Small artificial waterbodies are widespread and persistent emitters of methane and carbon dioxide.

Inland waters play an active role in the global carbon cycle and emit large volumes of the greenhouse gases (GHGs), methane (CH4 ) and carbon dioxide (CO2 ). A considerable body of research has improved emissions estimates from lakes, reservoirs and rivers but recent attention has been drawn to the importance of small, artificial waterbodies as poorly quantified but potentially important emission hotspots. Of particular interest are emissions from drainage ditches and constructed ponds. These waterbody types are prevalent in many landscapes and their cumulative surface areas can be substantial. Furthermore, GHG emissions from constructed waterbodies are anthropogenic in origin and form part of national emissions reporting, whereas emissions from natural waterbodies do not (according to Intergovernmental Panel on Climate Change guidelines). Here, we present GHG data from two complementary studies covering a range of land uses. In the first, we measured emissions from nine ponds and seven ditches over a full year. Annual emissions varied considerably: 0.1-44.3 g CH4  m-2  year-1 and -36-4421 g CO2  m-2  year-1 . In the second, we measured GHG concentrations in 96 ponds and 64 ditches across seven countries, covering subtropical, temperate and sub-arctic biomes. When CH4 emissions were converted to CO2  equivalents, 93% of waterbodies were GHG sources. In both studies, GHGs were positively related to nutrient status (C, N, P), and pond GHG concentrations were highest in smallest waterbodies. Ditch and pond emissions were larger per unit area when compared to equivalent natural systems (streams, natural ponds). We show that GHG emissions from natural systems should not be used as proxies for those from artificial waterbodies, and that artificial waterbodies have the potential to make a substantial but largely unquantified contribution to emissions from the Agriculture, Forestry and Other Land Use sector, and the global carbon cycle.

NK Cells Accumulate in Infected Tissues and Contribute to Pathogenicity of Ebola Virus in Mice.

Understanding the immune parameters responsible for survival following Ebola virus (EBOV) infection is paramount for developing countermeasures. In lethal EBOV infections, levels of both NK and T cells decline drastically in the circulation and lymphoid tissues before death. However, the fate of these lymphocytes in viral replication sites remains unknown. In this study, reverse transcription-PCR (RT-PCR) and fluorescence-activated cell sorting (FACS) analysis were used to investigate lymphocyte frequencies in various infected mouse tissues after challenge with mouse-adapted EBOV (MA-EBOV). A decrease in NK cell numbers from systemic circulation was observed concomitant to an increase of these cells in tissues that are supporting active replication of EBOV. Unexpectedly, NK accumulation in virus replication sites correlated with enhanced EBOV disease progression in specific conditions; at a high challenge dose, NK-depleted mice displayed lower viremia and liver damage and higher hepatic T cell levels. Upregulation of UL16 binding protein 1 (ULBP-1) was detected in hepatic T cells, suggesting that NK cells participate in their elimination. Overall, this study supports the concept that NK cells accumulate in EBOV-infected tissues and can contribute to viral pathogenicity.IMPORTANCE Ebola virus (EBOV) outbreaks can claim numerous lives and also devastate the local health infrastructure, as well as the economy, of affected countries. Lethal EBOV infection has been documented to decrease the levels of several immune cells in the blood that are necessary to defend the host. This decrease in immune cells is, however, not observed in individuals who survive EBOV infection. Having a better grasp of how these immune cells are lost is therefore of high importance to develop and improve new and existing therapeutics. The significance of our research is in identifying the mechanism responsible for the apparent loss of immune cells in lethal EBOV infection. This will allow therapeutic options aimed at preventing the loss of these immune cells, therefore allowing infected individuals to better fight the infection.

Problem-solving and learning in Carib grackles: individuals show a consistent speed-accuracy trade-off.

The generation and maintenance of within-population variation in cognitive abilities remain poorly understood. Recent theories propose that this variation might reflect the existence of consistent cognitive strategies distributed along a slow-fast continuum influenced by shyness. The slow-fast continuum might be reflected in the well-known speed-accuracy trade-off, where animals cannot simultaneously maximise the speed and the accuracy with which they perform a task. We test this idea on 49 wild-caught Carib grackles (Quiscalus lugubris), a tame opportunistic generalist Icterid bird in Barbados. Grackles that are fast at solving novel problems involving obstacle removal to reach visible food perform consistently over two different tasks, spend more time per trial attending to both tasks, and are those that show more shyness in a pretest. However, they are also the individuals that make more errors in a colour discrimination task requiring no new motor act. Our data reconcile some of the mixed positive and negative correlations reported in the comparative literature on cognitive tasks, suggesting that a speed-accuracy trade-off could lead to negative correlations between tasks favouring speed and tasks favouring accuracy, but still reveal consistent strategies based on stable individual differences.

