Showing papers in "The Journal of Experimental Biology in 1994"

Molecular physiology of norepinephrine and serotonin transporters

Abstract: Cocaine- and antidepressant-sensitive norepinephrine and serotonin transporters (NETs and SERTs) are closely related members of the Na+/Cl- transporter gene family, whose other members include transporters for inhibitory amino acid transmitters, neuromodulators, osmolytes and nutrients. Availability of cloned NET and SERT cDNAs has permitted rapid progress in the definition of cellular sites of gene expression, the generation of transporter-specific antibodies suitable for biosynthetic and localization studies, the examination of structure-function relationships in heterologous expression systems and a biophysical analysis of transporter function. In situ hybridization and immunocytochemical studies indicate a primary expression of NET and SERT genes in brain by noradrenergic and serotonergic neurons, respectively. Both NET and SERT are synthesized as glycoproteins, with multiple glycosylation states apparent for SERT proteins in the brain and periphery. N-glycosylation of NET and SERT appears to be essential for transporter assembly and surface expression, but not for antagonist binding affinity. Homology cloning efforts have revealed novel NET and SERT homologs in nonmammalian species that are of potential value in the delineation of the precise sites for substrate and antagonist recognition, including a Drosophila melanogaster SERT with NET-like pharmacology. Electrophysiological recording of human NETs and SERTs stably expressed in HEK-293 cells reveals that both transporters move charge across the plasma membrane following the addition of substrates; these currents can be blocked by NET-and SERT-selective antagonists as well as by cocaine.

Hydrodynamic aspects of design and attachment of a back-mounted device in penguins

Abstract: Wind tunnel and water tank experiments were carried out on a penguin model in order to optimise the shape and attachment of a back-mounted datalogger. Device-induced turbulence was minimised when the unit was placed in the most caudal position. Drag was further reduced by shaping the device to match the body contour. The hydrodynamic resistance of the package could be reduced by 65 % compared with an earlier unit. These results are discussed together with results from new studies on kinematics and energetics of underwater swimming of live instrumented penguins.

Dynamic and static stability in hexapedal runners

Abstract: Summary Stability is fundamental to the performance of terrestrial locomotion. Running cockroaches met the criteria for static stability over a wide range of speeds, yet several locomotor variables changed in a way that revealed an increase in the importance of dynamic stability as speed increased. Duty factors (the fraction of time that a leg spends on the ground relative to the stride period) decreased to 0.5 and below with an increase in speed. The duration of double support (i.e. when both tripods, or all six legs, were on the ground) decreased significantly with an increase in speed. All legs had similar touchdown phases in the tripod, but the shortest leg, the front one, lifted off before the middle and the rear leg, so that only two legs of the tripod were in contact with the ground at the highest speeds. Per cent stability margin (the shortest distance from the center of gravity to the boundaries of support, normalized to the maximum possible stability margin) decreased with increasing speed from 60 % at 10 cm s 21 to values less than zero at speeds faster than 50 cm s 21 , indicating instances of static instability at fast speeds. The center of mass moved rearward or posteriorly with respect to the base of support as speed increased. Moments about the center of mass, as shown by the center of pressure (the equivalent of a single ‘effective’ leg), were variable, but were balanced by opposing moments over a stride. Thus, hexapods can exploit the advantages of both static and dynamic stability. Static or quasi-static assumptions alone were insufficient to explain straight-ahead, constant-speed locomotion and may hinder discovery of behaviors that are dynamic, where kinetic energy and momentum can act as a bridge from one step to the next.

Studies of tropical tuna swimming performance in a large water tunnel - energetics

Abstract: Yellowfin tuna (Thunnus albacares) swimming kinematics was studied in a large water tunnel at controlled swimming velocities (U). Quantified kinematic variables included the tail-beat frequency, stride length (l), caudal amplitude, yaw, the propulsive wavelength, the speed of the propulsive wave (C) and the sweepback angle of the pectoral fins. In general, all variables, except the propulsive wavelength and consequently C, are comparable to values determined for other teleosts. The propulsive wavelength for the tunas (1.23­1.29 L, where L is fork length) is 30­60 % longer than in other cruise-adapted teleosts such as salmonids. The resulting thunniform swimming mode and the morphological and anatomical adaptations associated with the long propulsive wavelength (e.g. fusiform body shape, rigid vertebral column) act to minimize anterior resistance and maximize caudal thrust. The long propulsive wavelength also increases the maximum l which, in concert with the elevated muscle temperatures of tunas, increases their maximum swimming velocity.

Integrated responses to exhaustive exercise and recovery in rainbow trout white muscle: acid-base, phosphogen, carbohydrate, lipid, ammonia, fluid volume and electrolyte metabolism

