bluuebunny/crossref_metadata_2025 huggingface dataset:
Electrophysiological Analysis of the Neurotoxic Action of a Funnel-Web Spider Toxin, δ-Atracotoxin-HV1a, on Insect Voltage-Gated Na+ Channels
Authors: Grolleau Françoise , Stankiewicz Maria , Birinyi-Strachan Liesl , Wang Xiu-Hong , Nicholson Graham M. , Pelhate Marcel , Lapied Bruno
Published: April 2021

The effects of δ-ACTX-Hv1a, purified from the venom of the funnel-web spider Hadronyche versuta, were studied on the isolated giant axon and dorsal unpaired median (DUM) neurones of the cockroach Periplaneta americana under current- and voltage-clamp conditions using the double oil-gap technique for single axons and the patch-clamp technique for neurones. In parallel, the effects of the toxin were investigated on the excitability of rat dorsal root ganglion (DRG) neurones. In both DRG and DUM neurones, δ-ACTX-Hv1a induced spontaneous repetitive firing accompanied by plateau potentials. However, in the case of DUM neurones, plateau action potentials were facilitated when the membrane was artificially hyperpolarized. In cockroach giant axons, δ-ACTX-Hv1a also produced plateau action potentials, but only when the membrane was pre-treated with 3-4 diaminopyridine. Under voltage-clamp conditions, δ-ACTX-Hv1a specifically affected voltage-gated Na+ channels in both axons and DUM neurones. Both the current/voltage and conductance/ voltage curves of the δ-ACTX-Hv1a-modified inward current were shifted 10 mV to the left of control curves. In the presence of δ-ACTX-Hv1a, steady-state Na+ channel inactivation became incomplete, causing the appearance of a non-inactivating component at potentials more positive than −40 mV. The amplitude of this non-inactivating component was dependent on the holding potential. From this study, it is concluded that, in insect neurones, δ-ACTX-Hv1a mainly affects Na+ channel inactivation by a mechanism that differs slightly from that of scorpion α-toxins.
Passive Absorption of Hydrophilic Carbohydrate Probes by the House Sparrow Passer Domesticus
Authors: Chediack J. G. , Caviedes-Vidal E. , Karasov W. H. , Pestchanker M.
Published: April 2021

To evaluate the permeability of the intestine of the house sparrow Passer domesticus to hydrophilic compounds, we applied a pharmacokinetic technique to measure in vivo absorption of two carbohydrate probes, L-arabinose and D-mannitol. Probes were fed or injected, and blood and excreta were subsequently collected and analyzed by gas chromatography/mass spectrometry. Following injection, plasma probe concentration decreased in a log-linear fashion, implying single-compartment, first-order kinetics. Following oral administration, plasma probe concentrations increased, reached a maximum at 10 min and then decreased in log-linear fashion. Mannitol and arabinose absorption were calculated from the areas under the post-absorption plasma curve and the respective distribution spaces and elimination constants. The amounts absorbed increased linearly with the concentration administered (range 1–1000 mmol l−1 ), implying a passive process. The mouth-to-cloaca retention time of digesta, measured using the non-absorbable compound potassium ferrocyanide, was independent of probe concentration. On average, 69 % of the oral dose of probe was absorbed and this was independent of the concentration of probe administered. This paper supports an earlier report of substantial passive glucose absorption in house sparrows and offers a method to study the extent of hydrophilic solute absorption, which has importance for future research in areas as diverse as biomedical, ecological and evolutionary physiology.
Anaerobic Sulfur Metabolism in Thiotrophic Symbioses
Authors: Arndt C. , Gaill F. , Felbeck H.
Published: April 2021

Hydrogen sulfide is generally accepted to be the energy source for the establishment of sulfur-oxidizing symbiotic communities. Here, we show that sulfur-storing symbioses not only consume but also produce large amounts of hydrogen sulfide. The prerequisite for this process appears to be the absence of oxygen. Anaerobic sulfide production is widespread among different thiotrophic symbioses from vent and non-vent sites (Riftia pachyptila, Calyptogena magnifica, Bathymodiolus thermophilus, Lucinoma aequizonata and Calyptogena elongata). The extent of H2S generation correlates positively with the amount of elemental sulfur stored in the symbiont-bearing tissues of the hosts. Sulfide production starts a few hours after anoxia sets in, with H2S initially accumulating in the circulatory system before it is excreted into the surrounding environment. We propose that not sulfate but the elemental sulfur deposited in the symbionts serves as a terminal electron acceptor during anoxia and is reduced to sulfide. In anoxia-tolerant symbioses such as L. aequizonata, anaerobic sulfur respiration may be important for producing maintenance energy to help the species survive several months without oxygen. The increased levels of cysteine in the gills of L. aequizonata may be caused by a lack of reoxidation due to the absence of oxygen.
Magnetic Compass Orientation in the Blind Mole Rat Spalax Ehrenbergi
Authors: Kimchi Tali , Terkel Joseph
Published: April 2021

