bluuebunny/crossref_metadata_2025 huggingface dataset:
Mammalian K-ras2 Is a Corticosteroid-Induced Gene in Vivo
Authors: Brennan Francine E. , Fuller Peter J.
Published: March 2006

Aldosterone acts via the mineralocorticoid receptor to regulate gene expression. A number of aldosterone-induced genes have been characterized in the distal colon and/or the distal nephron. Using the Xenopus kidney-derived A6 cell line, the K-ras transcript of the K-ras gene was identified as aldosterone induced, with a role in epithelial sodium transport. This study sought to establish whether K-ras expression is also increased in mammalian epithelia in vivo in response to aldosterone. RNA was extracted from the kidney and distal colon of rats treated with aldosterone or dexamethasone. Northern blot analysis and real-time RT-PCR were performed using probes and primers specific for the K-rasA isoform and for total K-ras. The expression of both total K-ras and of the A isoform is induced in the distal colon by aldosterone and by dexamethasone. Given the relative abundances of the two isoforms, this would appear to indicate induction of both isoforms. The time course of the response is consistent with a primary transcriptional response. In contrast to the documented up-regulation in the amphibian kidney, we did not observe regulation by corticosteroids in the kidney. However, regulation in a subpopulation of cells cannot be excluded.
Paracrine and Autocrine Regulation of Epidermal Growth Factor-Like Factors in Cumulus Oocyte Complexes and Granulosa Cells: Key Roles for Prostaglandin Synthase 2 and Progesterone Receptor
Authors: Shimada Masayuki , Hernandez-Gonzalez Inmaculada , Gonzalez-Robayna Ignacio , Richards JoAnne S.
Published: March 2006

The molecular bridges that link the LH surge with functional changes in cumulus cells that possess few LH receptors are being unraveled. Herein we document that epidermal growth factor (EGF)-like factors amphiregulin (Areg), epiregulin (Ereg), and betacellulin (Btc) are induced in cumulus oocyte complexes (COCs) by autocrine and paracrine mechanisms that involve the actions of prostaglandins (PGs) and progesterone receptor (PGR). Areg and Ereg mRNA and protein levels were reduced significantly in COCs and ovaries collected from prostaglandin synthase 2 (Ptgs2) null mice and Pgr null (PRKO) mice at 4 h and 8 h after human chorionic gonadotropin, respectively. In cultured COCs, FSH/forskolin induced Areg mRNA within 0.5 h that peaked at 4 h, a process blocked by inhibitors of p38MAPK (SB203580), MAPK kinase (MEK) 1 (PD98059), and PTGS2 (NS398) but not protein kinase A (PKA) (KT5720). Conversely, AREG but not FSH induced Ptsg2 mRNA at 0.5 h with peak expression of Ptgs2 and Areg mRNAs at 4 h, processes blocked by the EGF receptor tyrosine kinase inhibitor AG1478 (AG), PD98059, and NS398. PGE2 reversed the inhibitory effects of AG on AREG-induced expression of Areg but not Ptgs2, placing Ptgs2 downstream of EGF-R signaling. Phorbol 12-myristate 13-acetate (PMA) and adenovirally expressed PGRA synergistically induced Areg mRNA in granulosa cells. In COCs, AREG not only induced genes that impact matrix formation but also genes involved in steroidogenesis (StAR, Cyp11a1) and immune cell-like functions (Pdcd1, Runx1, Cd52). Collectively, FSH-mediated induction of Areg mRNA via p38MAPK precedes AREG induction of Ptgs2 mRNA via ERK1/2. PGs acting via PTGER2 in cumulus cells provide a secondary, autocrine pathway to regulate expression of Areg in COCs showing critical functional links between G protein-coupled receptor and growth factor receptor pathways in ovulating follicles.
Roles of Purinergic P2X Receptors as Pacemaking Channels and Modulators of Calcium-Mobilizing Pathway in Pituitary Gonadotrophs
Authors: Zemkova Hana , Balik Ales , Jiang Yonghua , Kretschmannova Karla , Stojilkovic Stanko S.
Published: March 2006

Anterior pituitary cells release ATP and express several subtypes of purinergic P2 receptors, but their biophysical properties and roles in spontaneous and receptor-controlled electrical activity have not been characterized. Here we focused on extracellular ATP actions in gonadotrophs from embryonic, neonatal, and adult rats. In cells from all three age groups, the Ca2+-mobilizing agonist GnRH induced oscillatory, hyperpolarizing, nondesensitizing, and slow deactivating currents. In contrast, ATP induced nonoscillatory, depolarizing, slowly desensitizing, and rapidly deactivating current, indicating that these cells express cation-conducting P2X channels but not Ca2+-mobilizing P2Y receptors. The amplitudes of P2X current response and the rates of receptor desensitization were dependent on ATP concentration. The biophysical and pharmacological properties of P2X currents were consistent with the expression of P2X2 subtype of channels in these cells. ATP-induced rapid depolarization of gonadotrophs lead to initiation of firing in quiescent cells, an increase in the frequency of action potentials in spontaneously active cells, and a transient stimulation of LH release. ATP also influenced GnRH-induced current and membrane potential oscillations and LH release in an extracellular Ca2+-dependent manner. These inositol 1,4,5-triphosphate-dependent oscillations were facilitated, slowed, or stopped, depending of ATP concentration, the time of its application, and the level of Ca2+ content in intracellular stores. These results indicate that, in gonadotrophs, P2X receptors could operate as pacemaking channels and modulators of GnRH-controlled electrical activity and secretion.
Tribute to R. G. Boutilier: Skin colour and body temperature changes in baskingBokermannohyla alvarengai(Bokermann 1956)
Authors: Tattersall Glenn J. , Eterovick Paula C. , de Andrade Denis V.
Published: March 2006