Abstract: White muscle and arterial blood plasma were sampled at rest and during 4 h of recovery from exhaustive exercise in rainbow trout. A compound respiratory and metabolic acidosis in the blood was accompanied by increases in plasma lactate (in excess of the metabolic acid load), pyruvate, glucose, ammonia and inorganic phosphate levels, large elevations in haemoglobin concentration and haematocrit, red cell swelling, increases in the levels of most plasma electrolytes, but no shift of fluid out of the extracellular fluid (ECF) into the intracellular fluid (ICF) of white muscle. The decrease in white muscle pHi was comparable to that in pHe; both recovered by 4 h. Creatine phosphate and ATP levels were both reduced by 40% after exercise, the former recovering within 0.25 h, whereas the latter remained depressed until 4 h. Changes in creatine concentration mirrored those in creatine phosphate, whereas changes in IMP and ammonia concentration mirrored those in ATP. White muscle glycogen concentration was reduced 90% primarily by conversion to lactate; recovery was slow, to only 40% of resting glycogen levels by 4 h. During this period, most of the lactate and metabolic acid were retained in white muscle and there was excellent conservation of carbohydrate, suggesting that in situ glycogenesis rather than oxidation was the major fate of lactate. The redox state ([NAD+]/[NADH]) of the muscle cytoplasm, estimated from ICF lactate and pyruvate levels and pHi, remained unchanged from resting levels, challenging the traditional view of the 9anaerobic9 production of lactate. Furthermore, the membrane potential, estimated from levels of ICF and ECF electrolytes using the Goldman equation, remained unchanged throughout, challenging the view that white muscle becomes depolarized after exhaustive exercise. Indeed, ICF K+ concentration was elevated. Lactate was distributed well out of electrochemical equilibrium with either the membrane potential (Em) or the pHe-pHi difference, supporting the view that lactate is actively retained in white muscle. In contrast, H+ was actively extruded. Ammonia was distributed passively according to Em rather than pHe-pHi throughout recovery, providing a mechanism for retaining high ICF ammonia levels for adenylate resynthesis in situ. Although lipid is not traditionally considered to be a fuel for burst exercise, substantial decreases in free carnitine and elevations in acyl-carnitines and acetyl-CoA concentrations indicated an important contribution of fatty acid oxidation by white muscle during both exercise and recovery.

Odor plumes and how blue crabs use them in finding prey.

Abstract: Orientation of animals using chemical cues often takes place in flows, where the stimulus properties of odorants are affected by the characteristics of fluid motion. Kinematic analysis of movement patterns by animals responding to odor plumes has been used to provide insight into the behavioral and physiological aspects of olfactory-mediated orientation, particularly in terrestrial insects. We have used this approach in analyzing predatory searching by blue crabs in response to plumes of attractant metabolites released from the siphons of live clams in controlled hydrodynamic environments. Crabs proceed directly upstream towards clams in smooth-turbulent flows and show high locomotory velocities and few periods of motionlessness. Crabs assume more indirect trajectories and display slower locomotion and more stopping in rough-turbulent flows. This degradation of foraging performance is most pronounced as flow shifts from a smooth- to a rough-turbulent regime, where the change in hydraulic properties is associated with contraction of the viscous sublayer region of the boundary layer. Because flow in this region is quasilaminar, the viscous sublayer may be a particularly effective vehicle for chemical stimulus transmission, such that orientation is severely compromised when it is reduced or removed. Our results also suggest that rheotactic and chemical information are both necessary for successful orientation. Perception of chemical cues acts to bias locomotion upcurrent, and feedback from odorant stimulus distributions appears directly to regulate subsequent stopping and turning en route to prey. Although the mechanisms of orientation to odorant plumes displayed by insects and blue crabs are largely similar, blue crabs appear to rely more heavily on spatial and/or temporal aspects of chemical stimulus distributions than has been suggested for insect systems.

The effects of wing rotation on unsteady aerodynamic performance at low reynolds numbers

Abstract: The downstroke-to-upstroke transition of many insects is characterized by rapid wing rotation. The aerodynamic consequences of these rapid changes in angle of attack have been investigated using a mechanical model dynamically scaled to the Reynolds number appropriate for the flight of small insects such as Drosophila. Several kinematic parameters of the wing flip were examined, including the speed and axis of rotation, as well as the duration and angle of attack during the wing stroke preceding rotation. Alteration of these kinematic parameters altered force generation during the subsequent stroke in a variety of ways. 1. When the rotational axis was close to the trailing edge, the model wing could capture vorticity generated during rotation and greatly increase aerodynamic performance. This vortex capture was most clearly manifested by the generation of lift at an angle of attack of 0°;. Lift at a 0°; angle of attack was also generated following rotation about the leading edge, but only if the downstroke angle was large enough to generate a von Karman street. The lift may be due to an alteration in the effective angle of attack caused by the inter-vortex stream in the downstroke wake. 2. The maximum lift attained (over all angles of attack) was substantially elevated if the wing translated backwards through a wake generated by the previous stroke. Transient lift coefficient values of nearly 4 were obtained when the wing translated back through a von Karman street generated at a 76.5°; angle of attack. This effect might also be explained by the influence of the inter-vortex stream, which contributes a small component to fluid velocity in the direction of translation. 3. The growth of lift with angle of attack was significantly elevated following a 7.5 chord stroke with a 76.5°; angle of attack, although it was relatively constant under all other kinematic conditions. 4. The results also indicate the discrepancies between transient and time-averaged measures of performance that arise when unsteady mechanisms are responsible for force generation. Although the influence of wing rotation was strong during the first few chords of translation, averaging the performance over as little as 6.5 chords of motion greatly attenuated the effects of rotation. 5. Together, these modeling results suggest that the unsteady mechanisms generated by simple wing flips could provide an important source for the production of aerodynamic forces in insect flight. Furthermore, the extreme sensitivity to small variations in almost all kinematic parameters could provide a foundation for understanding the aerodynamic mechanisms underlying active flight control.