The blind mole rat Spalax ehrenbergi is a solitary, subterranean rodent that digs and inhabits a system of branching tunnels, with no above-ground exits, which it never leaves unless forced to. To survive, the mole rat must be able to orient efficiently in its tunnel system. The sensory channels available for spatial orientation in the subterranean environment are restricted in comparison with those existing above ground. This study examined the possibility that the mole rat is able to perceive and use the earth’s magnetic field to orient in space. Experiments were performed using a device constructed from a pair of electromagnetic ‘Helmholtz coils’, which create a magnetic field whose direction and strength can be altered. In the first experiment, we tested a group of mole rats (N=33) in an eight-armed maze under the earth’s natural magnetic field to determine whether they have directional preferences for the location of their sleeping nest, food chamber and toilet site. A second group of mole rats (N=30) was tested for their directional preference after the earth’s magnetic field had been experimentally shifted by 180 °. We found that the first group exhibited a significant preference (P<0.001) to build both their sleeping nest and their food store in the southern sector of the maze, whereas the second group shifted the location of their nests (P<0.01) and food store (P<0.05), to the northern sector of the maze, corresponding to the shift in the magnetic field. In the second experiment, we tested whether the magnetic compass orientation found in the first experiment depends on a light stimulus by testing a group of mole rats in the eight-armed maze under total darkness. No significant difference in directional preference between light and dark test conditions was observed. It can be concluded, therefore, that, in contrast to some amphibians and birds, magnetic compass orientation in the mole rat is independent of light stimulation.

In the third experiment, we examined whether mole rats (N=24) use the earth’s magnetic field as a compass cue to orient in a labyrinth. In the first stage (trials 1–13), the animals were trained to reach a goal box at the end of a complex labyrinth until all individuals had learned the task. In the second stage (trial 14), half the trained mole rats underwent another labyrinth trial under the earth’s natural magnetic field, while the other half were tested under a magnetic field shifted by 180 °. We found a significant decrease (P<0.001) in performance of the mole rats tested under the shifted magnetic field compared with the group tested under the natural magnetic field.

The findings from these experiments prove that the mole rat is able to perceive and use the earth’s magnetic field to orient in space.
Metabolic Rate and Thermal Stability During Honeybee Foraging at Different Reward Rates
Authors: Moffatt Luciano
Published: April 2021

During honeybee foraging, the stabilization of thoracic temperature (Tth) at elevated values is necessary to meet the power requirements of flight at different air temperatures (Ta). To understand how the bee achieves thermal stability at different reward rates, the metabolic rates of undisturbed foraging bees were measured at different Ta values and different sucrose solution flow rates. Metabolic heat production, calculated from the rate of carbon dioxide production, decreased linearly from 49.7 to 23.4 mW as Ta increased from 19 to 29 °C (sucrose flow rate 1.75 μl min−1, 50 % w/w). In contrast, crop load and inspection rate remained constant. Metabolic rate displayed a linear relationship with both Ta and the logarithm of the flow rate of sucrose solution (range analyzed 0.44–13.1 μl min−1, 50 % w/w). Metabolic rate decreased by 3.13±0.52 mW (mean ± S.E.M., N=37) for every 1 °C increase in Ta and increased by 4.36±1.13 mW for a doubling in flow rate. These changes in metabolic power output might be used to achieve thermal stability during foraging. It is suggested that the foraging bee might increase its Tth in accordance with the reward rate.
A Comparative Analysis of the Evolutionary Patterning and Mechanistic Bases of Lactate Dehydrogenase Thermal Stability in Porcelain Crabs, Genus Petrolisthes
Authors: Stillman Jonathon H. , Somero George N.
Published: April 2021

The kinetic properties of orthologous homologs (orthologs) of enzymes are typically correlated with environmental temperatures in species adapted to different thermal regimes, but correlations between adaptation temperature and enzyme thermal stability are less clear. Although the thermal stability of a protein is related chiefly to its primary structure (including post-translational modification), thermal stability can also be altered by extrinsic factors present in the intracellular milieu. Here, we present a comparative analysis of the thermal stability of lactate dehydrogenase (LDH) orthologs from 22 congeneric species of porcelain crab (genera Petrolisthes and Allopetrolisthes) from a broad range of thermal habitats. Interspecific diversity of LDH stability is high: temperatures required for a 50 % loss of activity in 10 min ranged from 65 to 75.5 °C, corresponding to half-lives of less than 1 min to more than 3 h at 70 °C. Although stability is positively correlated with maximal habitat temperature in some sister taxa, phylogenetic comparative analysis incorporating all 22 species does not indicate that the interspecific diversity of LDH stability represents an adaptive response to current thermal habitats. Examination of the mechanistic bases of LDH stabilization indicates that differences in stability are related both to properties of the LDH molecule itself (intrinsic stability) and to the effects of extrinsic protein(s). Intrinsic differences were shown by the unfolding of structure during heating, as measured by circular dichroism spectroscopy. Stabilizing effects of extrinsic proteins are implied by the results of cellular fractionation experiments that removed low-molecular-mass solutes and proteins from the muscle homogenates. We conclude that the overall structural stability and functional properties of proteins can evolve independently and that in vivo protein–protein interactions can provide another means to regulate protein stability selectively.
Detection of Cl− Flux in the Apical Microenvironment of Cultured Foetal Distal Lung Epithelial Cells
Authors: Land S. C. , Collett A.
Published: April 2021

