The first direct measurements of ventilatory flow and oxygen utilization after exhaustive exercise and voluntary feeding in a teleost fish, Oncorhynchus mykiss.

A new "less invasive" device incorporating an ultrasonic flow probe and a divided chamber, but no stitching of membranes to the fish, was employed to make the first direct measurements of ventilatory flow rate (V̇w) and % O2 utilization (%U) in juvenile rainbow trout (37 g, 8ºC) after exhaustive exercise (10-min chasing) and voluntary feeding (2.72% body mass ration). Under resting conditions, the allometrically scaled V̇w (300 ml kg-1 min-1 for a 37-g trout = 147 ml kg-1 min-1 for a 236-g trout exhibiting the same mass-specific O2 consumption rate, MO2) and the convection requirement for O2 (CR = 4.13 L mmol-1) were considerably lower, and the %U (67%) was considerably higher than in previous studies using surgically attached masks or the Fick principle. After exhaustive exercise, V̇w and MO2 approximately doubled whereas frequency (fr) and %U barely changed, so increased ventilatory stroke volume (Vsv) was the most important contributor to increased MO2. CR declined slightly. Values gradually returned to control conditions after 2-3 h. After voluntary feeding, short-term increases in V̇w, Vsv and MO2 were comparable to those after exercise, and fr again did not change. However, %U increased so CR declined even more. The initial peaks in V̇w, Vsv and MO2, similar to those after exercise, were likely influenced by the excitement and exercise component of voluntary feeding. However, in contrast to post-exercise fish, post-prandial fish exhibited second peaks in these same parameters at 1-3 h after feeding, and %U increased further, surpassing 85%, reflecting the true "specific dynamic action" response. We conclude that respiration in trout is much more efficient than previously believed.

Breathing versus feeding in the Pacific hagfish.

Hagfish represent the oldest extant connection to the ancestral vertebrates, but their physiology is not well understood. Using behavioural (video), physiological (respirometry, flow measurements), classical morphological (dissection, silicone injection) and modern imaging approaches (micro-MRI, DICE micro-CT), we examined the interface between feeding and the unique breathing mechanism (nostril opening, high-frequency velum contraction, low-frequency gill pouch contraction and pharyngo-cutaneous duct contraction) in the Pacific hagfish, Eptatretus stoutii. A video tour via micro-MRI is presented through the breathing and feeding passages. We have reconciled an earlier disagreement as to the position of the velum chamber, which powers inhalation through the nostril, placing it downstream of the merging point of the food and water passage, such that the oronasal septum terminates at the anterior end of the velum chamber. When feeding occurs by engulfment of large chunks by the dental plates, food movement through the chamber may transiently interfere with breathing. Swallowing is accelerated by peristaltic body undulation involving the ventral musculature, and is complete within 5 s. After a large meal (anchovy, 20% body mass), hagfish remain motionless, defaecating bones and scales at 1.7 days and an intestinal peritrophic membrane at 5 days. O2 consumption rate approximately doubles within 1 h of feeding, remaining elevated for 12-24 h. This is achieved by combinations of elevated O2 utilization and ventilatory flow, the latter caused by varying increases in velar contraction frequency and stroke volume. Additional imaging casts light on the reasons for the trend for greater O2 utilization by more posterior pouches and the pharyngo-cutaneous duct in fasted hagfish.

Brain and gills as internal and external ammonia sensing organs for ventilatory control in rainbow trout, Oncorhynchus mykiss.

