Characterization of nucleoside triphosphate diphosphohydrolase activity in Trichomonas gallinae and the influence of penicillin and streptomycin in extracellular nucleotide hydrolysis

Here we described an nucleoside triphosphate diphosphohydrolase (NTPDase) activity in living trophozoites of Trichomonas gallinae. The enzyme hydrolyzes a variety of purine and pyrimidine nucleoside di- and triphosphates in an optimum pH range of 6.0-8.0. This enzyme activity was activated by high concentrations of divalent cations, such as calcium and magnesium. Contaminant activities were ruled out because the enzyme was not inhibited by classical inhibitors of ATPases (ouabain, 5.0 mM sodium azide, oligomycin) and alkaline phosphatases (levamisole). A significant inhibition of ATP hydrolysis (38%) was observed in the presence of 20 mM sodium azide. Sodium orthovanadate inhibited ATP and ADP hydrolysis (24% and 78%), respectively. The apparent K(M) (Michaelis constant) values were 667.62+/-13 microM for ATP and 125+/-5.3 microM for ADP. V(max) (maximum velocity) values were 0.44+/-0.007 nmol Pi min(-1) per 10(6) trichomonads and 0.91+/-0.12 nmol Pi min(-1) per 10(6) trichomonads for ATP and ADP, respectively. Moreover, we showed a marked decrease in ATP, ADP and AMP hydrolysis when the parasites were grown in the presence of penicillin and streptomycin. The existence of an NTPDase activity in T. gallinae may be involved in pathogenicity, protecting the parasite from the cytolytic effects of the extracellular nucleotides.     

Comparative structural analyses of purified glycogen particles from rat liver,human skeletal muscle and commercial preparations

Glycogen is a cellular energy store that is crucial for whole body energy metabolism, metabolic regulation and exercise performance. To understand glycogen structure we have purified glycogen particles from rat liver and human skeletal muscle tissues and compared their biophysical properties with those found in commercial glycogen preparations. Ultrastructural analysis of commercial liver glycogens fails to reveal the classical α-rosette structure but small irregularly shaped particles. In contrast, commercial slipper limpet glycogen consists of β-particles with similar branching and chain lengths to purified rat liver glycogen together with a tendency to form small α-particles, and suggest it should be used as a source of glycogen for all future studies requiring a substitute for mammalian liver glycogen.     

The Apollo Number: Space Suits,Self-Support,and the Walk-Run Transition

Background

How space suits affect the preferred walk-run transition is an open question with relevance to human biomechanics and planetary extravehicular activity. Walking and running energetics differ; in reduced gravity (<0.5 g), running, unlike on Earth, uses less energy per distance than walking.

Methodology/Principal Findings

The walk-run transition (denoted *) correlates with the Froude Number (Fr = v²/gL, velocity v, gravitational acceleration g, leg length L). Human unsuited Fr* is relatively constant (∼0.5) with gravity but increases substantially with decreasing gravity below ∼0.4 g, rising to 0.9 in 1/6 g; space suits appear to lower Fr*. Because of pressure forces, space suits partially (1 g) or completely (lunar-g) support their own weight. We define the Apollo Number (Ap = Fr/M) as an expected invariant of locomotion under manipulations of M, the ratio of human-supported to total transported mass. We hypothesize that for lunar suited conditions Ap* but not Fr* will be near 0.9, because the Apollo Number captures the effect of space suit self-support. We used the Apollo Lunar Surface Journal and other sources to identify 38 gait events during lunar exploration for which we could determine gait type (walk/lope/run) and calculate Ap. We estimated the binary transition between walk/lope (0) and run (1), yielding Fr* (0.36±0.11, mean±95% CI) and Ap* (0.68±0.20).