A self-referencing Cl−-selective microelectrode (Cl− SrE) was developed and used to detect changes in the direction and magnitude of the Cl− flux (JCl) from the apical region of cultured foetal distal lung epithelial cells (FDLEs) as a function of external Cl− concentration ([Cl−]e) and in response to pharmacological challenges. The technique, which is similar to that developed for other ion-selective microelectrodes, centres on the oscillation of a Cl−-selective microelectrode between known points, micrometres apart, orthogonal to the plasma membrane. Application of the Fick principle to the differential voltage obtained per excursion amplitude (the referenced signal) yields the Cl− flux (pmol cm−2 s−1). A Cl− effusion gradient was used to confirm that empirical measurements of JCl using the Cl− SrE were statistically similar to predicted flux values calculated from the fall in [Cl−] with distance from the tip of the effusion source. Apical JCl was then measured as a function of [Cl−]e from polarised FDLE cultures grown on permeable supports. At [Cl−]e<50 mmol l−1, an apical-to-basolateral (inward) flux, maximal at 400 pmol cm−2 s−1, was observed; this reverted to a continuous basolateral-to-apical (outward) flux of 203 pmol cm−2 s−1 at [Cl]e>100 mmol l−1. At [Cl−]e>100 mmol l−1, isoproterenol (basolaterally applied, 10 μmol l−1) activated a Cl− influx of 561 pmol cm−2 s−1, whereas UTP (apically applied, 100 μmol l−1) stimulated a Cl− efflux of 300 pmol cm−2 s−1. In all cases, 50–70 % of JCl was abolished by Cl− channel blockade using 10 μmol l−1 diphenylamine-2-carboxylic acid (DPC) or 5-nitro-2-(3-phenylpropylamino)benzoic acid (NPPB). We conclude that the Cl− SrE resolves a Cl− gradient in the microenvironment of the apical region of lung epithelia that varies in both direction and magnitude as a function of external [Cl−]e and in response to Cl− channel blockade and to β2 adrenoreceptor and P2Y receptor agonists.
Energetics of Terrestrial Locomotion of the Platypus Ornithorhynchus Anatinus
Authors: Fish F. E. , Frappell P. B. , Baudinette R. V. , Macfarlane P. M.
Published: April 2021

The platypus Ornithorhynchus anatinus Shaw displays specializations in its limb structure for swimming that could negatively affect its terrestrial locomotion. Platypuses walked on a treadmill at speeds of 0.19–1.08 m s−1. Video recordings were used for gait analysis, and the metabolic rate of terrestrial locomotion was studied by measuring oxygen consumption. Platypuses used walking gaits (duty factor >0.50) with a sprawled stance. To limit any potential interference from the extensive webbing on the forefeet, platypuses walk on their knuckles. Metabolic rate increased linearly over a 2.4-fold range with increasing walking speed in a manner similar to that of terrestrial mammals, but was low as a result of the relatively low standard metabolic rate of this monotreme. The dimensionless cost of transport decreased with increasing speed to a minimum of 0.79. Compared with the cost of transport for swimming, the metabolic cost for terrestrial locomotion was 2.1 times greater. This difference suggests that the platypus may pay a price in terrestrial locomotion by being more aquatically adapted than other semi-aquatic or terrestrial mammals.
Mite Not Make it Home: Tracheal Mites Reduce the Safety Margin for Oxygen Delivery of Flying Honeybees
Authors: Harrison Jon F. , Camazine Scott , Marden James H. , Kirkton Scott D. , Rozo Albert , Yang Xiaolong
Published: April 2021

Many physiological systems appear to have safety margins, with excess capacity relative to normal functional needs, but the significance of such excess capacity remains controversial. In this study, we investigate the effects of parasitic tracheal mites (Acarapis woodi) on the safety margin for oxygen delivery and flight performance of honeybees. Tracheal mites did not affect the flight metabolic rate of honeybees in normoxic (21 % oxygen) or hyperoxic (40 % oxygen) air, but did reduce their metabolic rate relative to uninfected bees when flying in hypoxic air (5 or 10 % oxygen), demonstrating that mites reduced the safety margin for tracheal oxygen delivery. The negative effects of mites on flight metabolic rate in hypoxic atmospheres were graded with the number of mites per trachea. For example, in 10 % oxygen atmospheres, flight metabolic rate was reduced by 20 % by moderate mite infection and by 40 % by severe mite infection. Thus, the safety margin for oxygen delivery in honeybees allows them to retain normal flight metabolic rate and behavior despite tracheal mite infection under most conditions. However, the reduction in tracheal gas-exchange capacity may constrain activities requiring the highest metabolic rates, such as flying in cool weather. In support of this hypothesis, bees that were unable to return to the hive during late-winter flights showed significantly higher levels of mite infection than bees that returned safely.
Effects of Water Salinity on Acid–Base Balance in Decapod Crustaceans
Authors: Whiteley N. M. , Scott J. L. , Breeze S. J. , Mccann L.
Published: April 2021