Ammonia is both a respiratory gas and a toxicant in teleost fish. Hyperventilation is a well-known response to elevations of both external and internal ammonia levels. Branchial neuroepithelial cells (NECs) are thought to serve as internal sensors of plasma ammonia (peripheral chemoreceptors), but little is known about other possible ammonia-sensors. Here, we investigated whether trout possess external sensors and/or internal central chemoreceptors for ammonia. For external sensors, we analyzed the time course of ventilatory changes at the start of exposure to high environmental ammonia (HEA, 1 mM). Hyperventilation developed gradually over 20 min, suggesting that it was a response to internal ammonia elevation. We also directly perfused ammonia solutions (0.01-1 mM) to the external surfaces of the first gill arches. Immediate hypoventilation occurred. For central chemoreceptors, we injected ammonia solutions (0.5-1.0 mM) directly onto the surface of the hindbrain of anesthetized trout. Immediate hyperventilation occurred. This is the first evidence of central chemoreception in teleost fish. We conclude that trout possess both external ammonia sensors, and dual internal ammonia sensors (perhaps for redundancy), but their roles differ. External sensors cause short term hypoventilation, which would help limit toxic waterborne ammonia uptake. When fish cannot avoid HEA, the diffusion of waterborne ammonia into the blood will stimulate both peripheral (NECs) and central (brain) chemoreceptors, resulting in hyperventilation. This hyperventilation will be beneficial in increasing ammonia excretion via the Rh metabolon system in the gills not only after HEA exposure, but also after endogenous ammonia loading from feeding or exercise.

The osmorespiratory compromise in the fish gill.

August Krogh made fundamental discoveries about both respiratory gas exchange and osmo/iono-regulation in fish gills. Dave Randall and co-workers identified a tradeoff between these two functions such that high functional surface area and low diffusion distance would favour O2 uptake (e.g. exercise, hypoxia), whereas low functional surface area and high diffusion distance would favour osmo/iono-regulation (rest, normoxia). Today we call this concept the "osmorespiratory compromise" and realize that it is much more complex than originally envisaged. There are at least 6 mechanisms by which fish can change functional branchial area and diffusion distance. Three involve reorganizing blood flow pathways: (i) flow redistribution within the secondary (respiratory) lamellae; (ii) flow shunting between "respiratory" and "ionoregulatory" pathways in the filament; (iii) opening up more distal lamellae on the filament and closing non-respiratory pathways. Three more involve "reversible gill remodeling": (iv) proliferation of the interlamellar gill cell mass (ILCM); (v) proliferation of ionocytes up the sides of the lamellae; (vi) covering over the apical exposure of ionocytes by extension of pavement cells. In ways that remain incompletely understood, these mechanisms allow dynamic regulation of the osmorespiratory compromise, such that ion and water fluxes can be decoupled from O2 uptake during continuous exercise. Furthermore, hypoxia-tolerant species can reduce branchial ion and water fluxes below normoxic levels despite hyperventilating during hypoxia. In marine fish, the osmorespiratory conflict is intensified by the greater ionic and osmotic gradients from seawater to blood, but underlying mechanisms remain poorly understood.

The gaseous gastrointestinal tract of a seawater teleost, the English sole (Parophrys vetulus).

There has been considerable recent progress in understanding the respiratory physiology of the gastrointestinal tract (GIT) in teleosts, but the respiratory conditions inside the GIT remain largely unknown, particularly the luminal PCO2 and PO2 levels. The GIT of seawater teleosts is of special interest due to its additional function of water absorption linked to HCO3- secretion, a process that may raise luminal PCO2 levels. Direct measurements of GIT PCO2 and PO2 using micro-optodes in the English sole (Parophrys vetulus; anaesthetized, artificially ventilated, 10-12 °C) revealed extreme luminal gas levels. Luminal PCO2 was 14-17 mmHg in the stomach and intestinal segments of fasted sole, considerably higher than arterial blood levels of 5 mmHg. Moreover, feeding, which raised intestinal HCO3- concentration, also raised luminal PCO2 to 34-50 mmHg. All these values were higher than comparable measurements in freshwater teleosts, and also greater than environmental CO2 levels of concern in aquaculture or global change scenarios. The PCO2 values in subintestinal vein blood draining the GIT of fed fish (28 mmHg) suggested some degree of equilibration with high luminal PCO2, whereas subintestinal vein PO2 levels were relatively low (9 mmHg). All luminal sections of the GIT were virtually anoxic (PO2 ≤ 0.3 mmHg), in both fasted and fed animals, a novel finding in teleosts.

The internal CO2 threat to fish: high PCO2 in the digestive tract.