In amphibians solar basking far from water sources is relatively uncommon since the highly permeable amphibian skin does not represent a significant barrier to the accompanying risk of losing water by evaporation. A South American frog, Bokermannohyla alvarengai (Bokermann 1956), however,spends a significant amount of the day exposed to full sun and relatively high temperatures. The means by which this frog copes with potentially high rates of evaporative water loss and high body temperatures are unknown. Thus, in this study, skin colour changes, body surface temperature, and evaporative water loss rates were examined under a mixture of field and laboratory conditions to ascertain whether changes in skin reflectivity play an important role in this animal's thermal and hydric balance. Field data demonstrated a tight correlation between the lightness of skin colour and frog temperature,with lighter frogs being captured possessing higher body temperatures. Laboratory experiments supported this relationship, revealing that frogs kept in the dark or at lower temperatures (20°C) had darker skin colours,whereas frogs kept in the light or higher temperatures (30°C) had skin colours of a lighter hue. Light exhibited a stronger influence on skin colour than temperature alone, suggesting that colour change is triggered by the increase in incident solar energy and in anticipation of changes in body temperature. This conclusion is corroborated by the observation that cold,darkly coloured frogs placed in the sun rapidly became lighter in colour during the initial warming up period (over the first 5 min), after which they warmed up more slowly and underwent a further, albeit slower, lightening of skin colour. Surprisingly, despite its natural disposition to bask in the sun,this species does not possess a `waterproof' skin, since its rates of evaporative water loss were not dissimilar from many hylid species that live in arboreal or semi-aquatic environments. The natural history of B. alvarengai is largely unknown and, therefore, it is likely that the herein reported colour change and basking behaviour represent a complex interaction between thermoregulation and water balance with other ecologically relevant functions, such as crypsis.
Tribute to R. G. Boutilier: Acid–base transfer across fish gills
Authors: Randall D. J. , Tsui T. K. N.
Published: March 2006

The gills are the major site of acid–base regulation in most fish. Acid–base transfer across fish gills is dominated by carbon dioxide and ammonia excretion, especially the former. Bicarbonate buffering in the blood is less than that found in mammals; regulation of ventilation has little effect on CO2 levels in the blood and control of ventilation is not used to regulate body pH in fish. Proton ATPase (freshwater fish),Na+/H+ exchangers (marine fish) and anion exchangers(marine and freshwater fish) are located in the gills. These transporters contribute to the regulation of internal pH, but little is known about how this is done in fish. Fish kept in confined water volumes acidify their environment, largely due to CO2. This acidification augments ammonia excretion and reduces ammonia toxicity. The possible involvement of ammonia recycling in acid excretion is also discussed.
Tribute to R. G. Boutilier: The role for skeletal muscle in the hypoxia-induced hypometabolic responses of submerged frogs
Authors: West T. G. , Donohoe P. H. , Staples J. F. , Askew G. N.
Published: March 2006

Much of Bob Boutilier's research characterised the subcellular, organ-level and in vivo behavioural responses of frogs to environmental hypoxia. His entirely integrative approach helped to reveal the diversity of tissue-level responses to O2 lack and to advance our understanding of the ecological relevance of hypoxia tolerance in frogs. Work from Bob's lab mainly focused on the role for skeletal muscle in the hypoxic energetics of overwintering frogs. Muscle energy demand affects whole-body metabolism, not only because of its capacity for rapid increases in ATP usage, but also because hypometabolism of the large skeletal muscle mass in inactive animals impacts so greatly on in vivo energetics. The oxyconformance and typical hypoxia-tolerance characteristics (e.g. suppressed heat flux and preserved membrane ion gradients during O2 lack) of skeletal muscle in vitro suggest that muscle hypoperfusion in vivo is possibly a key mechanism for (i) downregulating muscle and whole-body metabolic rates and (ii) redistributing O2 supply to hypoxia-sensitive tissues. The gradual onset of a low-level aerobic metabolic state in the muscle of hypoxic, cold-submerged frogs is indeed important for slowing depletion of on-board fuels and extending overwintering survival time. However, it has long been known that overwintering frogs cannot survive anoxia or even severe hypoxia. Recent work shows that they remain sensitive to ambient O2 and that they emerge rapidly from quiescence in order to actively avoid environmental hypoxia. Hence, overwintering frogs experience periods of hypometabolic quiescence interspersed with episodes of costly hypoxia avoidance behaviour and exercise recovery.