Independent appearance of an innovative feeding behaviour in Antillean bullfinches.

Behavioural innovations have been largely documented in birds and are thought to provide advantages in changing environments. However, the mechanisms by which behavioural innovations spread remain poorly known. Two major mechanisms are supposed to play a fundamental role: innovation diffusion by social learning and independent appearance of the same innovation in different individuals. Direct evidence for the independent emergence of the same innovation in different individuals is, however, lacking. Here, we show that a highly localized behavioural innovation previously observed in 2000 in Barbados, the opening of sugar packets by Loxigilla barbadensis bullfinches, persisted more than a decade later and had spread to a limited area around the initial site. More importantly, we found that the same innovation appeared independently in other, more distant, locations on the same island. On the island of St-Lucia, 145 km from Barbados, we also found that the sister species of the Barbados bullfinch, the Lesser Antillean bullfinch Loxigilla noctis developed the same innovation independently. Finally, we found that a third species, the Bananaquit Coereba flaveola, exploited the bullfinches' technical innovation to benefit from this new food source. Overall, our observations provide the first direct evidence of the independent emergence of the same behavioural innovation in different individuals of the same species, but also in different species subjected to similar anthropogenic food availability.

Advances in hematopoietic stem cell culture.

Recent advances in our understanding of the earliest stages of hematopoietic cell differentiation, and how these may be manipulated under defined conditions in vitro, have set the stage for the development of robust bioprocess technology applicable to hematopoietic cells. Sensitive and specific assays now exist for measuring the frequency of hematopoietic stem cells with long-term in vivo repopulating activity from human as well as murine sources. The production of natural or engineered ligands through recombinant DNA and/or combinatorial chemistry strategies is providing new reagents for enhancing the productivity of hematopoietic cell cultures. Multifactorial and dose-response analyses have yielded new insight into the different types and concentrations of factors required to optimize the rate and the extent of amplification of specific subpopulations of primitive hematopoietic cells. In addition, the rate of cytokine depletion from the medium has also been found to be dependent on the types of cell present. The discovery of these cell-type-specific parameters affecting cytokine concentrations and responses has introduced a new level of complexity into the design of optimized hematopoietic bioprocess systems.

Cell division tracking and expansion of hematopoietic long-term repopulating cells.

The combined use of rigorous assays for quantitating transplantable stem cell numbers and precise cell labeling and tracking procedures have provided definitive evidence that stem cell self-renewal divisions can occur in vitro in the absence of stromal feeder layers. These findings set the stage for defining conditions that may alter the ability of these cells to maintain their primitive status when mitogenically activated.

Effects of somatostatin and hypothalamic ventromedial lesions on GH release induced by morphine.

Morphine in intravenous doses ranging from 10 mug/kg to 8 mg/kg was shown to be effective in stimulating GH release in the unanesthetized rat. The response to the log of the dose was linear over a range of 10 to 1000 mug/kg. Somatostatin (GH-release inhibiting factor) administered SC in a dose of 200 mug/kg 5 min before morphine prevented the GH rise. Neither inhibitors of catecholamine or serotonin synthesis nor blockage of alpha and beta-adrenergic receptors had any effect on the response. The response was partially blocked in animals with large hypothalamic ventromedial (VMN) lesions. Such lesions completely abolished the GH response to pentobarbital. These results indicate that morphine is a remarkably potent agent for stimulation of GH release but the precise mechanism and site of action of the drug remain to be determined.

Evidence that reduced growth hormone secretion observed in monosodium glutamate-treated rats is the result of a deficiency in growth hormone-releasing factor.