Our goal was to use novel fibreoptic sensors to make the first direct PCO2 measurements in the digestive tracts of live freshwater fish (anaesthetized, artificially ventilated, 12°C). PCO2 levels in gastrointestinal fluids were substantially higher than in blood, and were elevated after feeding. In the carnivorous, gastric rainbow trout, the mean PCO2 in various parts of the tract increased from 7-13 torr (1 torr = 0.1333 kPa) during fasting to 20-41 torr after feeding, relative to arterial levels of 3.5-4 torr. In the agastric, omnivorous goldfish, the mean gut levels varied from 10-13 torr in fasted animals to 14-18 torr in fed animals, relative to arterial levels of 5-7 torr. These elevated PCO2 values were associated with surprisingly high [Formula: see text] concentrations (greater than 40 mmol l-1) in the intestinal chyme. Incubations of food pellets with acid or water revealed endogenous PCO2 generation sufficient to explain gastric PCO2 in fed trout and anterior intestine PCO2 in fed goldfish. The impacts of possible equilibration with venous blood draining the tract are assessed. We conclude that fish are already coping with PCO2 levels in the internal gastrointestinal environment many-fold greater than those of current concern in the external environment for climate change and aquacultural scenarios.

Ventilatory sensitivity to ammonia in the Pacific hagfish (Eptatretus stoutii), a representative of the oldest extant connection to the ancestral vertebrates.

Ventilatory sensitivity to ammonia occurs in teleosts, elasmobranchs and mammals. Here, we investigated whether the response is also present in hagfish. Ventilatory parameters (nostril flow, pressure amplitude, velar frequency and ventilatory index, the last representing the product of pressure amplitude and frequency), together with blood and water chemistry, were measured in hagfish exposed to either high environmental ammonia (HEA) in the external sea water or internal ammonia loading by intra-vascular injection. HEA exposure (10 mmol l-1 NH4HCO3 or 10 mmol l-1 NH4Cl) caused a persistent hyperventilation by 3 h, but further detailed analysis of the NH4HCO3 response showed that initially (within 5 min) there was a marked decrease in ventilation (80% reduction in ventilatory index and nostril flow), followed by a later 3-fold increase, by which time plasma total ammonia concentration had increased 11-fold. Thus, hyperventilation in HEA appeared to be an indirect response to internal ammonia elevation, rather than a direct response to external ammonia. HEA-mediated increases in oxygen consumption also occurred. Responses to NH4HCO3 were greater than those to NH4Cl, reflecting greater increases over time in water pH and PNH3  in the former. Hagfish also exhibited hyperventilation in response to direct injection of isotonic NH4HCO3 or NH4Cl solutions into the caudal sinus. In all cases where hyperventilation occurred, plasma total ammonia and PNH3  levels increased significantly, while blood acid-base status remained unchanged, indicating specific responses to internal ammonia elevation. The sensitivity of breathing to ammonia arose very early in vertebrate evolution.

The effects of salinity and hypoxia exposure on oxygen consumption, ventilation, diffusive water exchange and ionoregulation in the Pacific hagfish (Eptatretus stoutii).

Hagfishes (Class: Myxini) are marine jawless craniate fishes that are widely considered to be osmoconformers whose plasma [Na+], [Cl-] and osmolality closely resemble that of sea water, although they have the ability to regulate plasma [Ca2+] and [Mg2+] below seawater levels. We investigated the responses of Pacific hagfish to changes in respiratory and ionoregulatory demands imposed by a 48-h exposure to altered salinity (25 ppt, 30 ppt (control) and 35 ppt) and by an acute hypoxia exposure (30 Torr; 4 kPa). When hagfish were exposed to 25 ppt, oxygen consumption rate (MO2), ammonia excretion rate (Jamm) and unidirectional diffusive water flux rate (JH2O, measured with 3H2O) were all reduced, pointing to an interaction between ionoregulation and gas exchange. At 35 ppt, JH2O was reduced, though MO2 and Jamm did not change. As salinity increased, so did the difference between plasma and external water [Ca2+] and [Mg2+]. Notably, the same pattern was seen for plasma Cl-, which was kept below seawater [Cl-] at all salinities, while plasma [Na+] was regulated well above seawater [Na+], but plasma osmolality matched seawater values. MO2 was reduced by 49% and JH2O by 36% during hypoxia, despite a small elevation in overall ventilation. Our results depart from the "classical" osmorespiratory compromise but are in accord with responses in other hypoxia-tolerant fish; instead of an exacerbation of gill fluxes when gas transfer is upregulated, the opposite happens.