In keeping with this flexible physiology and behaviour, muscle mechanical properties in frogs do not deteriorate during periods of overwintering quiescence. On-going studies inspired by Bob Boutilier's integrative mindset continue to illuminate the cost–benefit(s) of intermittent locomotion in overwintering frogs, the constraints on muscle function during hypoxia, the mechanisms of tissue-level hypometabolism, and the details of possible muscle atrophy resistance in quiescent frogs.
Cardiorespiratory modifications, and limitations, in post-smolt growth hormone transgenic Atlantic salmonSalmo salar
Authors: Deitch E. J. , Fletcher G. L. , Petersen L. H. , Costa I. A. S. F. , Shears M. A. , Driedzic W. R. , Gamperl A. K.
Published: March 2006

In recent years, there has been a great deal of interest in how growth hormone (GH) transgenesis affects fish physiology. However, the results of these studies are often difficult to interpret because the transgenic and non-transgenic fish had very different environmental/rearing histories. This study used a stable line of size-matched GH Atlantic salmon (Salmo salar) that were reared in a shared tank with controls (at 10°C, for∼9 months) to perform a comprehensive examination of the cardiorespiratory physiology of GH transgenic salmon, and serves as a novel test of the theory of symmorphosis. The GH transgenic salmon had a 3.6× faster growth rate,and 21 and 25% higher values for mass-specific routine and standard oxygen consumption (ṀO2),respectively. However, there was no concurrent increase in their maximum ṀO2, which resulted in them having an 18% lower metabolic scope and a 9% reduction in critical swimming speed. This decreased metabolic capacity/performance was surprising given that the transgenics had a 29% larger heart with an 18% greater mass-specific maximum in situ cardiac output, a 14% greater post-stress blood haemoglobin concentration, 5–10% higher red muscle and heart aerobic enzyme (citrate synthase or cytochrome oxidase) activities, and twofold higher resting and 1.7× higher post-stress, catecholamine levels. However, gill surface area was the only cardiorespiratory parameter that was not enhanced, and our data suggest that gill oxygen transfer may have been limiting. Overall, this research: (1) shows that there are significant metabolic costs associated with GH transgenesis in this line of Atlantic salmon; (2) provides the first direct evidence that cardiac function is enhanced by GH transgenesis; (3) shows that a universal upregulation of post-smolt (adult) GH transgenic salmon cardiorespiratory physiology, as suggested by symmorphosis, does not occur; and (4) supports the idea that whereas differences in arterial oxygen transport (i.e. cardiac output and blood oxygen carrying capacity) are important determinants of inter-specific differences in aerobicity, diffusion-limited processes must be enhanced to achieve substantial intra-specific improvements in metabolic and swimming performance.
Tribute to R. G. Boutilier: Evidence of a high activity carbonic anhydrase isozyme in the red blood cells of an ancient vertebrate, the sea lamprey Petromyzon marinus
Authors: Esbaugh A. J. , Tufts B. L.
Published: March 2006

Carbonic anhydrase (CA) is a multi-functional enzyme that catalyzes the hydration/dehydration of carbon dioxide. In the red blood cell (rbc), CA is necessary to facilitate the transport of carbon dioxide out of the body. Results from earlier biochemical studies indicate that ancient vertebrates,such as agnathans, possess a low activity rbc CA isozyme, whereas more recently evolved vertebrates, such as teleost fish, possess a high activity isozyme. At present, however, the changes in the molecular structure that have resulted in this large increase in catalytic efficiency are unknown. The objective of the current study was therefore to determine the molecular structure of rbc CA in lampreys and compare it to that of teleosts in an effort to ascertain how this important enzyme became more efficient over evolutionary time. Isolation and sequencing of cytoplasmic CA from rbc and gill showed only a single isozyme of 789 bp (262 amino acids). This isozyme was also found in brain and kidney, with no evidence of additional cytoplasmic CA isozymes in other tissues. Phylogenetic analysis grouped this isozyme closely to vertebrate CA VII, which is ancestral to the rbc isozymes in other vertebrates. Interestingly, active site analysis revealed a structure similar to high activity isozymes. A comparative kinetic analysis of CA from rbc lysates and CA fusion proteins showed that the traditional method of determining the turnover number may not be appropriate for all vertebrate CAs. In contrast to previous evidence, lamprey CA was found to be a high activity isozyme. These results suggest that the critical functional characteristics of rbc CA have been highly conserved throughout vertebrate evolution.
Significance of a basal melanin layer to production of non-iridescent structural plumage color: evidence from an amelanotic Steller's jay(Cyanocitta stelleri)
Authors: Shawkey Matthew D. , Hill Geoffrey E.
Published: March 2006

Non-iridescent structural plumage color is typically produced by coherent scattering of light within a matrix of keratin and air (a `spongy layer') in feather barbs. It remains unclear what role, if any, the basal melanin layer underlying this spongy layer plays in the production of coloration. Amelanism in birds with structural color is a `natural experiment' in which melanin pigmentation is lost, allowing us to identify the effects of the loss of melanin on structural color production. Here we use full-spectrum spectrometry, transmission electron microscopy and Fourier analysis to compare the color and nanostructure of an amelanotic Steller's jay (Cyanocitta stelleri Gmelin) feather with a normal blue Steller's jay feather. As a control, we also examined a white domestic chicken (Gallus gallusLinnaeus) feather. The pale amelanotic jay feather had a broad reflectance curve with a blue/green peak, while the typical blue feather had a typical distinct bell-shaped blue curve with a UV/violet peak. The white chicken feather had a typical white reflectance curve with no discrete peaks. Electron microscopy revealed that both the amelanotic and blue feather barbs contained well-formed spongy layers that were of the correct size and arrangement to produce their measured peak reflectance values, whereas the chicken feather had no spongy layer. The washed-out color of the amelanotic jay feather was thus most probably caused by the loss of the basal melanin layer, suggesting that melanin functions to absorb incoherently scattered white light from the feather barb thereby increasing the purity of the color produced by the spongy layer.
Interaction between non-specific electrostatic forces and humoral factors in haemocyte attachment and encapsulation in the edible cockle,Cerastoderma edule
Authors: Wootton Emma C. , Dyrynda Elisabeth A. , Ratcliffe Norman A.
Published: March 2006