Conclusions/Significance

The Apollo Number explains 60% of the difference between suited and unsuited Fr*, appears to capture in large part the effects of space suits on the walk-run transition, and provides several testable predictions for space suit locomotion and, of increasing relevance here on Earth, exoskeleton locomotion. The knowledge of how space suits affect gait transitions can be used to optimize space suits for use on the Moon and Mars.     

The 5-HT(1A) receptor agonist 8-OH-DPAT prevents prefrontocortical glutamate and serotonin release in response to blockade of cortical NMDA receptors

We studied the role of 5-HT(1A) receptors in controlling the release of glutamate (GLU) in the medial prefrontal cortex (mPFC) of conscious rats with the in vivo microdialysis technique. The effect of the 5-HT(1A) receptor agonist 8-hydroxy-2-(di-n-propylamino)tetralin infused in the prefrontal cortex was examined under basal conditions and on the rise of extracellular GLU (+106%) induced by co-infusion of the competitive N-methyl-d-aspartate receptor antagonist 3-[(R)-2-carboxypiperazin-4yl]-propyl-1-phosphonic acid (CPP). 8-OH-DPAT (0.3 and 3 microm) had no effect on basal extracellular GLU, but the higher concentration completely abolished the rise of extracellular GLU induced by CPP. CPP also increased extracellular serotonin (5-HT) in the mPFC (+50%) and this effect was antagonized by 3 microm 8-OH-DPAT which, by itself, had no effect on basal 5-HT release. The effects of 8-OH-DPAT on extracellular GLU and 5-HT were reversed by the 5-HT(1A) receptor antagonist WAY100 635 (100 microm), indicating a selective involvement of 5-HT(1A) receptors. WAY100 635 had no effect by itself. These results show that the stimulation of cortical 5-HT(1A) receptors prevents the CPP-evoked rise of extracellular GLU and 5-HT and suggest that these effects may contribute to the ability of intracortical 8-OH-DPAT to counteract cognitive deficits caused by the blockade of NMDA receptors.     

Neonatal exposure to UVR alters skin immune system development, and suppresses immunity in adulthood

Neonates have a developing immune response, with a predisposition towards induction of tolerance. As the immune system develops, immunity rather than tolerance is induced, with this development of immunity occurring in response to external factors such as the environment. As ultraviolet radiation (UVR) suppresses immunity, it is likely that the effect of UVR on the neonatal immune system would be augmentation of the suppressive response. In support, childhood exposure to UVR has been linked with an increased incidence of melanoma; consistent with an increase in suppression. To address this, phenotypic and functional immune system studies were undertaken at 8 weeks after one single exposure of solar-simulated UVR to mice, when mice had reached adulthood. Subtle changes were observed in cell populations resident in the skin-draining lymph nodes (LNs) and there also appeared to be a subtle, but not statistically significant, increase in the production of interleukin-10 and interferon-γ. Importantly, these changes also corresponded with significant suppression of the contact hypersensitivity response in irradiated mice compared with their control counterparts. This suppression was apparent when antigen sensitisation occurred during the neonatal or adult period, and thus did not appear to be analogous to UVR-induced suppression in adults. Although the percentage of T regulatory cells was increased in the skin-draining LNs, they were induced in a different manner to those induced following adult UVR exposure, with no increase in function on a per-cell basis. It therefore appears that one single neonatal exposure to UVR alters development of the immune system, leading to long-term implications for induction of immunity.     

Genome-wide studies of copy number variation and exome sequencing identify rare variants in BAG3 as a cause of dilated cardiomyopathy