The ventilation mechanism of the Pacific hagfish Eptatretus stoutii.

We made anatomical and physiological observations of the breathing mechanisms in Pacific hagfish Eptatretus stoutii, with measurements of nostril flow and pressure, mouth and pharyngo-cutaneous duct (PCD) pressure and velum and heart impedance and observations of dye flow patterns. Resting animals frequently exhibit spontaneous apnea. During normal breathing, water flow is continuous at a high rate (~125 ml kg-1 min-1 at 12°C) powered by a two-phase unidirectional pumping system with a fast suction pump (the velum, ~22 min-1 ) for inhalation through the single nostril and a much slower force pump (gill pouches and PCD ~4.4 min-1 ) for exhalation. The mouth joins the pharynx posterior to the velum and plays no role in ventilation at rest or during swimming. Increases in flow up to >400 ml kg-1 min-1 can be achieved by increases in both velum frequency and stroke volume and the ventilatory index (product of frequency x nostril pressure amplitude) provides a useful proxy for ventilatory flow rate. Two types of coughing (flow reversals) are described. During spontaneous swimming, ventilatory pressure and flow pulsatility becomes synchronised with rhythmic body undulations.

A putative corticosteroid hormone in Pacific lamprey, Entosphenus tridentatus.

Great efforts have been put forth to elucidate the mechanisms of the stress response in vertebrates and demonstrate the conserved response across different vertebrate groups, ranging from similarities in the activation of the hypothalamic-pituitary-adrenal axis to the release and role of corticosteroids. There is however, still very little known about stress physiology in the Pacific lamprey (Entosphenus tridentatus), descendants of the earliest vertebrate lineage, the agnathans. In this paper we demonstrate that 11-deoxycortisol, a steroid precursor to cortisol in the steroidogenic pathway, may be a functional corticosteroid in Pacific lamprey. We identified the putative hormone in Pacific lamprey plasma by employing an array of methods such as RIA, HPLC and mass spectrometry analysis. We demonstrated that plasma levels of 11-deoxycortisol significantly increased in Pacific lamprey 0.5 and 1 h after stress exposure and that lamprey corticotropin releasing hormone injections increased circulating levels of 11-deoxycortisol, suggesting that the stress response is under the control of the HPA/I axis as it is in higher vertebrates. A comprehensive understanding of vertebrate stress physiology may help shed light on the evolution of the corticosteroid signaling system within the vertebrate lineage.

Trophic transfer of micro- and nanoplastics and toxicity induced by long-term exposure of nanoplastics along the rotifer (Brachionus plicatilis)-marine medaka (Oryzias melastigma) food chain.

Microplastics (MPs) and nanoplastics (NPs) are emerging pollutants in the ocean, but their transfer and toxicity along the food chains are unclear. In this study, a marine rotifer (Brachionus plicatilis)-marine medaka (Oryzias melastigma) food chain was constructed to evaluate the transfer of polystyrene MPs and NPs (70 nm, 500 nm, and 2 µm, 2000 µg/L) and toxicity of 70 nm PS-NPs (0, 20, 200, and 2000 µg/L) on marine medaka after long-term food chain exposure. The results showed that the amount of 70 nm NPs accumulated in marine medaka was 1.24 µg/mg, which was significantly higher than that of 500 nm NPs (0.87 µg/mg) and 2 µm MP (0.69 µg/mg). Long-term food chain exposure to NPs caused microflora dysbiosis, resulting in activation of toll-like receptor 4 (TLR4) pathway, which induced liver inflammation. Moreover, NPs food chain exposure increased liver and muscle tissue triglyceride and lactate content, but decreased the protein, sugar, and glycogen content. NPs food chain exposure impaired reproductive function and inhibited offspring early development, which might pose a threat to the sustainability of marine medaka population. Overall, the study revealed the transfer of MPs and NPs and the effects of NPs on marine medaka along the food chain.