In invertebrates, encapsulation is the common immune defence reaction towards foreign bodies, including multicellular parasites, which enter the haemocoel and are too large to be phagocytosed. This immune response has been most extensively studied in insects, in which it is highly complex, involving a diversity of cellular and molecular processes, but little is known of this process in bivalve molluscs. Non-specific physicochemical properties are known to influence parasite–haemocyte interactions in many invertebrates, and these may provide the common basis of encapsulation on which highly specific biochemical interactions are imposed. The present study uses synthetic beads and thread to mimic inactive metacercarial cysts of trematodes, and thus investigates factors involved in the basic, non-specific mechanisms of cell attachment and encapsulation in the edible cockle, Cerastoderma edule. Results showed that positively charged targets stimulated the most vigorous response, and further detailed experiments revealed that non-specific electrostatic forces and humoral plasma factors have a synergistic role in haemocyte attachment and the encapsulation response of C. edule.
Effect of osmotic stress on expression of a putative facilitative urea transporter in the kidney and urinary bladder of the marine toad,Bufo marinus
Authors: Konno Norifumi , Hyodo Susumu , Matsuda Kouhei , Uchiyama Minoru
Published: March 2006

Anuran amphibians accumulate a large amount of urea in their extracellular fluids to avoid a severe dehydration under dry and hyper-saline environments. To clarify the mechanisms of urea retention, we examined structure and distribution of the urea transporter (UT) in the kidney of the marine toad(Bufo marinus), and its expression in the kidney and urinary bladder following exposure to dry and hyper-saline conditions by means of cDNA cloning, semi-quantitative RT-PCR, immunoblot analysis and immunohistochemistry. The Bufo UT cDNA cloned from the kidney encodes a 390-amino-acid residue protein, which is 80% identical to Rana esculenta UT with the functional characteristics of a urea transporter. The Bufo UT mRNA was abundantly expressed in the kidney and urinary bladder, but not in the skin. In immunoblot analysis using a specific antibody raised against the Bufo UT, a 52 kDa protein similar to the glycosylated forms of mammalian UT-A2 (∼55 kDa) was detected in extracts from plasma membrane fractions of the kidney and urinary bladder. When toads were acclimated to dry and hyper-saline environments for 7 days, UT mRNA expression was upregulated in the kidney and urinary bladder and there was an elevated plasma urea concentration and osmolality. Immunohistochemistry showed that the UT was specifically localized on the apical membrane of the early distal tubule, known to be the diluting segment, in the kidney and the epithelial cells of urinary bladder. Immunoreactive cells were not detected along the late distal tubule, the connecting tubule or the collecting duct in the kidney. The present findings suggest that the Bufo UT probably contributes to urea transport in the kidney and urinary bladder in response to hyperosmotic stresses such as body fluid hypertonicity and dehydration.
The effects of social experience on the behavioral response to unexpected touch in crayfish
Authors: Song Cha-Kyong , Herberholz Jens , Edwards Donald H.
Published: March 2006

Crayfish fight and form a dominance hierarchy characterized by a pattern of repeated agonistic interactions between animals with a consistent outcome of winner and loser. Once a dominance hierarchy is established, dominant animals display an elevated posture with both claws held laterally and forward,whereas subordinate animals display a more prone posture with both claws extended forward and down. Dominant animals behave aggressively towards the subordinate opponent, often approaching and attacking, whereas subordinate animals behave submissively by tailflipping and retreating. To evaluate whether the differences in social behavior are accompanied by differences in responses to non-social stimuli, we exposed socially naïve and experienced crayfish (Procambarus clarkii) to an unexpected touch in different social conditions. Socially naïve animals turned to confront the source of a unilateral touch with raised claws and elevated posture. Dominant animals also turned to face the stimulus source with raised claws and elevated posture, both when tested alone and in the presence of a subordinate opponent. Subordinate animals displayed this orienting response only while separated from their dominant partners. When paired with their dominant partners, subordinates avoided the stimulus source by walking rapidly forwards or backwards. When the subordinate animals were later tested again, first while semi-separated from the dominant and later while fully separated, they displayed a mixed pattern of avoidance and orienting responses. These results indicate that the behavioral responses of subordinate crayfish to touch depend on their social status, their current social conditions and their recent social history.
Representation of behaviourally relevant information by blowfly motion-sensitive visual interneurons requires precise compensatory head movements
Authors: Kern R. , van Hateren J. H. , Egelhaaf M.
Published: March 2006