Extracellular acid–base balance in decapod crustaceans is influenced by water salinity, although the nature of this relationship is unclear. In euryhaline crabs, a decrease in salinity results in a metabolic alkalosis in the haemolymph and an increase in salinity results in a metabolic acidosis. Alterations in acid–base status by external changes in salinity are thought to be secondary to the adjustments required for ionic and osmotic regulation. In the present study, acid–base adjustments in the haemolymph of Eriocheir sinensis after transfer to 30 % sea water accompanied alterations in muscle pH and [HCO3−], as an initial acidosis coincided with an alkalosis in the leg muscle. By 48 h transfer, haemolymph pH increased as muscle pH and HCO3− declined. Haemolymph [Cl−] decreased significantly 3 h after transfer to a new steady state but haemolymph [Na+] and muscle [Na+] and [Cl−] remained unchanged. Muscle free amino acid concentration increased twofold 6 h after transfer, followed by a 2.5-fold increase in the haemolymph after 24 h. In contrast, 30 % sea water had no effect on haemolymph acid–base adjustments in the osmoconforming crab, Necora puber, which lacks ion and osmo-regulatory mechansims. Collectively these observations support the view that salinity-induced alterations in acid–base status are caused by adjustments consistent with cell volume regulation.
Ontogeny of Decapod Crustacean Hemocyanin: Effects of Temperature and Nutrition
Authors: Terwilliger Nora B. , Dumler Karen
Published: April 2021

Hemocyanin is present throughout the decapod crustacean’s life, usually as one-hexamer and two-hexamer oligomers. Hemocyanins of some decapod crustaceans undergo changes in subunit composition and oxygen affinity during development. Maternal hemocyanin is taken up from the hemolymph via endocytosis by the oocyte. Embryo hemocyanin differs in subunit composition from hemocyanin of oocyte and adult crab and may represent the onset of hemocyanin synthesis. Complex changes in expression of hemocyanin subunits occur through megalopa and early juvenile stages of the crab Cancer magister, culminating in the pattern of adult hemocyanin. The influences of food availability and temperature on development, growth and hemocyanin ontogeny in early juvenile C. magister have been studied. Crabs were raised in warm or cold sea water and fed high or low levels of food for 6 months. While intermolt period was shorter in crabs fed high food levels, especially those raised in warm water, crabs reared in cold water with high food levels attained the largest sizes. Thus increased food availability affects growth more than increased temperature. Adult hemocyanin appeared at about the same number of weeks after the start of the experiment for crabs in the warm water/high food, warm water/low food and cold water/high food groups, even though warm water/low food crabs had molted fewer times. Crabs in the cold water/low food group expressed adult hemocyanin much later than the other groups. Molt stage and maturation from juvenile to adult are not absolutely coupled, and food availability has a greater influence than temperature on hemocyanin ontogeny.
Binding of Urate and Caffeine to Haemocyanin Analysed by Isothermal Titration Calorimetry
Authors: Menze Michael A. , Hellmann Nadja , Decker Heinz , Grieshaber Manfred K.
Published: April 2021

Haemocyanin serves as an oxygen carrier in the haemolymph of decapod crustaceans. The oxygen-binding behaviour of the pigment is modulated by the two major anaerobic metabolites, L-lactate and urate. The binding of these two metabolites to haemocyanin has been investigated mainly indirectly by following the effector-induced changes in the oxygen-binding properties of the respiratory pigment. Only a few direct investigations of effector binding, employing ultracentrifugation techniques and equilibrium dialysis, have been carried out. No evidence for cooperative binding for either effector was detected using these methods. However, isothermal titration calorimetry (ITC) offers a useful tool to gain additional insight into the binding of effectors to these highly allosterically regulated macromolecules. By applying the ITC method to the fully oxygenated dodecameric haemocyanin of the lobster Homarus vulgaris, cooperativity in binding has been found for the urate analogue caffeine but not for urate itself: using urate and the urate analogue caffeine as ligands, two conformations of the oxygenated pigment were detected.
Mechanical Forces Imposed on Echinoid Eggs During Spawning: Mitigation of Forces by Fibrous Networks within Egg Extracellular Layers
Authors: Thomas Florence I. M. , Bolton Toby F. , Sastry Ann Marie
Published: April 2021

Many echinoderms spawn large numbers of eggs in rapid spawning bouts directly into the water column. During spawning, the eggs pass from the gonad along a narrow oviduct and through a rigid gonopore before reaching the water. As the eggs traverse this pathway, they are exposed to shear stress induced by the development of a velocity gradient within the fluid formed by the eggs. In some species, the diameter of the eggs is larger than the diameter of the gonopore. In these cases, the eggs also experience strain resulting from compression of the egg as it passes through the relatively small gonopore. The magnitude of shear stress experienced by eggs differs among species depending upon the viscosity of the eggs. The degree of strain experienced by eggs differs among species depending upon the relative sizes of the egg and gonopore and also changes within species as they grow. Recent evidence suggests that these forces have the propensity to damage eggs, thereby reducing their fertilizability, or to destroy them. Experimental evidence shows that the jelly coat around the eggs can protect them from shear stresses and reduce the strain imposed on them under a compressive force. Echinoderm eggs are surrounded by a jelly coat that has a composite structure of irregularly arranged fibrous networks embedded in a jelly matrix. These fibrous networks have characteristics that are similar to those of engineered and biological materials that are known to reduce the transmission of mechanical forces to other structures. Given this similarity to other materials, three possible mechanisms for the action of the jelly coat may be identified. First, the apparent viscosity of the eggs of echinoids declines as the rate at which they are sheared during spawning increases. This reduction in viscosity with increasing shear rate (shear-thinning) may, in part, be due to the structure of the jelly coat and its resultant non-Newtonian nature. Second, experimental evidence indicates that the jelly coat preferentially deforms under a compressive load, reducing the transmission of that load to the egg. Third, the jelly (but not the fibers in the coating) may deform in a nearly viscous manner. In this case, the fibers may serve to provide an elastic ‘backbone’ to the layer and remain in place to stiffen the outer layer of the egg. The composite structure of the jelly coat and the morphology of the fibrous network are likely to be critical to all these mechanisms.
Ca2+ in Specification of Vegetal Cell Fate in Early sea Urchin Embryos
Authors: Yazaki I.
Published: April 2021