The present experiments were designed to examine various aspects of GH secretion in adult male rats given monosodium glutamate (MSG; 4 mg/g BW, sc) during the neonatal period. MSG-treated animals sustained lesions localized to the hypothalamic arcuate nuclei (ARC) and had reduced nasal-anal lengths and body weights. Anterior pituitary (AP) weights were decreased, but AP concentrations of GH and PRL were not significantly altered. Analysis of pulsatile GH secretion showed depressed GH pulses and prolonged GH trough periods. Mean 5-h plasma GH levels were reduced, whereas PRL levels were not affected. Morphine sulfate (MS) at doses of 0.01, 0.1, 1.0, and 3.0 mg/kg induced a prompt rise in GH during the 45 min after drug administration in controls. MSG-treated animals showed a significant rise in GH only with 1.0 and 3.0 mg/kg MS. A significant elevation in PRL was found in both control and MSG-treated animals after 1.0 and 3.0 mg/kg MS. The pentobarbital-induced rise in GH was also blunted in MSG-treated animals. MSG-treated animals which were administered antisomatostatin serum showed elevated GH trough and mean GH levels, with no apparent effect on GH peak levels. In view of the mechanisms by which MS and pentobarbital act to increase GH secretion, the present data suggest that the GH regulatory deficit observed in MSG-treated rats is due to a relative loss of GH-releasing factor secondary to ARC damage.

Neuropharmacological regulation of episodic growth hormone and prolactin secretion in the rat.

The effects on GH and PRL secretion of several pharmacological agents known to modify central neurotransmitter action were determined in unanesthetized male rats. Phenoxybenzamine, an alpha-adrenergic blocker (5 mg/kg iv), abolished episodic GH secretion and caused elevation of serum PRL levels. Propranolol, a beta-adrenergic blocker (5 mg/kg iv), had no effect on GH secretion and caused a small but significant depression in PRL levels. Parachlorophenylalanine methyl ester, an inhibitor of tryptophan hydroxylase (300-350 mg/kg ip), resulted in significant inhibition of GH pulsatile secretion and suppressed PRL levels. Methysergide hydrogen maleinate (25 mg/kg iv), a serotonin receptor antagonist, also inhibited GH secretion, but produced a transient stimulation in PRL levels. Atropine sulfate (2 mg/kg iv) caused significant suppression in GH secretion, but had no effect on PRL. Picrotoxin, a gamma-aminobutyric acid antagonist, in a subconvulsive dose of 1-3 mg/kg iv, also depressed GH episodic secretion but did not affect PRL levels. These results indicate that several neurotransmitters, i.e., norepinephrine, serotonin, acetylcholine, and gamma-aminobutyric acid, found in high concentration in the hypothalamus, influence GH and PRL secretion.

Cell cycle distribution of primitive haematopoietic cells stimulated in vitro and in vivo.

A novel approach is used to study the proliferating behaviour of primitive haematopoietic cell populations in response to different stimuli. A mathematical model based on the average proportion of apoptotic, dividing and quiescent cells in primitive haematopoietic cell populations is developed to describe the mitotic history of 5- (and 6-) carboxyfluorescein diacetate succinimidyl ester-labelled cells. The cell cycle distributions in different cytokine-supplemented cultures of primitive human and mouse bone marrow cells are determined and compared with those found in vivo. The results indicate that a combination of flt-3 ligand, Steel factor and interleukin-11 or hyper-interleukin-6 provide a level of mitogenic stimulation similar to that existing in vivo after a myeloablative radiation dose. The comparison of the cell cycle distribution obtained for different cultures of human bone marrow CD34(+)(45RA/71)(-) cells demonstrates that the addition of flt-3 ligand in these cultures decreases apoptosis significantly but does not reduce quiescence. In addition, in vivo and in vitro, it was found that more than 3 days of stimulation are required to recruit a maximum number of quiescent cells into active cell cycle. These kinetics of cell cycle activation are found to be similar to those identified for the haematopoietic stem cells compartment in the same cultures. This mathematical analysis provides a useful tool for the development of haematopoietic stem cell culture processes for clinical applications.

Multiple regulatory elements control the basal promoter activity of the human alpha 4 integrin gene.