Flying blowflies shift their gaze by saccadic turns of body and head,keeping their gaze basically fixed between saccades. For the head, this results in almost pure translational optic flow between saccades, enabling visual interneurons in the fly motion pathway to extract information about translation of the animal and thereby about the spatial layout of the environment. There are noticeable differences between head and body movements during flight. Head saccades are faster and shorter than body saccades, and the head orientation is more stable between saccades than the body orientation. Here, we analyse the functional importance of these differences by probing visual interneurons of the blowfly motion pathway with optic flow based on either head movements or body movements, as recorded accurately with a magnetic search coil technique. We find that the precise head–body coordination is essential for the visual system to separate the translational from the rotational optic flow. If the head were tightly coupled to the body,the resulting optic flow would not contain the behaviourally important information on translation. Since it is difficult to resolve head orientation in many experimental paradigms, even when employing state-of-the-art digital video techniques, we introduce a `headifying algorithm', which transforms the time-dependent body orientation in free flight into an estimate of head orientation. We show that application of this algorithm leads to an estimated head orientation between saccades that is sufficiently stable to enable recovering information on translation. The algorithm may therefore be of practical use when head orientation is needed but cannot be measured.
Swim speeds and stroke patterns in wing-propelled divers: a comparison among alcids and a penguin
Authors: Watanuki Yutaka , Wanless Sarah , Harris Mike , Lovvorn James R. , Miyazaki Masamine , Tanaka Hideji , Sato Katsufumi
Published: March 2006

In diving birds, the volume and resulting buoyancy of air spaces changes with dive depth, and hydrodynamic drag varies with swim speed. These factors are important in the dive patterns and locomotion of alcids that use their wings both for aerial flight and underwater swimming and of penguins that use their wings only for swimming. Using small data-loggers on free-ranging birds diving to 20–30 m depth, we measured depth at 1 Hz and surge and heave accelerations at 32–64 Hz of four species of alcids (0.6–1.0 kg mass) and the smallest penguin species (1.2 kg). Low- and high-frequency components of the fluctuation of acceleration yielded estimates of body angles and stroke frequencies, respectively. Swim speed was estimated from body angle and rate of depth change. Brünnich's (Uria lomvia) and common(Uria aalge) guillemots descended almost vertically, whereas descent of razorbills (Alca torda), rhinoceros auklets (Cerorhinca monocerata) and little penguins (Eudyptula minor) was more oblique. For all species, swim speed during descent was within a relatively narrow range. Above depths of 20–30 m, where they were all positively buoyant, all species ascended without wing stroking. During descent, little penguins made forward accelerations on both the upstroke and downstroke regardless of dive depth. By contrast, descending alcids produced forward accelerations on both upstroke and downstroke at depths of <10 m but mainly on the downstroke at greater depths; this change seemed to correspond to the decrease of buoyancy with increasing depth. The magnitude of surge (forward)acceleration during downstrokes was smaller, and that during upstrokes greater, in little penguins than in alcids. This pattern presumably reflected the proportionally greater mass of upstroke muscles in penguins compared with alcids and may allow little penguins to swim at less variable instantaneous speeds.
How well do specialist feeders regulate nutrient intake? Evidence from a gregarious tree-feeding caterpillar
Authors: Despland Emma , Noseworthy Meghan
Published: March 2006

Nutritional regulation is a powerful mechanism used by generalist feeders to obtain the balance of nutrients they require from nutritionally diverse,perhaps unbalanced, foods. We examined nutritional regulation in a species with a narrow individual diet breadth: the forest tent caterpillar, Malacosoma disstria. Fourth instar caterpillars were provided with artificial foods consisting of different ratios of protein to digestible carbohydrate in no-choice, choice and compensatory feeding experiments. In the no-choice test, caterpillars were confined to a single food source of varying protein/carbohydrate ratio for the duration of the fourth larval stadium. Caterpillars performed best on equal-ratio and slightly protein-biased diets. Significant reductions in performance were only observed on extremely protein-or carbohydrate-biased diets. Daily consumption of the three acceptable intermediate diets was consistent with volumetric regulation, but the timing of the moult to the next instar appeared linked instead to protein intake. In the choice test, caterpillars were provided with two complementary foods, one biased toward protein and the other toward carbohydrate, for the duration of the stadium. The caterpillars fed randomly from the two food sources presented to them, except for the extremely protein-biased diet (P:C ratio of 35:7),which they avoided. The compensatory feeding experiment tested whether forest tent caterpillars deprived of either protein or digestible carbohydrate would select a food containing the deficient nutrient. Insects were conditioned on either protein-only, carbohydrate-only, protein-and-carbohydrate or no-nutrient foods, then offered a choice between protein-only and carbohydrate-only foods. Unlike previously studied generalist feeders, our caterpillars did not compensate for protein deficiency and showed only very weak evidence of compensation for carbohydrate deficiency. Forest tent caterpillars are colonial trail-laying forest folivores that are generally confined to a single host plant and hence do not experience much diversity in food nutrient ratios. We show that forest tent caterpillars do not independently regulate protein and carbohydrate intake. These findings are consistent with predictions that nutritional regulation abilities should be less important in animals with narrower diet breadths.
Requirement of the fixed end for spontaneous beating in flagella
Authors: Fujimura Miki , Okuno Makoto
Published: March 2006