The malpighian tubules of Drosophila melanogaster: a novel phenotype for studies of fluid secretion and its control.

Abstract: The insect renal (Malpighian) tubule has long been a model system for the study of fluid secretion and its neurohormonal control (Maddrell, 1981; Maddrell and O’Donnell, 1992). Classical physiology suggests a model for tubular secretion of iso-osmotic fluid in most insects, in which ions are thought to enter basally either through a series of ion channels (Na+, K+ and Cl2) or through a bumetanide-sensitive Na+/K+/2Cl2 cotransport. Apical fluxes are energised by a plasma-membrane H+-pumping V-ATPase, driving secretion of Na+ or K+ through one or more exchangers, at least one of which is amiloride-sensitive and appears to be closely similar to the Na+/H+ exchanger of vertebrates (Maddrell and O’Donnell, 1992). Cl2 follows passively, perhaps through apical Cl2 channels. Water follows the major ions, and haemolymph solutes diffuse across the tubule wall passively via a paracellular route. There are also transcellular active transport processes for certain metabolites or toxins, such as acylamides (Maddrell et al. 1974) and plant alkaloids (Maddrell, 1976; O’Donnell et al. 1983). To extend studies of ion transport beyond that revealed by the techniques of classical physiology, it is necessary to adopt a molecular genetic approach, in which the relevant genes are identified, characterised and mutated. However, the generalised molecular genetic dissection of the major genes responsible for a stimulus–secretion pathway in any vertebrate epithelium remains a daunting task, as the genetic tools available are relatively unsophisticated. Encouraged by previous physiological work on the larger, but less well genetically mapped, Drosophila hydei (Bertram et al. 1991; Wessing and Eichelberg, 1978; Wessing et al. 1987), we have investigated the feasibility of adapting the established tubule secretion assay to the study of tubules of larval and adult Drosophila melanogaster. The results (i) demonstrate that these Malpighian tubules, to

Mechanical determinants of the minimum energy cost of gradient running in humans.

Abstract: The metabolic cost and the mechanical work of running at different speeds and gradients were measured on five human subjects. The mechanical work was partitioned into the internal work (Wint) due to the speed changes of body segments with respect to the body centre of mass and the external work (Wext) due to the position and speed changes of the body centre of mass in the environment. Wext was further divided into a positive part (W+ext) and a negative part (W-ext), associated with the energy increases and decreases, respectively, over the stride period. For all constant speeds, the most economical gradient was -10.6 +/-0.5% (S.D., N = 5) with a metabolic cost of 146.8 +/- 3.8 ml O2 kg-1 km-1. At each gradient, there was a unique W+ext/W-ext ratio (which was 1 in level running), irrespective of speed, with a tendency for W-ext and W+ext to disappear above a gradient of +30% and below a gradient of -30%, respectively. Wint was constant within each speed from a gradient of -15% to level running. This was the result of a nearly constant stride frequency at all negative gradients. The constancy of Wint within this gradient range implies that Wint has no role in determining the optimum gradient. The metabolic cost C was predicted from the mechanical experimental data according to the following equation: [formula: see text] where eff- (0.80), eff+ (0.18) and effi (0.30) are the efficiencies of W-ext, W+ext and Wint, respectively, and el- and el+ represent the amounts of stored and released elastic energy, which are assumed to be 55J step-1. The predicted C versus gradient curve coincides with the curve obtained from metabolic measurements. We conclude that W+ext/W-ext partitioning and the eff+/eff- ratio, i.e. the different efficiency of the muscles during acceleration and braking, explain the metabolic optimum gradient for running of about -10%.

Regulation of feeding behavior in adult Drosophila melanogaster varies with feeding regime and nutritional state.

Abstract: The regulation of feeding behavior in adult Drosophila melanogaster includes such elements as ingestion responsiveness, volume ingested in a single meal, food storage in the crop and rate of defecation. Our results suggest that feeding behavior varies in a manner dependent on feeding regime (food-deprived or ad-libitum-fed) and nutritional state. Fed flies that are subsequently food-deprived become increasingly more responsive to food stimuli over time and, when offered 1% agar diets containing different concentrations of sucrose, ingest greater amounts of diets that have higher sucrose concentrations. When fed ad libitum for 72 h on these same diets, D. melanogaster maintained much smaller crops on average than food-deprived flies fed a single meal. Additionally, ad-libitum-fed flies are grouped into two categories depending on the concentration of sucrose in the diet. Flies fed for 72 h on 1% agar diets having 50 mmoll-1 sucrose or more are not affected by the concentration of sucrose in the diet, while flies fed on diets of 15 or 25 mmoll-1 sucrose increase ingestion responsiveness, crop size and the rate of defecation with decreasing concentrations of sucrose in the diet. Flies fed on even lower sucrose concentrations (5 or 10 mmoll-1 sucrose) for 27-72 h exhibit both a shift over time to larger crop sizes and increased mortality over those of flies fed 15 mmoll-1 sucrose. These data suggest that flies fed ad libitum are capable of modulating their feeding behavior in response to their nutritional state.

Effects of zinc on the kinetics of branchial calcium uptake in freshwater rainbow trout during adaptation to waterborne zinc.