Arginine vasotocin (AVT) triggers courtship behavior without exposure to external stimuli and modulates the olfactory response of male Hynobius leechii salamanders.

To study the function of arginine vasotocin (AVT) in a specific courtship behavior (body undulation) in male Hynobius leechii, we injected various doses of AVT or an AVT V1a or V2 receptor antagonist into breeding and non-breeding males. After these injections, we placed the males alone or with breeding females in Petri dishes and measured the incidence and frequency of body undulation. Additionally, to test whether AVT modulates the olfactory response of males, we exposed breeding males that were injected with AVT to female odors and measured the same response. Both breeding and non-breeding males intraperitoneally injected with 50 or 100 µg of AVT exhibited body undulation. Additionally, breeding males intraperitoneally injected with 50 or 100 µg of AVT exhibited an increased frequency of body undulation when exposed to female odors. The intraperitoneal injection of 5, 25, or 50 µg of the AVT V1a or V2 receptor antagonist did not significantly decrease the incidence or frequency of body undulation of 100 µg AVT-injected breeding males. However, in a post-hoc experiment, the breeding males that were subcutaneously injected with 100 µg of the AVT V1a receptor antagonist exhibited a significant decrease in the frequency of body undulation, induced by exposure to females. Additionally, a central injection of 1 µg of AVT into the brain induced body undulation in breeding males. Our results show that AVT not only induces a specific courtship behavior in male H. leechii via AVT V1a receptors but also modulates the olfactory response.

Nanoplastics induce more severe multigenerational life-history trait changes and metabolic responses in marine rotifer Brachionus plicatilis: Comparison with microplastics.

Micro/nanoplastics (MPs/NPs) have attracted global attention for their potential adverse impacts on marine ecosystems. This study investigated the impacts of MPs/NPs (70 nm, 500 nm, and 2 µm) on population growth and life-history traits of marine rotifer (Brachionus plicatilis), and further explored the differences from the aspects of nutrient accumulation and metabolomic profiles. The results showed that 200 and 2000 µg/L 70 nm NPs significantly suppressed population growth, and negatively affected life span, the first spawning and breeding time, and fecundity in F0-F2 generation rotifers. Whereas 500 nm NPs and 2 µm MPs showed no effect on population growth 200 µg/L and only changed the life-history traits at the highest concentration. Moreover, 70 nm NPs were more easily accumulated in the rotifers and reduced food ingestion and nutrient accumulation, which caused more severe disruption on purine-pyrimidine metabolism, tricarboxylic acid cycle, and protein synthesis pathway compared to 500 nm NPs. Thus, the smaller the size of the plastic particles, the stronger the toxicity to the rotifers. This study provided new insights into the toxicity of MPs/NPs on marine zooplankton and proposed that metabolomics was powerful to explore the toxicity mechanisms of MPs/NPs.

Understanding ventilation and oxygen uptake of Pacific hagfish (Eptatretus stoutii), with particular emphasis on responses to ammonia and interactions with other respiratory gases.