It is well known that any part of a flagellum has the ability to bend. However, it is not clearly understood how flagella generate successive bending waves spontaneously. Some micromanipulation experiments have suggested that the base of the flagellum is required. By contrast, spontaneous bending waves could be generated in computer simulation work if the microtubules were tied together at one end. We hypothesized that the basal structure of flagella can only act as a tied end when the outer doublet microtubules are tightly bound together so as not to slide. We developed a new technique for introducing local inhibition at any position on the demembranated and reactivated flagellum. The flagellum maintained spontaneous beating when the local inhibition was introduced at any position on it. In addition, spontaneous beating occurred without the basal body when an artificial fixed region was introduced to the flagellum. We conclude that the axoneme, a bundle of microtubules, requires the fixed end for spontaneous beating.
Functional consequences of activity-dependent synaptic enhancement at a crustacean neuromuscular junction
Authors: Stein Wolfgang , Smarandache Carmen R. , Nickmann Melanie , Hedrich Ulrike B. S.
Published: March 2006

This study provides evidence that activity-dependent synaptic enhancement at a neuromuscular junction modifies the characteristics of force production of the receiving muscle during rhythmic motor neuron discharge patterns. Long-lasting augmentation of the excitatory junction potentials (EJPs)quickens and strengthens the muscle response to a given motor pattern. We used the muscle gm6 of the crab Cancer pagurus to study the functional consequences and temporal dynamics of facilitation and augmentation. This stomach muscle is driven by the rhythmic activity of the gastric mill central pattern generator in the stomatogastric nervous system. We tested the response of this muscle to rhythmic motor drive using a variety of gastric mill-like stimulations.

EJPs recorded in muscle gm6 were initially small but are summated and facilitated strongly with continuous stimulation. Facilitation increased with shorter interspike intervals and possessed a time constant of decay <1 s.

During gastric mill rhythms, motor neuron activity was by contrast represented by bursts of activity with intermittent pauses of several seconds. Recordings in intact animals and in the isolated nervous system showed a great variability in firing frequency and temporal distribution of motor neuron bursts. Train stimulations with various stimulus frequencies (5 Hz, 10 Hz, 20 Hz) and inter-train intervals (2 s, 4 s, 8 s, 16 s, 32 s) revealed that augmentation acted in addition to facilitation. Augmentation increased muscle EJPs during stimulations with inter-train intervals of 16 s or less. The effects of augmentation increased with shorter inter-train intervals, but were independent of stimulus frequency.

Augmentation also contributed to the electrical response of the muscle during gastric mill rhythms, which were obtained in vitro and in vivo, and was also reflected by an increase of muscle force and the slope of force development during repetitive train stimulation. We conclude that the augmentation of EJPs at the neuromuscular junction tunes the muscle response to support force production during rhythmic motor patterns.
Extracellular pH changes activate the p38-MAPK signalling pathway in the amphibian heart
Authors: Stathopoulou Konstantina , Gaitanaki Catherine , Beis Isidoros
Published: March 2006

We investigated the activation of the p38-MAPK signalling pathway during extracellular pH changes in the isolated perfused amphibian heart. Extracellular alkalosis (pH 8.5 or 9.5) maximally activated p38-MAPK within 2 min (4.17- and 3.20-fold, respectively) and this effect was reversible since the kinase phosphorylation levels decreased upon reperfusing the heart with normal Tris–Tyrode's buffer. Extracellular acidosis also activated p38-MAPK moderately, but persistently (1.65-fold, at 1 min and 1.91-fold, at 60 min). The alkalosis-induced p38-MAPK activation depended upon the Na+/H+ exchanger (NHE) and Na+/K+-ATPase, because it was abolished when the NHE inhibitors amiloride and HOE642 and the Na+/K+-ATPase inhibitor, ouabain, were used. Our studies also showed that extracellular alkalosis (pH 8.5) induced MAPKAPK2 phosphorylation (2.59-fold, 2 min) and HSP27 phosphorylation (5.33-fold, 2 min) in a p38-MAPK-dependent manner, as it was inhibited with 1 μmol l–1 SB203580. Furthermore,immunohistochemical studies of the phosphorylated forms of p38-MAPK and HSP27 revealed that these proteins were localised in the perinuclear region and dispersedly in the cytoplasm of ventricular cells during alkalosis. Finally,alkalosis induced the increase of HSP70 protein levels (1.52-fold, 5 min), but independently of p38-MAPK activation. These data indicate that the p38-MAPK signalling pathway is activated by extracellular pH changes and in the case of alkalosis this activation may have a protective role.
Kinematics of foraging dives and lunge-feeding in fin whales
Authors: Goldbogen Jeremy A. , Calambokidis John , Shadwick Robert E. , Oleson Erin M. , McDonald Mark A. , Hildebrand John A.
Published: March 2006