In sea urchin embryos, the first specification of cell fate occurs at the fourth cleavage, when small cells (the micromeres) are formed at the vegetal pole. The fate of other blastomeres is dependent on the receipt of cell signals originating from the micromeres. The micromeres are fated to become skeletogenic cells and show the ability to induce the endoderm (the archenteron) in the neighbouring cells during the 16-to 60-cell stage. Several molecules involved in signaling pathways, i.e. Notch for mesoderm specification, bone morphogenic protein (BMP) for ectoderm specification and β-catenin for endoderm specification, are spatially and temporally expressed during development. In the micromeres, β-catenin increases and subsequently localizes to the nuclei under the regulation of TCF, a nuclear binding partner of β-catenin, until the 60-cell stage. However, the mechanisms activating these signaling substances are still unclear. In this article, I demonstrate some specific properties of the membrane and cytoplasm of micromeres including new findings on intracellular Ca2+ concentration, and propose a mechanism by which the functional micromeres are autonoumously formed. The possible roles of these in the specification of vegetal cell fate in early development are discussed.
The Pharmacology of γ-Aminobutyric Acid and Acetylcholine Receptors at the Echinoderm Neuromuscular Junction
Authors: Devlin C. Leah
Published: April 2021

This review describes the various subtypes of γ-aminobutyric acid (GABA) receptors found at the echinoderm neuromuscular junction (NMJ), based on pharmacological and physiological studies. The review focuses mainly on holothurian GABA receptors at the NMJ located between the radial nerve and longitudinal muscle of the body wall (LMBW) and compares them to GABA receptors described at other echinoderm NMJs. Since a primary action of GABA on the holothurian LMBW is to modulate contractile responses to the excitatory neurotransmitter, acetylcholine (ACh), the pharmacology of echinoderm nicotinic ACh receptors (nAChRs) and muscarinic ACh receptors (mAChRs) is also addressed.

GABA responses have been described in the asteroids, echinoids and holothuroids but not in the other echinoderm classes. Some actions of GABA on echinoderm muscle include regulation of basal tone and spontaneous rhythmic contractions and modulation of cholinergic responses. Both GABA A and B receptor subtypes are present at the echinoderm NMJ, a feature also common to the arthropods, molluscs and chordates. Echinoderm GABA A receptors may mediate the excitatory responses to GABA. The GABA A receptor antagonist bicuculline has a paradoxical effect on contractility, stimulating large protracted contractions of the LMBW. The GABA A agonist muscimol potentiates cholinergic contractions of the holothurian LMBW. Another population of GABA receptors is inhibitory and is sensitive to the GABA B agonist baclofen and GABA B antagonists phaclofen and 2-OH-saclofen. The pre- and/or postsynaptic location of the GABA A and B receptors is not currently known. The folded GABA analogue 4-cis-aminocrotonic acid has no effect on the contractility of the holothurian LMBW so GABA C receptors are probably lacking in this preparation.

Pharmacological studies have shown that distinct nAChRs and mAChRs are colocalized in numerous echinoderm muscle preparations. Most recently, nAChR agonists were used to characterize pharmacologically receptors at the holothurian LMBW that bind ACh. Nicotinic AChRs with unique pharmacological profiles are localized both pre- and postsynaptically at this NMJ, where their physiological action is to enhance muscle tone. Muscarinic agonists also have excitatory actions on the LMBW but their action is to stimulate phasic, rhythmic contractions of the muscle. The location of mAChRs at the echinoderm NMJ, however, is unknown.

Since most of the studies described in the present review have used whole-mount preparations consisting largely of a combination of muscle fibers, neurons and connective tissue, it is extremely difficult to determine pharmacologically the exact location of the various receptor subtypes. Additional electrophysiological studies on isolated neurons and muscle fibers are therefore required to clearly define extra-, pre- and/or postsynaptic sites for the receptor subtypes at the echinoderm NMJ.
Ca2+ Transport by the Sarcoplasmic Reticulum Ca2+-ATPase in sea Cucumber (Ludwigothurea Grisea) Muscle
Authors: Landeira-Fernandez Ana Maria
Published: April 2021