Dilated cardiomyopathy commonly causes heart failure and is the most frequent precipitating cause of heart transplantation. Familial dilated cardiomyopathy has been shown to be caused by rare variant mutations in more than 30 genes but only ~35% of its genetic cause has been identified, principally by using linkage-based or candidate gene discovery approaches. In a multigenerational family with autosomal dominant transmission, we employed whole-exome sequencing in a proband and three of his affected family members, and genome-wide copy number variation in the proband and his affected father and unaffected mother. Exome sequencing identified 428 single point variants resulting in missense, nonsense, or splice site changes. Genome-wide copy number analysis identified 51 insertion deletions and 440 copy number variants > 1 kb. Of these, a 8733 bp deletion, encompassing exon 4 of the heat shock protein cochaperone BCL2-associated athanogene 3 (BAG3), was found in seven affected family members and was absent in 355 controls. To establish the relevance of variants in this protein class in genetic DCM, we sequenced the coding exons in BAG3 in 311 other unrelated DCM probands and identified one frameshift, two nonsense, and four missense rare variants absent in 355 control DNAs, four of which were familial and segregated with disease. Knockdown of bag3 in a zebrafish model recapitulated DCM and heart failure. We conclude that new comprehensive genomic approaches have identified rare variants in BAG3 as causative of DCM.     

Hypercholesterolemia increases mitochondrial oxidative stress and enhances the MPT response in the porcine myocardium: beneficial effects of chronic exercise

Hypercholesterolemia has been suggested to have direct negative effects on myocardial function due to increased reactive oxygen species (ROS) generation and increased myocyte death. Mitochondrial permeability transition (MPT) is a significant mediator of cell death, which is enhanced by ROS generation and attenuated by exercise training. The purpose of this study was to investigate the effect of hypercholesterolemia on the MPT response of cardiac mitochondria. We tested the hypothesis that familial hypercholesterolemic (FH) pigs would have an enhanced MPT response and that exercise training could reverse this phenotype. MPT was assessed by mitochondrial swelling in response to 10-100 μM Ca(2+). FH pigs did show an increased MPT response to Ca(2+) that was associated with decreases in the expression of the putative MPT pore components mitochondrial phosphate carrier (PiC) and cyclophilin-D (CypD). FH also caused increased oxidative stress, depicted by increased protein nitrotyrosylation, as well as decreased levels of reduced GSH in cardiac mitochondria. Expression of the mitochondrial antioxidant enzymes manganese superoxide dismutase (MnSOD), thioredoxin-2 (Trx2), and peroxiredoxin-3 (Prx3) was greatly reduced in the FH pigs. In contrast, cytosolic catalase expression and activity were increased. However, chronic exercise training was able to normalize the MPT response in FH pigs, reduce mitochondrial oxidative stress, and return MnSOD, Trx2, Prx3, and catalase expression/activities to normal. We conclude that FH reduces mitochondrial antioxidants, increases mitochondrial oxidative stress, and enhances the MPT response in the porcine myocardium, and that exercise training can reverse these detrimental alterations.     

Assembly and plasticity of the glutamatergic postsynaptic specialization

Glutamate mediates most excitatory synaptic transmission in the brain. Synaptic strength at glutamatergic synapses shows a remarkable degree of use-dependent plasticity and such modifications may represent a physiological correlate to learning and memory. Glutamate receptors and downstream enzymes are organized at synapses by cytoskeletal proteins containing multiple protein-interacting domains. Recent studies demonstrate that these 'scaffolding' proteins within the postsynaptic specialization have the capacity to promote synaptic maturation, influence synapse size, and modulate glutamate receptor function.     

Leachate from a Waste Disposal Centre reduces the initiation of arbuscular mycorrhiza,and spread of hyphae in soil

Irrigating arbuscular mycorrhizal plants with leachate from a waste disposal centre was examined in a set of experiments. Application of leachate at half the average concentration found in the field or higher reduced the growth of the host plant, the initiation of mycorrhizal colonisation, and hyphal growth of the mycorrhizal fungus Glomus pellucidum through the soil. Causes of these reductions are complex and interrelated, and associated with the electrical conductivity of the solution, toxicity of specific ions, in particular NH₄ ⁺, NH₃ and HCO₃ ⁻, and changes in the pH causing shifts in the equilibria of the soil solution. This revised version was published online in June 2006 with corrections to the Cover Date.