It has been suggested that expression of the genes encoding the alpha 4/beta 1 integrin increases during wound healing of the cornea. As a first step in understanding the mechanisms required to stimulate alpha 4 gene expression during this process, we defined the minimal upstream sequence required to direct basal promoter activity for this gene. Using deletion analyses of the alpha 4 gene upstream sequence, we identified two functionally important negative regulatory elements. Dimethylsulfate (DMS) methylation interference assays provided evidence for the binding of a single nuclear protein to tandemly repeated homologous cis-acting elements (designated alpha 4.1 and alpha 4.2) from the alpha 4 basal promoter that share the core sequence 5'-GTGGGT-3'. The formation of a protection only at alpha 4.1 in DNase I footprinting suggested that it is the primary target element for the binding of nuclear proteins. Three distinct nuclear proteins bound a double-stranded oligonucleotide bearing the DNA sequence of alpha 4.1 to produce specific DNA-protein complexes (R1 to R3) in gel-shift assays, from which that producing R3 was identified as the protein yielding DNase I protection at alpha 4.1. Detailed mutational analysis of alpha 4.1 and alpha 4.2 indicated that both elements positively regulate gene expression in primary cultures of corneal epithelial cells and Jurkat tissue culture cells, which is consistent with the deletion analysis. However, when transiently transfected into pituitary GH4C1, the alpha 4.2 mutants yielded increased chloramphenicol acetyl transferase activity therefore demonstrating that these elements have the ability to function either as positive or negative regulators of gene transcription in a manner that is dependent on the type of cell transfected.

Characterization of human hematopoietic cells with short-lived in vivo repopulating activity.

Recent studies with purified hematopoietic stem cells in vitro support a model of stem cell self-renewal control that involves distinct mechanisms regulating permissiveness to and execution of lineage restriction. Such a model predicts the existence of phenotypically separable populations of hematopoietic cells that are pluripotent and either capable or incapable of extensive self-renewal. Such populations have been previously described in the mouse. We describe here the first evidence that such cells can now be identified in humans using different types of immunodeficient mice as hosts.

Introduction to stem cell biology in vitro. Threshold to the future.

Transplantable hematopoietic cells with multilineage reconstituting ability can be quantitated in suspensions of human or murine cells using similar assay procedures. The incorporation into these assays of stringently defined functional endpoints ensures a high degree of specificity for the cells detected. Application of these assays to stem cell-containing suspensions after they have been stimulated for several days with defined cytokines in vitro, or by a mixture of defined and/or undefined factors in vivo, has shown that net amplifications in these populations can be obtained under both circumstances. Such studies have allowed cytokine conditions that support stem cell self-renewal divisions to be identified and have also provided evidence that stem cell regeneration can be manipulated both in vitro and in vivo by altering the molecular milieu of the responding cells. These observations pave the way to future delineation of mechanisms that control the normal behavior, pathology and future clinical exploitation of hematopoietic stem cell populations.

Relations of diabetes intrusiveness and personal control to symptoms of depression among adults with diabetes.

The generalizability of a model linking illness characteristics to psychosocial well-being was tested in a cross-sectional study of 237 adults with type 2 diabetes. It was hypothesized that diabetic complications increase illness intrusiveness, which in turn increases depressive symptomatology either directly or indirectly by reducing personal control over health outcomes. Illness intrusiveness was defined as the result of disruptions of valued activities and interests due to constraints imposed by the illness. An excellent fit of this model to the data was found using structural equation modeling. The model explained 65% of the variance in depressive symptomatology. Assessment of an alternative model excluding personal control suggested that the extent to which diabetes intrudes in life, rather than diabetic complications per se or personal control, is a key factor in relation to depressive symptomatology in individuals with diabetes.

Structural and in vivo mechanical characterization of canine patellar cartilage: a closed chondromalacia patellae model.

The purpose of this project was to study the relationship between the structure of the patellar cartilage and its response to static compressive loading with a closed chondromalacia patellae model. An animal model was used to induce degeneration of the patella that was monitored quantitatively and qualitatively as a function of time. Ten adult mongrel dogs had their left patellofemoral groove replaced by a customized metallic implant covered with a thin film of polyethylene for periods of 3 months (five dogs) and 6 months (five dogs). An indenter was designed to perform mechanical indentation testing on the patellar cartilage in situ. The animals were anesthetized and the response of patellar cartilage to a static compressive load of 4.5 MPa was monitored for 20 min and its relaxation after load removal for 20 min. Indentation tests were performed every 3 months of the implantation period. At the end of the implantation period, the patellae were processed for histology, and sections were stained with Safranin-O indicative of the proteoglycans content. Macroscopically, no apparent degeneration or fibrillation of the patellar surfaces was observed after 3 or 6 months of implantation. However, the patellar surface showed a change in coloration after 6 months. A 17 +/- 3% and 37 +/- 8% deformation of the cartilage were calculated for the 3-month and 6-month specimens, respectively. Histologically, a progressive loss of proteoglycans was observed in the matrix as a function of implantation time. These results indicated that an increase in cartilage compliance is associated with an intrinsic remodeling of the cartilage matrix and that these changes might occur without external signs of degeneration and can be quantified.

