condensation pressure and the pore radius can be derived from the state equation and used to predict the pore radius from a condensation pressure, or vice versa.

Tension regulation during lengthening and shortening actions of the human soleus muscle

In the present study we investigated tension regulation in the human soleus (SOL) muscle during controlled lengthening and shortening actions. Eleven subjects performed plantar flexor efforts on an ankle torque motor through 30 degrees of ankle displacement (75 degrees-105 degrees internal ankle angle) at lengthening and shortening velocities of 5, 15 and 30 degrees s(-1). To isolate the SOL from the remainder of the triceps surae, the subject's knee was flexed to 60 degrees during all trials. Voluntary plantar flexor efforts were performed under two test conditions: (1) maximal voluntary activation (MVA) of the SOL, and (2) constant submaximal voluntary activation (SVA) of the SOL. SVA trials were performed with direct visual feedback of the SOL electromyogram (EMG) at a level resulting in a torque output of 30% of isometric maximum. Angle-specific (90 degrees ankle angle) torque and EMG of the SOL, medial gastrocnemius (MG) and tibialis anterior (TA) were recorded. In seven subjects from the initial group, the test protocol was repeated under submaximal percutaneous electrical activation (SEA) of SOL (to 30% isometric maximal effort). Lengthening torques were significantly greater than shortening torques in all test conditions. Lengthening torques in MVA and SVA were independent of velocity and remained at the isometric level, whereas SEA torques were greater than isometric torques and increased at higher lengthening velocities. Shortening torques were lower than the isometric level for all conditions. However, whereas SVA and SEA torques decreased at higher velocities of shortening, MVA torques were independent of velocity. These results indicate velocity- and activation-type-specific tension regulation in the human SOL muscle.

Incubation of eggs of the Australian broad-shelled turtle, Chelodina expansa (Testudinata : Chelidae), at different temperatures: effects on pattern of oxygen consumption and hatchling morphology

Incubation temperature influences embryonic development and the morphology of resultant hatchlings in many species of turtle but few studies have addressed its effect on oxygen consumption and total embryonic energy expenditure. Eggs of the Australian broad-shelled river turtle, Chelodina expansa, were incubated at constant temperatures of 24 degrees C and 28 degrees C to determine the effect of temperature on oxygen consumption, embryonic energy expenditure and hatchling morphology. All embryos at both incubation temperatures experienced a period of developmental diapause immediately after oviposition. Once this initial diapause was broken, embryos underwent a further period of developmental arrest when the embryo was still very small and had minimal oxygen consumption (