Fin whales are among the largest predators on earth, yet little is known about their foraging behavior at depth. These whales obtain their prey by lunge-feeding, an extraordinary biomechanical event where large amounts of water and prey are engulfed and filtered. This process entails a high energetic cost that effectively decreases dive duration and increases post-dive recovery time. To examine the body mechanics of fin whales during foraging dives we attached high-resolution digital tags, equipped with a hydrophone, a depth gauge and a dual-axis accelerometer, to the backs of surfacing fin whales in the Southern California Bight. Body pitch and roll were estimated by changes in static gravitational acceleration detected by orthogonal axes of the accelerometer, while higher frequency, smaller amplitude oscillations in the accelerometer signals were interpreted as bouts of active fluking. Instantaneous velocity of the whale was determined from the magnitude of turbulent flow noise measured by the hydrophone and confirmed by kinematic analysis. Fin whales employed gliding gaits during descent, executed a series of lunges at depth and ascended to the surface by steady fluking. Our examination of body kinematics at depth reveals variable lunge-feeding behavior in the context of distinct kinematic modes, which exhibit temporal coordination of rotational torques with translational accelerations. Maximum swimming speeds during lunges match previous estimates of the flow-induced pressure needed to completely expand the buccal cavity during feeding.
Enhanced Discrimination of Benign from Malignant Prostatic Disease by Selective Measurements of Cleaved Forms of Urokinase Receptor in Serum
Authors: Piironen Timo , Haese Alexander , Huland Hartwig , Steuber Thomas , Christensen Ib Jarle , Brünner Nils , Danø Keld , Høyer-Hansen Gunilla , Lilja Hans
Published: March 2006

Background: Early detection of prostate cancer (PCa) centers on measurements of prostate-specific antigen (PSA), but current testing practices suffer from lack of specificity and generate many unnecessary prostate biopsies. Soluble urokinase plasminogen activator receptor (uPAR) is present in blood in both intact and cleaved forms. Increased uPAR in blood is correlated with poor prognosis in various cancers, but uPAR has not been shown to be useful in PCa diagnostics. We assessed the ability of immunoassays for specific uPAR forms to discriminate PCa from benign conditions.

Methods: We measured total PSA (tPSA), free PSA (fPSA), intact uPAR [uPAR(I-III)], intact uPAR + cleaved uPAR domains II+III [uPAR(I-III) + uPAR(II-III)], and cleaved uPAR domain I [uPAR(I)] in sera from 224 men with and 166 men without PCa. We assessed differences in serum concentrations between the PCa and noncancer groups within the entire cohort and in men with tPSA concentrations of 2–10 μg/L. The diagnostic accuracy of individual analytes and analyte combinations was explored by logistic regression and ROC analyses and evaluations of sensitivity and specificity pairs.

Results: Serum uPAR(I) and uPAR(II-III) were higher in PCa than in benign disease. In men with tPSA between 2 and 10 μg/L, the combination of %fPSA with the ratio uPAR(I)/uPAR(I-III) had a greater area under the ROC curve (0.73) than did %fPSA (0.68).

Conclusions: Specific measurements of different uPAR forms in serum improve the specificity of PCa detection. The uPAR forms may therefore be complementary to PSA for PCa detection, most importantly in men with moderately increased PSA.
The Antibody Configurations of Cardiac Troponin I Assays May Determine Their Clinical Performance
Authors: James Stefan , Flodin Mats , Johnston Nina , Lindahl Bertil , Venge Per
Published: March 2006

Background: Previous studies have shown superior clinical performance of the cardiac troponin I (cTnI) assay from Beckman-Coulter Diagnostics. This assay had a unique combination of monoclonal antibodies with 2 monoclonal antibodies directed against epitopes near the NH2 terminus of the heart-specific region of troponin I. The approach has been adopted by the new cTnI assay from Abbott Diagnostics. The aim of our study was to investigate whether this approach affects the clinical performance of cTnI assays.

Methods: Cardiac troponin concentrations were measured in a random sample of patients with unstable coronary artery disease included in the GUSTO IV trial (n = 696) by the AccuTnI (Beckman-Coulter Diagnostics), Architect cTnI (Abbott Diagnostics), Immulite 2500 cTnI (Diagnostics Products Corporation), and Elecsys 2010 cTnT (Roche Diagnostics) assays and related to the 1-year mortality. The primary cutoff concentrations were based on the 99th percentile upper reference limits and an imprecision (CV) ≤10%.

Results: The sensitivities of the AccuTnI and Architect cTnI assays in identifying patients who died within 1 year were equal and were significantly higher (P <0.05) than those of the Immulite 2500 cTnI and the Elecsys cTnT assays. The concordance between the AccuTnI and Architect cTnI assays was 97%, but concordances between the Architect cTnI and the Elecsys cTnT assays were 89%–92% with more at-risk patients (P <0.01 to P <0.001) identified by the Architect cTnI assay.

Conclusions: The Architect cTnI assay has clinical performance similar to that of the AccuTnI, probably as a result of the inclusion of a monoclonal antibody against troponin I epitope 41–49 in the assay.
Rec A-independent homologous recombination induced by a putative fold-back tetraplex DNA
Authors: Shukla Arun Kumar , Roy Kunal B.
Published: March 2006

We have recently reported that a GC-rich palindromic repeat sequence presumably adopts a stable fold-back tetraplex DNA structure under supercoiling. To establish the biological significance of this structure, we inserted this sequence between two direct repeat sequences, separated by 200 bp, in a plasmid. We then investigated the effect of this sequence on homologous recombination events. Here we report that the putative fold-back DNA tetraplex structure induces homologous recombination between direct repeat sequences. Interestingly, this recombination event is independent of recA, a major driving force for homologous recombination. We think that the fold-back structure forces the repeat sequences to come into close proximity and therefore leads to strand exchange. Although triplex-induced recombination has been well documented, our results for the first time directly establish the potential of a tetraplex structure to induce recA-independent homologous recombinationin vivo. This finding might have a significant implication for site-directed gene deletion in the context of the correction of genetic defects.
Pressure- and temperature-induced unfolding studies: thermodynamics of core hydrophobicity and packing of ribonuclease A
Authors: Font Josep , Benito Antoni , Torrent Joan , Lange Reinhard , Ribó Marc , Vilanova Maria
Published: March 2006