Abstract: The effects of sublethal waterborne Zn2+ (150 micrograms l-1 = 2.3 mumol-1) on the kinetics of unidirectional Ca2+ influx were studied in juvenile freshwater rainbow trout during chronic exposure (60 days) at a water [Ca2+] of 1.0 mmol l-1. An unexposed group held under identical conditions served as control. The presence of Zn2+ in the water increased the apparent Km for Ca2+ influx by up to 300% with only a small inhibitory effect (35% at most) on the maximum rate of uptake (Jmax). These results, in combination with earlier data showing that Ca2+ competitively inhibits Zn2+ uptake, suggest that Zn2+ and Ca2+ compete for the same uptake sites. Acute withdrawal of Zn2+ after 3h of exposure resulted in a 23-fold reduction in Km for Ca2+, but a persistent small depression of Jmax. During prolonged exposure to Zn2+, the apparent Km for Ca2+ remained greatly elevated and Jmax remained slightly depressed. The actual Ca2+ influx in hard water ([Ca2+] = 1.0 mmol l-1) decreased marginally and paralleled the small changes in Jmax. The increases in apparent Km had a negligible influence on the actual Ca2+ influx because Km values (38–230 mumol l-1), even when elevated by Zn2+, remained below the water [Ca2+] (1000 mumol l-1). Rainbow trout exposed to Zn2+ exhibited a slower rate of protein synthesis in the gills (measured on day 23) and an increased tolerance to Zn2+ challenge (measured on both days 27 and 50). Unidirectional Zn2+ influx, measured at the end of the exposure period, was significantly reduced in the Zn2+-exposed fish. There were no changes in hepatic or branchial Zn2+, Cu2+ or metallothionein concentrations. We hypothesize that, during exposure to sublethal [Zn2+] in hard water, the fish may change the Km for a mutual Ca2+/Zn2+ carrier so as to reduce markedly Zn2+ influx without greatly altering Ca2+ influx. This reduced Zn2+ influx, rather than metallothionein induction, may be the basis of adaptation to elevated concentrations of waterborne Zn2+.

IMMUNOLOCALIZATION OF H + -ATPase IN THE GILL EPITHELIA OF RAINBOW TROUT

Abstract: The localization of proton pumps (H+-ATPase) in gill epithelia of rainbow trout [Oncorhynchus mykiss (Walbaum)] was elucidated by immunofluorescence microscopy, using rabbit polyclonal antibodies against the 70 kDa subunit of H+-ATPase purified from clathrin-coated vesicles of bovine brain. In the gill epithelia of freshwater trout, the immunostaining was uniformly distributed along the lamellae and generally concentrated in apical regions. It is concluded, therefore, that H+-ATPase is located in the apex of both chloride cells and epithelial cells of freshwater fish. Hypercapnic treatment resulted in a non-polarized and restrictive distribution of H+-ATPase in the chloride cell. No fluorescent staining was observed in the gill epithelium of seawater-adapted rainbow trout, except in some unidentified anucleate surface material. The presence of the 70 kDa subunit in fish gill epithelia was confirmed by Western blot. These results support the proposed role of a proton pump in sodium uptake in freshwater fish and demonstrate that the H+-ATPase in fish gills is of the vacuolar type, antigenically similar to the H+-ATPase in mammalian brain and kidney.

Energy savings in formation flight of pink-footed geese

Abstract: Fifty-four skeins of pink-footed geese (Anser brachyrhynchus) were photographed from directly underneath to eliminate the effects of perspective distortion, and the wing-tip spacings (the distance between adjacent birds' wing tips perpendicular to the flight path at maximum wingspan) and depths (the distance between adjacent birds' body centres parallel to the flight path) were measured at the same time as local wind speeds. The photographs were used to test for savings in induced power from wing positioning relative to the predicted positions of vortices generated by other wings, using a theoretical model. The mean wing-tip spacing corresponded to a saving in induced power of 14 %, less than one-third of the maximum possible. The saving in total power might be as low as 2.4 %. The high variation in wing-tip spacing suggests that pink-footed geese found difficulty maintaining position and thus adopted a strategy of flying outboard of the optimal position that maximises savings. This may minimise the risk of straying into a zone where savings are negative. There was a significant correlation between depth and wing-tip spacing, supporting an alternative communication hypothesis, whereby the birds position themselves to obtain maximum information on their neighbour's position. In high winds, there was little change in wing-tip spacing variation but a decrease in depth variation, suggesting a shift towards more regularly spaced skeins.

Ultraviolet photoreception contributes to prey search behaviour in two species of zooplanktivorous fishes

Abstract: We tested the hypothesis that ultraviolet photoreception contributes to prey search in small juvenile rainbow trout (Oncorhynchus mykiss) and pumpkinseed sunfish (Lepomis gibbosus) while foraging on Daphnia pulex. Small individuals of these species are ultraviolet-photosensitive zooplanktivores. For both species, prey pursuit distances and angles were larger under full-spectrum illumination than under ultraviolet-absent illumination. The same was true for the distances and angles associated with repositioning movements (i.e. those not leading to the location of a prey item). Thus, ultraviolet photoreception contributes to prey search and detection in these fishes. We argue that the most likely mechanism underlying this enhancement of prey search abilities is improved target contrast.