In muscle cells, the excitation–contraction cycle is triggered by an increase in the concentration of free cytoplasmic Ca2+. The Ca2+-ATPase present in the membrane of the sarcoplasmic reticulum (SR) pumps Ca2+ from the cytosol into this intracellular compartment, thus promoting muscle relaxation. The microsomal fraction derived from the longitudinal smooth muscle of the body wall from the sea cucumber Ludwigothurea grisea retains a membrane-bound Ca2+-ATPase that is able to transport Ca2+ mediated by ATP hydrolysis. Immunological analyses reveal that monoclonal antibodies against sarco-endoplasmic reticulum Ca2+-ATPase (SERCA1 and SERCA2a) cross-react with a 110 kDa band, indicating that the sea cucumber Ca2+-ATPase is a SERCA-type ATPase. Like the mammalian Ca2+-ATPase isoforms so far described, the enzyme also shows a high affinity for Ca2+ and ATP, has an optimum pH of approximately 7.0 and is sensitive to thapsigargin and cyclopiazonic acid, specific inhibitors of the SERCA pumps. However, unlike the mammalian SERCA isoforms, concentrations of ATP above 2 mmol l−1 inhibit Ca2+ transport, but not ATP hydrolysis, in sea cucumber vesicles, suggesting that high ATP concentrations uncouple the Ca2+-ATPase. Another unique feature observed with the sea cucumber Ca2+-ATPase is the high dependence of maximal activity on K+ or Na+. Similar activation promoted by these cations was observed with various mammalian Ca2+-ATPase preparations when they were incubated in the presence of low concentrations of sulphated polysaccharides. In control experiments, K+ and Na+ have almost no effect on Ca2+ transport, but in the presence of heparin or fucosylated chondroitin sulphate, the activity of the different mammalian Ca2+-ATPases is inhibited and they are activated by either K+ or Na+ in a manner similar to the native sea cucumber ATPase. These results led us to investigate the possible occurrence of a highly sulphated polysaccharide on vesicles from the SR of sea cucumber smooth muscle that could act as an ‘endogenous’ Ca2+-ATPase inhibitor. In fact, SR vesicles derived from the sea cucumber, but not from rabbit muscle, contain a highly sulphated polysaccharide. After extraction and purification of these polysaccharide molecules, their effect was tested on vesicles obtained from rabbit muscle. This compound inhibited Ca2+ uptake in rabbit SR vesicles, at concentrations lower than heparin, and restored the dependence on monovalent cations. These results strongly suggest that the sea cucumber Ca2+-ATPase is activated by monovalent cations because of the presence of endogenous sulphated polysaccharides.
Control of Cardiovascular Function and its Evolution in Crustacea
Authors: Mcmahon Brian R.
Published: April 2021

Work in the last decade has shown that crustacean open circulatory systems are highly efficient and controlled in a complex manner. Control occurs at several levels. Myocardial contraction is initiated in the cardiac ganglion but constantly modulated by the central nervous system, both directly via the cardioregulatory nerves and indirectly via the neurohormonal system. Heart rate and stroke volume can be controlled independently and measurements of both are needed to assess cardiac output accurately. Haemolymph outflow from many arthropod hearts is via a complex multiarterial distribution system, and the regional distribution of cardiac output is tightly controlled via cardioarterial valves at the base of each artery. These valves contain innervated muscle, and differential contraction serves to regulate the efflux of oxygenated haemolymph into a particular system. The major influence on both the evolution and control of arthropod open blood vascular systems is efficiency of oxygen uptake and delivery. This influence is illustrated by reference to a variety of crustacean and other arthropod types.
Lactate Sequestration in the Carapace of the Crayfish Austropotamobius Pallipes During Exposure in air
Authors: Jackson D. C. , Wang T. , Koldkjaer P. , Taylor E. W.
Published: April 2021

When held in air for up to 24 h, crayfish accumulated Ca2+ and Mg2+ in their haemolymph in direct proportion to raised levels of lactate. K+ levels were highly variable, with elevated levels associated with morbidity. Lactate accumulation in the haemolymph was reflected in proportional increases in lactate levels in the carapace and muscle. Pieces of carapace incubated in saline containing elevated levels of lactate accumulated lactate to up to half the dissolved concentration. Measured levels in the carapace, relative to its water content, implied that lactate accumulated in the carapace in a combined form, possibly complexed to calcium. The exoskeleton seems to provide a reserve of buffering capacity and a sink for lactate during anaerobic metabolism. A similar mechanism has been identified in pond turtles.
Novel Subcellular and Molecular Tools to Study Ca2+ Transport Mechanisms During the Elusive Moulting Stages of Crustaceans: Flow Cytometry and Polyclonal Antibodies
Authors: Wheatly Michele G. , Zhang Zhiping , Weil Jennifer R. , Rogers James V. , Stiner La’tonia M.
Published: April 2021

Our understanding of calcium homeostasis during the crustacean moulting cycle derives from research on intermoult animals that has been extrapolated to other stages. In terms of transepithelial Ca2+ flux, the more interesting stages are those surrounding ecdysis since crustaceans experience a sizeable negative calcium balance in immediate premoult and a significant positive calcium balance in immediate postmoult. These stages are elusive in the sense that larger species such as lobsters are rarely captured at this time, and smaller species such as blue crabs and crayfish are seldom synchronized in their moulting cycle. The reductionist approaches employed in cellular physiology, such as vesicle techniques, employ pooling of fresh tissues from many organisms. Examination of the elusive moulting stages requires more sensitive approaches that can utilize tissue from an individual crustacean to characterize Ca2+ pumps (Sarco/Endoplasmic Reticulum Ca2+-ATPase, SERCA; Plasma Membrane Ca2+-ATPase, PMCA) and the Na+/Ca2+ eXchanger (NCX). An emerging subcellular approach described in this paper is to use flow cytometry as a technique to monitor Ca2+ uptake into Fluo-3-loaded membrane vesicles. This paper illustrates the utility of this technique for measuring ATP-dependent Ca2+ uptake into hepatopancreatic basolateral membrane vesicles. Obstacles to progress in molecular studies have not been limited by synchronization of moulting since tissue can be snap-frozen and collected from many animals over time. Here, the problem has been the lack of specific antibodies that hybridize with the Ca2+ transporters of interest so that they can be localized within epithelia. In this paper, we introduce polyclonal antibodies raised in rabbits against crayfish SERCA, PMCA and NCX. Immunocytochemistry of SERCA in muscle, PMCA in antennal gland and NCX in heart confirms the specificity of the antibodies.
Neuroendocrine Regulation of Osmoregulation and the Evolution of Air-Breathing in Decapod Crustaceans
Authors: Morris Stephen
Published: April 2021