Methylene blue administration fails to confer neuroprotection in two amyotrophic lateral sclerosis mouse models.

Approximately 20% cases of familial amyotrophic lateral sclerosis (ALS) are caused by mutations in the gene encoding Cu/Zn superoxide dismutase (SOD1). Recent studies have shown that methylene blue (MB) was efficient in conferring protection in several neurological disorders. MB was found to improve mitochondrial function, to reduce reactive oxygen species, to clear aggregates of toxic proteins, and to act as a nitric oxide synthase inhibitor. These pleiotropic effects of relevance to ALS pathogenesis led us to test MB in two models of ALS, SOD1(G93A) mice and TDP-43(G348C) transgenic mice. Intraperitoneal administration of MB at two different doses was initiated at the beginning of disease onset, at 90 days of age in SOD1(G93A) and at 6 months of age in TDP-43(G348C) mice. Despite its established neuroprotective properties, MB failed to confer protection in both mouse models of ALS. The lifespan of SOD1(G93A) mice was not affected by MB treatment. The declines in motor function, reflex score, and body weight of SOD1(G93A) mice remained unchanged. MB treatment had no effect on motor neuron loss and aggregation or misfolding of SOD1. A combination of MB with lithium also failed to provide benefits in SOD1(G93A) mice. In TDP-43(G348C) mice, MB failed to improve motor function. Cytosolic translocation of TDP-43, ubiquitination and inflammation remained also unchanged after MB treatment of TDP-43(G348C) mice.

Polysaccharide concentration and molecular weight effects on the oxygen mass transfer in a reciprocating plate bioreactor.

In most polysaccharide fermentations, the nature of the fermentation broth changes drastically with time and, as a result, the overall oxygen mass transfer coefficient (K(L)a) can vary by orders of magnitude. To obtain a better understanding of this phenomenon, an experimental program was devised to study the respective influence of molecular weight and concentration of dextran solutions on K(L)a. Experiments were conducted in a reciprocating plate bioreactor. This bioreactor uses a stack of perforated plates that is reciprocated axially in the column and it is therefore well suited for mixing viscous liquid broths and providing uniform overall mass transfer coefficients. The variation of K(L)a with the power input per unit volume and the superficial gas velocity were obtained for three ranges of molecular weights and five concentrations of dextran. In every medium, two regimes of operation were observed as a function of the power input per unit volume: a first regime, at low power inputs per unit volume where K(L)a remains constant until a threshold of power input is attained; and a second regime, which is characterized by a steep increase of K(L)a as a function of the power input per unit volume. The presence of dissolved biological macromolecules, not only because of their effect on the rheology of the medium but also because their effect on the gas-liquid interface, has a significant impact on K(L)a. It was found that, generally, small concentrations of polysaccharide favor oxygen mass transfer despite the increase in medium viscosity. However, the respective influence of polysaccharide concentration and molecular weight was different for the two regimes of operation. (c) 1996 John Wiley & Sons, Inc.

Rheological properties of the human lumbar spine ligaments.

The purpose of this study is to provide a better understanding of the rheological properties of the lumbar spinal ligaments under subfailure physiological loads. Non-destructive tests including an hysteresis experiment, stress-relaxation and stepwise load-relaxation tests were used to investigate the time-dependent properties of the interspinous-supraspinous ligament complex. Using a reduced relaxation function, the viscoelastic behaviour over the experimental time-scale was described by a linear function of the logarithm of time. Internal damping of ligament substance dissipates about 36% of the mechanical energy applied during physiological loading. Local elastic stiffness is found to be two to four times global stiffness of the bone-ligament-bone complex. These physical parameters (stiffness, energy dissipation, hysteresis, relaxation, etc) can be used to improve computer models of the lumbar spinal column.

Physiological changes associated with unilateral pulmonary ventilation during operations on the lung.

No important changes in the respiratory parameters were observed during one-lung anaesthesia. However, this kind of thoracic intervention can be accompanied by a dramatic fall in arterial oxygen tension. Methods to avoid tissue hypoxia have been described. In our series no complications whatever occurred in the operative and post-operative periods which could be related to the oxygenation of the patient.