AMMONIA EXCRETION IN FRESHWATER RAINBOW TROUT (ONCORHYNCHUS MYKISS) AND THE IMPORTANCE OF GILL BOUNDARY LAYER ACIDIFICATION: LACK OF EVIDENCE FOR Na+/NH4+ EXCHANGE

Abstract: Net ammonia fluxes (JAmm) were measured in adult freshwater rainbow trout in vivo under a variety of conditions designed to inhibit unidirectional sodium uptake (JinNa; low external [NaCl], 10(-4) mol l-1 amiloride), alter transbranchial PNH3 and NH4+ gradients [24 h continuous (NH4)2SO4 infusion, or exposure to 1 mmol l-1 external total ammonia at pH 8] and prevent gill boundary layer acidification (5 mmol l-1 Hepes buffer). Inhibition of JinNa with amiloride or low external [NaCl] under normal conditions reduced JAmm by about 20 %, but did not prevent the net excretion of ammonia during exposure to high concentrations of external ammonia. Increasing the buffer capacity of the ventilatory water with Hepes buffer (pH 8) reduced JAmm by 36 % and abolished the effect of amiloride on ammonia excretion. No evidence could be found to support a directly coupled apical Na+/NH4+ exchange. We suggest that any dependence of ammonia excretion on sodium uptake is caused by alteration of transbranchial PNH3 gradients within the gill microenvironment secondary to changes in net H+ excretion. Under normal conditions (pH 8, low external ammonia) gill boundary layer acidification facilitates over one-third of the total ammonia excretion. During exposure to high concentrations of external ammonia in poorly buffered water, estimates of transbranchial PNH3 gradients from measurements of bulk water pH and total ammonia concentration (TAmm) may be grossly in error because of boundary layer acidification. Prevention of boundary layer acidification with Hepes buffer during exposure to high cocncentrations of external ammonia revealed that the local transbranchial PNH3 gradient at the gill may in fact be positive (blood to water), negating the need for an active NH4+ transport mechanism. In freshwater trout, NH3 diffusion may account for all ammonia excretion under all experimental conditions used in the present study.

Detection of magnetic inclination angle by sea turtles: a possible mechanism for determining latitude

Abstract: For animals that migrate long distances, the magnetic field of the earth provides not only a possible cue for compass orientation, but a potential source of world-wide positional information. At each location on the globe, the geomagnetic field lines intersect the earth's surface at a specific angle of inclination. Because inclination angles vary with latitude, an animal able to distinguish between different field inclinations might, in principle, determine its approximate latitude. Such an ability, however, has never been demonstrated in any animal. We studied the magnetic orientation behavior of hatchling loggerhead sea turtles (Caretta caretta L.) exposed to earth-strength magnetic fields of different inclinations. Hatchlings exposed to the natural field of their natal beach swam eastward, as they normally do during their offshore migration. In contrast, those subjected to an inclination angle found on the northern boundary of the North Atlantic gyre (their presumed migratory path) swam south-southwest. Hatchlings exposed to an inclination angle found near the southern boundary of the gyre swam in a northeasterly direction, and those exposed to inclination angles they do not normally encounter, or to a field inclination found well within the northern and southern extremes of the gyre, were not significantly oriented. These results demonstrate that sea turtles can distinguish between different magnetic inclination angles and perhaps derive from them an approximation of latitude. Most sea turtles nest on coastlines that are aligned approximately north­south, so that each region of nesting beach has a unique inclination angle associated with it. We therefore hypothesize that the ability to recognize specific inclination angles may largely explain how adult sea turtles can identify their natal beaches after years at sea.

Ca2+ transport in Saccharomyces cerevisiae.

Abstract: Cytosolic free Ca2+ is maintained at submicromolar levels in budding yeast by the activity of Ca2+ pumps and antiporters. We have recently identified the structural genes for two Ca2+ pumps, PMC1 [correction of PCM1] and PMR1, which are required for Ca2+ sequestration into the vacuole and secretory organelles, respectively. The function of either Ca2+ pump is sufficient for yeast viability, but deletion of both genes is lethal because of elevation of cytosolic [Ca2+] and activation of calcineurin, a Ca(2+)- and calmodulin-dependent protein phosphatase. Calcineurin activation decreases Ca2+ sequestration in the vacuole by a putative Ca2+ antiporter and may also increase Ca2+ pump activity. These regulatory processes can affect the ability of yeast strains to tolerate high extracellular [Ca2+]. We propose a model in which the cellular response to changes in the environmental levels of Ca2+ is mediated by calmodulin and calcineurin which, in turn, modulate the various types of Ca2+ transporters.

Characterization of macrophages and neutrophilic granulocytes from the pronephros of carp (cyprinus carpio)

Abstract: To analyse the functional activity of different leucocyte types, carp pronephros cells were separated on Percoll density gradients and by use of fluorescence-activated cell sorting. Cell populations were characterised by light and electron microscopy and by flow cytometry. Fractions enriched in macrophages and neutrophilic granulocytes were subsequently analysed for phagocytic activity in vitro by quantification of the uptake of Escherichia coli bacteria or yeast cells, and for respiratory burst response by measurement of the production of the reactive oxygen intermediates O2· and H2O2. Both cell types showed very active in vitro phagocytosis and production of both O2· and H2O2. When activated with phorbol myristate acetate or bacteria, it was evident that the neutrophilic granulocytes were significantly more active than the macrophages. Analysis of single-cell respiratory burst activity in fish phagocytes was investigated after preloading of cells with dihydrorhodamine123. Cells were subsequently separated and analysed for fluorescence using flow cytometry. Both the macrophage-enriched fraction and the granulocyte-enriched fraction appeared to consist of active and inactive subpopulations. In comparison with the inactive populations, active populations had characteristic high forward/sideward scatter profiles.