Gills are the primary organ for salt transport, but in land crabs they are removed from water and thus ion exchanges, as well as CO2 and ammonia excretion, are compromised. Urinary salt loss is minimised in land crabs by redirecting the urine across the gills where salt reabsorption occurs. Euryhaline marine crabs utilise apical membrane branchial Na+/H+ and Cl−/HCO3− exchange powered by a basal membrane Na+/K+-ATPase, but in freshwater crustaceans an apical V-ATPase provides for electrogenic uptake of Cl− in exchange for HCO3−. The HCO3− is provided by carbonic anhydrase facilitating CO2 excretion while NH4+ can substitute for K+ in the basal ATPase and for H+ in the apical exchange. Gecarcinid land crabs and the terrestrial anomuran Birgus latro can lower the NaCl concentration of the urine to 5 % of that of the haemolymph as it passes across the gills. This provides a filtration–reabsorption system analogous to the vertebrate kidney.

Crabs exercise hormonal control over branchial transport processes. Aquatic hyper-regulators release neuroamines from the pericardial organs, including dopamine and 5-hydroxytryptamine (5-HT), which via a cAMP-mediated phosphorylation stimulate Na+/K+-ATPase activity and NaCl uptake. Freshwater species utilise a V-ATPase, and additional mechanisms of control have been suggested. Crustacean hyperglycaemic hormone (CHH) has now also been confirmed to have effects on hydromineral regulation, and a putative role for neuropeptides in salt and water balance suggests that current models for salt regulation are probably incomplete. In a terrestrial crabs there may be controls on both active uptake and diffusive loss. The land crab Gecarcoidea natalis drinking saline water for 3 weeks reduced net branchial Na+ uptake but not Na+/K+-ATPase activity, thus implying a reduction in diffusive Na+ loss. Further, in G. natalis Na+ uptake and Na+/K+-ATPase were stimulated by 5-HT independently of cAMP. Conversely, in the anomuran B. latro, branchial Na+ and Cl− uptake and Na+/K+-ATPase are inhibited by dopamine, mediated by cAMP. There has been a multiple evolution of a kidney-type system in terrestrial crabs capable of managing salt, CO2 and NH3 movements.
A shot in the dark: the silent quest of a free-flying phonotactic fly
Authors: Müller Pie , Robert Daniel
Published: April 2021

To reproduce, females of the parasitoid fly Ormia ochracea detect and localise a calling male cricket upon which they deposit their endoparasitic larvae. Calling male crickets are therefore subject to both sexual and natural selection by simultaneously attracting mates and phonotactic parasitoids. The possible strategy of song interruption employed by the cricket host to reduce his attractiveness to acoustic parasitoids was tested in the laboratory by examining the fly’s phonotactic quest in response to synthetic cricket songs. Phonotactic flight trajectories were recorded in three dimensions with a stereo infrared video tracking system while the sound stimulus was controlled on-line as a function of the fly’s position in space. Within a single flight, three distinct phases could be observed: a take-off phase, a cruising phase, during which course and altitude were rather constant, and a landing phase characterised by a spiralling descent towards the sound source. The flies showed remarkable phonotactic accuracy in darkness; they landed at a mean distance of 8.2 cm from the centre of the loudspeaker after a flight distance of approximately 4 m. The present data illustrate the fly’s surprising ability to gauge the direction and distance of a sound source in three dimensions and, subsequently, to find it in darkness and silence.
ATP-regenerating system in the cilia of Paramecium caudatum
Authors: Noguchi Munenori , Sawadah Taketoshi , Akazawa Toshikazu
Published: April 2021

The energy supply for eukaryotic ciliary and flagellar movement is thought to be maintained by ATP-regenerating enzymes such as adenylate kinase, creatine kinase and arginine kinase. In this study, the energy-supplying system for the ciliary movement of Paramecium caudatum was examined. Arginine kinase and adenylate kinase activities were detected in the cilia. To demonstrate that phosphoarginine satisfactorily supplies high-energy phosphate compounds into the narrow ciliary space, we prepared an intact ciliated cortical sheet from live Paramecium caudatum. These cortical sheets, with an intact ciliary membrane, produced a half-closed system in which each cilium was covered with a ciliary membrane with an opening to the cell body. Ciliary beating on the intact cortical sheets was induced by perfusing not only ATP but also ADP. Addition of phosphoarginine (0.2 mmol l−1 ) increased the beat frequency. A further increase in beat frequency was observed in 0.4 mmol l−1 phosphoarginine, and this was enhanced when the cilia were reactivated with relatively low concentrations of ATP. We have demonstrated that phosphoarginine supplies energy as a ‘phosphagen’ for ciliary beating in Paramecium caudatum, suggesting that phosphoarginine functions not only as a reservoir of energy but also as a transporter of energy in these continuously energy-consuming circumstances.