The hagfishes are an ancient and evolutionarily important group, with breathing mechanisms and gills very different from those of other fishes. Hagfish inhale through a single nostril via a velum pump, and exhale through multiple separate gill pouches. We assessed respiratory performance in E. stoutii (31 ppt, 12 ºC, 50-120 g) by measuring total ventilatory flow ([Formula: see text]) at the nostril, velar (respiratory) frequency (fr), and inspired (PIO2) and expired (PEO2) oxygen tensions at all 12 gill pouch exits plus the pharyngo-cutaneous duct (PCD) on the left side, and calculated ventilatory stroke volume (S[Formula: see text]), % O2 utilization, and oxygen consumption (MO2). At rest under normoxia, spontaneous changes in [Formula: see text] ranged from apnea to > 400 ml kg-1 min-1, due to variations in both fr and S[Formula: see text]; "normal" [Formula: see text] averaged 137 ml kg-1 min-1, MO2 was 718 µmol kg-1 h-1, so the ventilatory convection requirement for O2 was about 11 L mmol-1. Relative to anterior gill pouches, lower PEO2 values (i.e. higher utilization) occurred in the more posterior pouches and PCD. Overall, O2 utilization was 34% and did not change during hyperventilation but increased to > 90% during hypoventilation. Environmental hypoxia (PIO2 ~ 8% air saturation, 1.67 kPa, 13 Torr) caused hyperventilation, but neither acute hyperoxia (PIO2 ~ 275% air saturation, 57.6 kPa, 430 Torr) nor hypercapnia (PICO2 ~ 1% CO2, 1.0 kPa, 7.5 Torr) significantly altered [Formula: see text]. MO2 decreased in hypoxia and increased in hyperoxia but did not change in hypercapnia. Acute exposure to high environmental ammonia (HEA, 10 mM NH4HCO3) caused an acute decrease in [Formula: see text], in contrast to the hyperventilation of long-term HEA exposure described in a previous study. The hypoventilatory response to HEA still occurred during hypoxia and hyperoxia, but was blunted during hypercapnia. Under all treatments, MO2 increased with increases in [Formula: see text]. Overall, there were lower convection requirements for O2 during hyperoxia, higher requirements during hypoxia and hypercapnia, but unchanged requirements during HEA. We conclude that this "primitive" fish operates a flexible respiratory system with considerable reserve capacity.

Is ammonia excretion affected by gill ventilation in the rainbow trout Oncorhynchus mykiss?

Ammonia (NH3 + NH4+) is the major nitrogenous waste in teleost fish. NH3 is also the third respiratory gas, playing a role in ventilatory control. However it is also highly toxic. Normally, ammonia excretion through the gills occurs at about the same rate as its metabolic production, but the branchial transport mechanisms have long been controversial. An influential review in this journal has claimed that ammonia excretion in fish is probably limited by diffusion rather than by convection, so that increases in ventilation would have negligible effect on the rate of ammonia excretion. Why then should elevated plasma ammonia stimulate ventilation? The diffusion-limitation argument was made before the discovery of Rhesus (Rh) glycoproteins and the associated metabolon in the gills, which serve to greatly increase branchial ammonia permeability under conditions of ammonia loading. Therefore, we hypothesized here that (i) in accord with the diffusion-limitation concept, changes in ventilation would not affect the rate of ammonia excretion under conditions where branchial Rh metabolon function would be low (resting trout with low plasma ammonia levels). However, we also hypothesized that (ii) in accord with convective limitation, changes in ventilation would influence the rate of ammonia excretion under conditions where diffusion limitation was removed because branchial Rh metabolon function would be high (ammonia-loaded trout with high plasma ammonia levels). We used variations in environmental O2 levels to manipulate ventilation in trout under control or ammonia-loaded conditions - i.e. hyperventilation in moderate hypoxia or hypoventilation in moderate hyperoxia. In accord with hypothesis (i), under resting conditions, ammonia excretion was insensitive to experimentally induced changes in ventilation. Ammonia-loading by NH4HCO3 infusion for 30h + increased the gill mRNA expressions of two key metabolon components (Rhcg2, V-H+-ATPase or HAT), together with a 7.5-fold increase in plasma ammonia concentration and a 3-fold increase in ammonia excretion rate. In accord with hypothesis (ii), in these fish, hypoxia-induced increases in ventilation elevated the ammonia excretion rate and lowered plasma ammonia, while hyperoxia-induced decreases in ventilation reduced the ammonia excretion rate, and elevated plasma ammonia concentration. We conclude that under conditions of natural ammonia loading (e.g. meal digestion, post-exercise recovery), diffusion-limitation is removed by Rh metabolon upregulation, such that the stimulation of ventilation by elevated plasma ammonia can play an important role in clearing the potentially toxic ammonia load.

F-series prostaglandin function as sex pheromones in the Korean salamander, Hynobius leechii.