Underwater swimming at low energetic cost by pygoscelid penguins

Abstract: Energetic requirements of under-water swimming in pygoscelid penguins were studied in Antarctica, using respirometry together with a 21 m long swim canal and externally attached devices recording the swimming speed and dive duration of unrestrained animals. Field measurements were compared with measurements of the hydrodynamic properties of an Adelie penguin model in a circulating water tank. Minimium transport costs during underwater swimming in Adelie (Pygoscelis adeliae), chinstrap (P. antarctica) and gentoo (P. papua) penguins averaged 4.9, 3.7 and 7.6 J kg-1 m-1, respectively, at their preferred swimming speeds of 2.2, 2.4 and 1.8 m s-1, allowing the birds to dive aerobically for 110, 130 and 93 s, respectively. From the swim canal measurements, we calculated a drag coefficient (CD) of 0.0368 for a typical Adelie penguin at 2.2 m s-1. This value is significantly lower than the CD of 0.04 of an ideal spindle and the CD of 0.0496 measured on the model in the laboratory. The reasons for this difference are discussed.

The effects of muscle fatigue on bone strain

Abstract: There is anecdotal evidence that bone strains may increase to the point that bone becomes susceptible to rapid failure when muscles become fatigued. To determine whether neuromuscular response could be a factor in accelerating bone failure, we tested the hypothesis that muscle fatigue causes a significant increase in peak principal and shear strains in bone. Ten adult foxhounds were subjected to rigorous exercise that caused muscular fatigue while myoelectrical activity of the quadriceps and hamstrings and strain on the distal tibia were monitored simultaneously. Ground reaction forces on the dog hindlimbs were measured before and after strain gauges had been applied to the tibia. The data show a significant shift to lower median myoelectrical frequencies in the quadriceps, indicating muscular fatigue, following the 20 min exercise period. In conjunction with this shift, peak principal and shear strains increased on both compressive and tensile cortices of the tibia and shear strain on the tensile cortex increased significantly (P = 0.02). The largest changes were along the anterior and anterolateral surfaces of the tibia, where peak principal strain increased by an average of 26-35% following muscular fatigue. The cross-sectional strain distribution was calculated at the gauge site at peak strain at the beginning of the exercise period and at peak strain after 20 min of exercise. These data show a change in strain distribution when muscle becomes fatigued. Strains on the posterior cortex of the bone showed the greatest change. Correlation analysis demonstrated a significant inverse association between median myoelectrical frequency and bone strain after 20 min of exercise (Spearman r2 = 1.00; P = 0.05). These data show that muscle fatigue may be associated with increased bone strain.

Kinematics of pectoral fin locomotion in the bluegill sunfish lepomis macrochirus

Abstract: The pectoral fins of ray-finned fishes are flexible and capable of complex movements, and yet little is known about the pattern of fin deformation during locomotion. For the most part, pectoral fins have been modeled as rigid plates. In order to examine the movements of different portions of pectoral fins, we quantified the kinematics of pectoral fin locomotion in the bluegill sunfish Lepomis macrochirus using several points on the distal fin edge and examined the effects of swimming speed on fin movements. We simultaneously videotaped the ventral and lateral views of pectoral fins of four fish swimming in a flow tank at five speeds ranging from 0.3 to 1.1 total lengths s-1. Four markers, placed on the distal edge of the fin, facilitated field-by-field analysis of kinematics. We used analyses of variance to test for significant variation with speed and among the different marker positions. Fin beat frequency increased significantly from 1.2 to 2.1 Hz as swimming speed increased from 0.3 to 1.0 total lengths s-1. Maximal velocities of movement for the tip of the fin during abduction and adduction generally increased significantly with increased swimming speed. The ratio of maximal speed of fin retraction to swimming speed declined steadily from 2.75 to 1.00 as swimming speed increased. Rather than the entire distal edge of the fin always moving synchronously, markers had phase lags as large as 32 with respect to the dorsal edge of the fin. The more ventral and proximal portions of the fin edge usually had smaller amplitudes of movement than did the more dorsal and distal locations. With increased swimming speed, the amplitudes of the lateral and longitudinal fin movements generally decreased. We used two distal markers and one basal reference point to determine the orientation of various planar fin elements. During early adduction and most of abduction, these planar fin elements usually had positive angles of attack. Because of fin rotation, angles of attack calculated from three-dimensional data differed considerably from those estimated from a simple lateral projection. As swimming speed increased, the angles of attack of the planar fin elements with respect to the overall direction of swimming approached zero. The oscillatory movements of the pectoral fins of bluegill suggest that both lift- and drag-based propulsive mechanisms are used to generate forward thrust. In addition, the reduced frequency parameter calculated for the pectoral fin of Lepomis (sigma=0.85) and the Reynolds number of 5x10(3) indicate that acceleration reaction forces may contribute significantly to thrust production and to the total force balance on the fin.

The olfactory memory of the honeybee Apis mellifera. II. Blocking between odorants in binary mixtures.