In this work we demonstrate that heat and pressure induce only slightly different energetic changes in the unfolded state of RNase A. Using pressure and temperature as denaturants on a significant number of variants, and by determining the free energy of unfolding at different temperatures, we estimated the stability of variants unable to complete the unfolding transition owing to the experimental conditions required for pressure experiments. The overall set of results allowed us to map the contributions to stability of the hydrophobic core residues of RNase A, with the positions most critical for stability being V54, V57, I106 and V108. We also show that the stability differences can be attributed to both hydrophobic interactions and packing density with an equivalent energetic magnitude. The main hydrophobic core of RNase A is tightly packed, as shown by the small-to-large and isosteric substitutions. In addition, we found that large changes in the number of methylene groups have non-additive positive stability interaction energies that are consistent with exquisite tight core packing and rearrangements of van der Waals' interactions in the protein interior, even after drastic deleterious substitutions.
The Development of Procedures for Charpy Impact Testing
Authors: Siewert TA , McCowan CN
Published: March 2006

This paper provides a broad overview of the progress in procedural improvements for Charpy impact testing. It includes a short summary of early developments, a discussion of topics that have been the subject of recent research, and a description of the importance of direct and indirect verification procedures. The need for standard procedures was recognized soon after the test was developed, and the early discoveries help to build the framework for our current procedures. Nevertheless, even after all these years of procedure development, researchers still find the need to learn more about certain aspects of the test procedures. Recent research seems to be concentrated in several broad categories: properties of the specimen (e.g., surface finish, tolerances, and miniature sizes for special applications), the anvils and striker (e.g., radii and surface finish), and general test procedures (e.g., time to reach test temperature and suitability for cryogenic testing).
Influence of Cold Rolling Threads Before or After Heat Treatment on the Fatigue Resistance of High Strength Fine Thread Bolts for Multiple Preload Conditions
Authors: Bradley NJ , Stephens RI , Horn NJ , Gradman JJ , Arkema JM , Borgwardt CS
Published: March 2006

SI class 12.9 high strength steel bolts were used to investigate the fatigue behavior of bolt threads rolled before/after heat treatment at five different preload values. Bolts used were 3/8 UNRF-24 (fine) and preloads were taken as 1, 50, 75, 90, and 100 % of roll before heat treatment proof stress. Since proof stress was lowered 10 % for roll after heat treatment, these preloads for roll after heat treatment bolts were then actually 1.1, 55, 83, 100, and 110 % of proof stress (to keep load the same). The tests produced a range of R ratios (R=Smin/Smax) between 0.03 and 0.92. Maximum near surface residual compressive stresses, obtained via x-ray diffraction, ranged from −500 to −1000 MPa. Axial loads were applied through the nut and all fatigue failures occurred at the first thread of the nut/bolt interface with crescent shaped cracks dominating in most tests. Multiple ratchet marks (separate crack nucleation sites) occurred for roll before heat treatment bolts, while fewer or no ratchet marks, were evident for the roll after heat treatment. Scanning electron microscopy evaluation indicated all fatigue crack growth regions contained multiple fatigue facets, while final fracture regions contained ductile dimpling. Cyclic creep/ratcheting was monitored and little, or none, was observed for preload tests equal to or less than 75 %. Replication data indicated a log-normal distribution on life was very reasonable. The fatigue resistance for fine threads rolled after heat treatment with preload stresses of 1 % (R ratio less than 0.05) caused very large increases (158 %) in 107 cycles fatigue strength compared to roll before heat treatment. This is in agreement with other roll before/after low R-ratio results. The roll after heat treatment bolts when tested at the higher proof loads had 107 fatigue strengths of 69 to 30 % increase. These increases are much less than the 158 % at 1 % preload, but still significant. Constant life Haigh diagrams at 105 and 107 cycles were in qualitative agreement with VDI 2230 bolt preload guidelines.
Promoted Combustion Testing of Pure and Ceramic-Coated Metals in High Pressure Oxygen by the Federal Institute of Materials Research and Testing (BAM)
Authors: Binder C , Kasch T , Brock T , Beck U , Weise M , Share M
Published: March 2006

The new promoted ignition combustion test apparatus at BAM matches the requirements of ASTM test method G 124 [ASTM G 124, Standard Test Method for Determining the Combustion Behavior of Metallic Materials in Oxygen-Enriched Atmospheres, 2003] and of EIGA publication, IGC Doc 13/02/E [EIGA Gas Association Document IGC Doc 13/02/E, Oxygen Pipeline Systems, 2003]. The test apparatus is most likely the only one in Europe. The test chamber allows investigations with pure oxygen or oxygen mixtures at pressures up to 500 bar and at temperatures up to 400°C at static or flowing gas conditions. The ignition device consists of an electrical ignition wire and a promoter. IR-sensors, distributed over the whole length of the rod, measure and record the burning temperature and allow measurement of the burn rate. A video system records the experiment for visual characterization of the burning behavior. BAM test results are compared with published data. Results on investigations with coated rods are presented. Ceramic coatings seem to be a solution for the use of certain less compatible metallic materials at higher oxygen pressures.