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Mechanics of limb bone loading during terrestrial locomotion in the green iguana (Iguana iguana) and American alligator (Alligator mississippiensis)
Authors: Blob Richard W. , Biewener Andrew A.
Published: April 2021

In vivo measurements of strain in the femur and tibia of Iguana iguana (Linnaeus) and Alligator mississippiensis (Daudin) have indicated three ways in which limb bone loading in these species differs from patterns observed in most birds and mammals: (i) the limb bones of I. iguana and A. mississippiensis experience substantial torsion, (ii) the limb bones of I. iguana and A. mississippiensis have higher safety factors than those of birds or mammals, and (iii) load magnitudes in the limb bones of A. mississippiensis do not decrease uniformly with the use of a more upright posture. To verify these patterns, and to evaluate the ground and muscle forces that produce them, we collected three-dimensional kinematic and ground reaction force data from subadult I. iguana and A. mississippiensis using a force platform and high-speed video. The results of these force/kinematic studies generally confirm the loading regimes inferred from in vivo strain measurements. The ground reaction force applies a torsional moment to the femur and tibia in both species; for the femur, this moment augments the moment applied by the caudofemoralis muscle, suggesting large torsional stresses. In most cases, safety factors in bending calculated from force/video data are lower than those determined from strain data, but are as high or higher than the safety factors of bird and mammal limb bones in bending. Finally, correlations between limb posture and calculated stress magnitudes in the femur of I. iguana confirm patterns observed during direct bone strain recordings from A. mississippiensis: in more upright steps, tensile stresses on the anterior cortex decrease, but peak compressive stresses on the dorsal cortex increase. Equilibrium analyses indicate that bone stress increases as posture becomes more upright in saurians because the ankle and knee extensor muscles exert greater forces during upright locomotion. If this pattern of increased bone stress with the use of a more upright posture is typical of taxa using non-parasagittal kinematics, then similar increases in load magnitudes were probably experienced by lineages that underwent evolutionary shifts to a non-sprawling posture. High limb bone safety factors and small body size in these lineages could have helped to accommodate such increases in limb bone stress.
Reproductive behaviour in the male cricket Gryllus bimaculatus DeGeer. I. Structure and function of the genitalia
Authors: Kumashiro Mikihiko , Sakai Masaki
Published: April 2021

We have investigated the morphology and physiology of the genitalia of the male cricket to establish a basis for neuroethological study of its reproductive behaviour. First, the structure of the phallic complex, including the dorsal pouch, guiding rod, epiphallus, ventral lobes and median pouch, are described, as are the muscles, cuticle, membranes and biomechanics of copulation. The innervation and sensory receptors have also been examined. Second, the functional role of the muscle in each genital organ has been determined by direct observation of muscle contraction during spontaneous or evoked movements and by analysis of the changes in movements after the ablation of the muscle. Third, for the flexible membranous organs, the ventral lobes and median pouch, the passages for haemolymph and their dynamic properties have been examined using petroleum jelly. Fourth, the sequence of coordinated motor actions performed by the internal and external genital organs, which were induced in both restrained and dissected males using newly developed techniques, has been analyzed during tethered copulation and spermatophore formation. As a result, the mechanisms of copulation and spermatophore formation are now more fully understood.
Gliding flight in a jackdaw: a wind tunnel study
Authors: Rosén Mikael , Hedenström Anders
Published: April 2021

We examined the gliding flight performance of a jackdaw Corvus monedula in a wind tunnel. The jackdaw was able to glide steadily at speeds between 6 and 11 m s−1. The bird changed its wingspan and wing area over this speed range, and we measured the so-called glide super-polar, which is the envelope of fixed-wing glide polars over a range of forward speeds and sinking speeds.

The glide super-polar was an inverted U-shape with a minimum sinking speed (Vms) at 7.4 m s−1 and a speed for best glide (Vbg) at 8.3 m s−1. At the minimum sinking speed, the associated vertical sinking speed was 0.62 m s−1. The relationship between the ratio of lift to drag (L:D) and airspeed showed an inverted U-shape with a maximum of 12.6 at 8.5 m s−1. Wingspan decreased linearly with speed over the whole speed range investigated. The tail was spread extensively at low and moderate speeds; at speeds between 6 and 9 m s−1, the tail area decreased linearly with speed, and at speeds above 9 m s−1 the tail was fully furled. Reynolds number calculated with the mean chord as the reference length ranged from 38 000 to 76 000 over the speed range 6–11 m s−1.

Comparisons of the jackdaw flight performance were made with existing theory of gliding flight. We also re-analysed data on span ratios with respect to speed in two other bird species previously studied in wind tunnels. These data indicate that an equation for calculating the span ratio, which minimises the sum of induced and profile drag, does not predict the actual span ratios observed in these birds. We derive an alternative equation on the basis of the observed span ratios for calculating wingspan and wing area with respect to forward speed in gliding birds from information about body mass, maximum wingspan, maximum wing area and maximum coefficient of lift. These alternative equations can be used in combination with any model of gliding flight where wing area and wingspan are considered to calculate sinking rate with respect to forward speed.