Abstract: Proboscis extension conditioning of honeybee workers was used to study the processing of odorants when bees were conditioned to binary mixtures. Responses to a set of pure floral odors and pheromones after conditioning have already been described. When bees are conditioned to certain mixtures of odorants, the response to both components is equal to that when they are tested alone. However, mixtures of an aliphatic aldehyde and an alcohol elicit asymmetric response patterns; that is, the response to the aldehyde is much stronger than that to the alcohol. A bee9s response to the alcohol after it had been trained in an aldehyde background is significantly lower than when the bee is trained to respond to the same alcohol in the background of another odorant. Such response patterns are not necessarily caused by a behavioral decrement resulting from a compound-unique perceptual effect produced by the mixture. Furthermore, studies of blocking show that behavioral acquisition in response to one component can be hindered or blocked by pretraining with the other component. These results suggest that honeybees can perceive the individual components of some binary mixtures. The similarities in neural processing in olfactory systems of vertebrates and invertebrates mean that such studies could elucidate behavioral mechanisms of olfaction in a wide phylogenetic spectrum of animals.

Effects of environmental temperature on the metabolic and acid-base responses of rainbow trout to exhaustive exercise

Abstract: In vivo experiments were conducted to determine how the physiological response to exhaustive exercise in rainbow trout (Oncorhynchus mykiss) is affected by environmental temperature. The white muscle acid­base status (e.g. pH, PCO2, HCO3- and deltaH+m) and metabolite (e.g. lactate, phosphocreatine, ATP and glycogen) content, and the acid­base status and lactate concentrations in the blood, were measured at rest and during recovery from burst exercise in rainbow trout acclimated to either 5 or 18 °C. Trout acclimated to the warmer temperature had higher resting levels of white muscle phosphocreatine (PCr) and also utilized a greater proportion of their muscle ATP and glycogen stores during burst activity compared with trout acclimated to the colder temperature. Recovery of muscle PCr and glycogen levels was independent of acclimation temperature, but muscle ATP levels recovered faster at 18 °C. Exhaustive exercise resulted in a similar lactacidosis in the muscle of trout acclimated to either temperature. In contrast, temperature had a marked influence on the lactacidosis in the blood. Blood lactate and metabolic proton concentrations following exercise were about twofold greater in fish acclimated to 18 °C than in fish acclimated to 5 °C. Despite the more severe acidosis and the greater lactate accumulation in the plasma of fish acclimated to warmer temperatures, the time required for recovery of these variables was similar to that at 5 °C. Taken together, these results suggest that acclimation temperature does not significantly affect anaerobic capacity in rainbow trout, but may account for much of the documented variability in the dynamics of the lactacidosis in blood following exhaustive exercise in fish.

Maintenance of dynamic strain similarity and environmental stress factor in different flow habitats: thallus allometry and material properties of a giant kelp

Abstract: We have focused on the giant kelp Nereocystis luetkeana to examine the mechanical scaling of benthic marine organisms loaded in tension by hydrodynamic forces. If we consider simply the allometry of the kelp's morphological characters, we conclude that their stipes are underscaled relative to the blade area they support (i.e. that the kelp do not maintain stress or elastic similarity as they grow). However, a closer look at the characteristics of these kelp in the field reveals (1) that they have different blade shapes (and hence drag coefficients) and stipe material properties in different hydrodynamic environments, and (2) that they show a decrease in drag coefficient as they become larger. One consequence of these adjustments of blade and stipe morphology is that the maximum stresses in N. luetkeana stipes, when the kelp are pulled by peak tidal currents in their respective habitats, are similar for kelp of different sizes and for kelp from different sites. Hence, sessile organisms such as these kelp can, via their growth responses in different mechanical environments, show a phenomenon analogous to dynamic strain similarity. In addition, N. luetkeana also maintain a constant environmental stress factor, the ratio of the stress required to break a component of an organism (in this case the stipe) at some stage in its life to the maximum stress normally encountered in the habitat by that component during that stage (in this case, stress due to the drag on a kelp exposed to the peak tidal currents typical of the site at which it lives), both between habitats and as they grow.

Allatostatin peptides in the crab stomatogastric nervous system: inhibition of the pyloric motor pattern and distribution of allatostatin-like immunoreactivity.

Abstract: The effects of four Diploptera punctata allatostatin peptides on the stomatogastric nervous system of the crab Cancer borealis were studied. All of the peptides had similar actions on the activity of neurons involved in rhythmic movements of the pyloric region of the stomach, decreasing the frequency of the pyloric rhythm in a dose-dependent manner. Diploptera allatostatin 3 (D-AST-3) was slightly more effective than the others. The absolute change in the frequency of the pyloric rhythm depended on the starting frequency, demonstrating that the effect of D-AST-3 depends on the preceding physiological state of the preparation. The largest decreases were observed when the starting frequency was slower than 0.8 Hz. Whole-mount immunocytochemistry with anti-Diploptera allatostatin 1 antibodies demonstrated the presence of allatostatin-like peptides in the paired commissural ganglia, the unpaired oesophageal ganglion and the stomatogastric ganglion, and in their connecting and motor nerves. Dense processes were labeled in the stomatogastric ganglion, 12-19 cell bodies and neuropil staining were found in each commissural ganglion, two cell bodies were stained in the oesophageal ganglion and two pairs of cell bodies, the gastropyloric receptor neurons, were stained in peripheral nerves.

Biochemistry and molecular biology of the vesicular monoamine transporter from chromaffin granules.