In order to test whether prostaglandins (PGs) function as sex pheromones in Hynobius leechii, a salamander that externally fertilizes its eggs, we conducted electro-olfactogram (EOG) studies with 19 PGs, liquid chromatography tandem mass spectrometry (LC-MS/MS) analyses of female and male holding waters, and behavioral tests on selected PGs. Of the 19 PGs tested, only three induced strong EOG responses from both males and ovulated females: 15-epi-prostaglandin F2alpha (15(R)-PGF2alpha), 15-keto-prostaglandin F2alpha (15K-PGF2alpha), and 13,14-dihydro-15-keto-prostaglandin F2alpha (13,14-dh-15K-PGF2alpha). In the LC-MS/MS studies, samples of holding water from ovulated females contained higher concentrations of 15(R)-PGF2alpha, PGF2alpha, and 13,14-dh-15K-PGF2alpha than those from males or oviposited females. In the behavioral tests, only 15(R)-PGF2alpha and ovulated female holding water induced significant reproductive behavior from male salamanders. These results suggest that F-series prostaglandins function as sex pheromones in amphibians.

Acute temperature effects on metabolic rate, ventilation, diffusive water exchange, osmoregulation, and acid-base status in the Pacific hagfish (Eptatretus stoutii).

The Pacific hagfish (Eptatretus stoutii) is a representative of the most basal extant craniates, and is a marine osmoconformer with an extremely low metabolic rate (MO2 = 475 µmol O2/kg/h at 12 °C). We investigated potential physiological trade-offs associated with compensatory changes in gill ventilation and perfusion when 12 °C-acclimated hagfish were acutely exposed to 7 °C or 17 °C, as reflected in diffusive unidirectional water flux ([Formula: see text], measured with tritiated water: 3H2O), net ammonia flux (Jamm), and plasma ion and acid-base status. [Formula: see text] was high (~ 1.4 L/kg/h at 12 °C) in comparison to marine teleosts and elasmobranchs. MO2 increased linearly with temperature (R2 = 0.991), and was more sensitive (Q10 = 3.22) in the 12-7 °C range than either Jamm (1.86) or [Formula: see text] (1.35), but the pattern reversed from 12 to 17 °C (Q10s: MO2 = 2.77, Jamm = 2.88, [Formula: see text] = 4.01). Heart rate, ventilatory index (a proxy for total ventilation), and coughing frequency also increased but with different patterns. At 17 °C, plasma [Ca2+] and [Mg2+] decreased, although osmolality increased, associated with elevations in plasma [Na+] and [Cl-]. Blood pH and PCO2 were unaffected by acute temperature changes while [HCO3-] increased. Hyperoxia (PO2 > 300 Torr) attenuated the increase in [Formula: see text] at 17 °C, did not affect Jamm, and had diverse effects on plasma ion and acid-base status. Our results suggest a clear osmorespiratory compromise occurring for the diffusive water fluxes as a result of acute temperature changes in this osmoconformer.

Habitat use and home range of the endangered gold-spotted pond frog (Rana chosenica).

Because of their complex life styles, amphibians and reptiles living in wetlands require both aquatic and terrestrial buffer zones in their protected conservation areas. Due to steep declines in wild populations, the gold-spotted pond frog (Rana chosenica) is listed as vulnerable by the IUCN. However, lack of data about its movements and use of habitat prevents effective conservation planning. To determine the habitat use and home range of this species, we radio-tracked 44 adult frogs for 37 days between 10 July and 4 Nov. 2007 to observe three different populations in the breeding season, non-breeding season, and late fall. The gold-spotted pond frog was very sedentary; its daily average movement was 9.8 m. Frogs stayed close to breeding ponds (within 6.6 m), and did not leave damp areas surrounding these ponds, except for dormancy migration to terrestrial sites such as dried crop fields. The average distance of dormancy migration of seven frogs from the edge of their breeding ponds was 32.0 m. The average size of an individual's home range was 713.8 m(2) (0.07 ha). The year-round population home range, which accounts for the home ranges of a population of frogs, was determined for two populations to be 8,765.0 m(2) (0.88 ha) and 3,700.9 m(2) (0.37 ha). Our results showed that to conserve this endangered species, appropriately sized wetlands and extended terrestrial buffer areas surrounding the wetlands (at least 1.33 ha, diameter 130 m) should be protected.