Abstract: Prior to secretion, monoamines (catecholamines, serotonin, histamine) are concentrated from the cytoplasm into vesicles by vesicular monoamine transporters (VMAT). These transporters also carry non-physiological compounds, e.g. the neurotoxin methyl-4-phenylpyridinium. VMAT acts as an electrogenic antiporter (exchanger) of protons and monoamines, using a proton electrochemical gradient. Vesicular transport is inhibited by specific ligands, including tetrabenazine, ketanserin and reserpine. The mechanism of transport and the biochemistry of VMAT have been analyzed with the help of these tools, using mainly the chromaffin granules from bovine adrenal glands as a source of transporter. Although biochemical studies did not suggest a multiplicity of VMATs, two homologous but distinct VMAT genes have recently been cloned from rat, bovine and human adrenal glands. The VMAT proteins are predicted to possess 12 transmembrane segments, with both extremities lying on the cytoplasmic side. They possess N-glycosylation sites in a putative luminal loop and phosphorylation sites in cytoplasmic domains. In rat, VMAT1 is expressed in the adrenal gland whereas VMAT2 is expressed in the brain. In contrast, we found that the bovine adrenal gland expressed both VMAT1 and VMAT2. VMAT2 corresponds to the major transporter of chromaffin granules, as shown by partial peptidic sequences of the purified protein and by a pharmacological analysis of the transport obtained in transfected COS cells (COS cells are monkey kidney cells possessing the ability to replicate SV-40-origin-containing plasmids). We discuss the possibility that VMAT1 may be specifically addressed to large secretory granules vesicles, whereas VMAT2 may also be addressed to small synaptic vesicles; species differences would then reflect the distinct physiological roles of the small synaptic vesicles in the adrenal gland.

Spontaneous swimming activity of atlantic cod gadus morhua exposed to graded hypoxia at three temperatures

Abstract: The spontaneous swimming activity of Atlantic cod Gadus morhua was investigated at graded levels of hypoxia at three temperatures (5, 10 and 15 °C) by using a computerized system monitoring animal activity. The fish were tested individually, and swimming distance was used as a measure of activity. No significant effect of temperature on swimming distance in normoxic water was found. At all temperatures, activity level decreased with decreasing oxygen saturation. Swimming behaviour at normoxia and 50 % and 25 % oxygen saturation is described. No apparent avoidance of hypoxic water was found based on the distribution of swimming speeds and turning angles. The possible benefits of a decreased activity level in a hypoxic environment are discussed.

Neurotransmitter transporters: Three important gene families for neuronal function

Abstract: Three distinct gene families encode transporter proteins that aid in temporal and spatial buffering of neurotransmitter and neurotransmitter metabolite concentrations and allow neurons to cycle and recycle transmitter molecules. Analyses of these gene families and their products are likely to enhance understanding of the molecular neurobiology of neuronal function and may elucidate contributors to the genetic etiologies of neurological and psychiatric disease.

Flight physiology of neotropical butterflies: allometry of airspeeds during natural free flight

Abstract: Airspeed measurements during natural free flight were made on a total of 270 neotropical butterflies representing 62 species. Morphological data were obtained from the same individuals for which airspeeds had been determined. Flight speed was positively correlated with body mass, thoracic mass and wing loading. Controlling for body mass, higher wing loadings were correlated with increased flight speed. Flight speed and wing aspect ratio were negatively correlated. No consistent correlations were found between airspeed and wing length, wing area or body length. Released butterflies and butterflies encountered in natural free flight did not differ substantially in flight speed allometry. The observed scaling of flight speeds was similar to that derived for a much smaller sample of butterflies flying in an insectary, although absolute values of flight speed were approximately three times higher in natural flight and correlation coefficients of allometric regressions were typically lower. These results suggest that butterfly airspeeds under natural conditions can reasonably be predicted from morphological measurements, and that studying flight in enclosed spaces preserves the allometry of flight speeds.

Hindlimb kinematics during terrestrial locomotion in a salamander (dicamptodon tenebrosus)

Abstract: A quantitative study of hindlimb kinematics during terrestrial locomotion in a non-specialized salamander was undertaken to allow comparisons with limb movements in other groups of tetrapods. Five Dicamptodon tenebrosus were videotaped at 200 fields s-1 walking on a treadmill. Coordinates of marker points on the salamander's midline, pelvic girdle and left hindlimb were digitized through at least three strides at both a walk (0.77 SVL s-1, where SVL is snout­vent length) and a trot (2.90 SVL s-1). Marker coordinates were used to compute kinematic variables summarizing trunk flexion, pelvic girdle rotation, femoral protraction/retraction and knee flexion/extension. The stride is characterized by uninterrupted trunk and pelvic girdle oscillation, femoral retraction throughout stance phase, and knee flexion in early stance followed by extension. Mean angular excursions are: trunk, 66 °; pelvic girdle, 38.5 °; pelvic girdle­femur, 106 °; and knee, 65 °. The hindlimb and pelvic girdle also show a complicated pattern of lateral movement related to knee flexion/extension and periods of support by the contralateral hindlimb during the step cycle. Dicamptodon shares the following features of the hindlimb step cycle with other tetrapod taxa: rotation of the pelvic girdle through a 30­40 ° arc, femoral retraction beginning simultaneously with and persisting throughout stance phase, flexion of the knee in early stance, and extension of the knee in late stance.