Despite the abundance of oxygen in atmospheric air relative to water, the initial loss of respiratory surface area and accumulation of carbon dioxide in the blood of amphibious fishes during emersion may result in hypoxemia. Given that the ability to respond to low oxygen conditions predates the vertebrate invasion of land, we hypothesized that amphibious fishes maintain O 2 uptake and transport while emersed by mounting a co-opted hypoxia response. We acclimated the amphibious fish Kryptolebias marmoratus, which are able to remain active for weeks in both air and water, for 7 days to normoxic brackish water (15‰, ~21 kPa O 2 ; control), aquatic hypoxia (~3.6 kPa), normoxic air (~21 kPa) or aerial hypoxia (~13.6 kPa). Angiogenesis in the skin and bucco-opercular chamber was pronounced in air-versus water-acclimated fish, but not in response to hypoxia. Aquatic hypoxia increased the O 2 -carrying capacity of blood via a large (40%) increase in red blood cell density and a small increase in the affinity of hemoglobin for O 2 (P 50 decreased 11%). In contrast, air exposure increased the hemoglobin O 2 affinity (decreased P 50 ) by 25% without affecting the number of red blood cells. Acclimation to aerial hypoxia both increased the O 2 -carrying capacity and decreased the hemoglobin O 2 affinity . These results suggest that O 2 transport is regulated both by O 2 availability and also, independently, by air exposure. The ability of the hematological system to respond to air exposure independent of O 2 availability may allow extant amphibious fishes, and may also have allowed primitive tetrapods to cope with the complex challenges of aerial respiration during the invasion of land.KEY WORDS: Hemoglobin, Oxygen-carrying capacity, Hemoglobin-oxygen affinity, Air-breathing organ, Air-breathing fish, Mangrove rivulus INTRODUCTIONThe transition from aquatic to terrestrial life represents a major step in vertebrate evolution because the physical conditions between these environments are dramatically different. Oxygen solubility is relatively low in water and even well-oxygenated aquatic environments contain only about 3% of the O 2 available in atmospheric air (Dejours, 1988). Low concentrations of aquatic O 2 , particularly in hypoxic habitats, have often been hypothesized to be one of the driving forces behind the evolution of amphibious or terrestrial life histories because invasion of land would allow animals to exploit the O 2 -rich aerial environment (Graham, 1997). RESEARCH ARTICLEDepartment of Integrative Biology, University of Guelph, Guelph, ON N1G 2W1, Canada.*Author for correspondence (aturko@uoguelph.ca) Received 4 July 2014; Accepted 15 September 2014Taking advantage of aerial O 2 presents several challenges for fishes. Water-breathing fishes exchange respiratory gases across the gills but during emersion the gill lamellae typically collapse and coalesce, reducing the surface area available for respiration. Accumulation of CO 2 in the blood of emersed fishes, resulting from the low solubility of CO 2 in...

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The inner egg capsule of embryos of the yellow-spotted salamander (Ambystoma maculatum) are routinely colonized by green algae, such as Oophila amblystomatis, that supply O(2) in the presence of light and may consume nitrogenous wastes, forming what has been proposed to be a mutualistic relationship. Given that A. maculatum have been reported to breed in acidic (pH <5.0) and neutral lakes, we hypothesized that low water pH would negatively affect these symbiotic organisms and alter the gradients within the jelly mass. Oxygen gradients were detected within jelly masses measured directly in a natural breeding pond (pH 4.5-4.8) at midday in full sunlight. In the lab, embryo jelly masses reared continuously at pH 4.5 had lower P(O)₂and higher ammonia levels relative to jelly masses held at pH 8.0 (control). Ammonia and lactate concentrations in embryonic tissues were approximately 37%-93% higher, respectively, in embryos reared at water pH 4.5 compared with pH 8.0. Mass was also reduced in embryos reared at pH 4.5 versus pH 8.0. In addition, light conditions (24 h light, 12L : 12D, or 24 h dark) and embryonic position (periphery vs. center) in the jelly mass affected P(O)₂but not ammonia gradients, suggesting that algal symbionts generate O(2) but do not significantly impact local ammonia concentrations, regardless of the pH of the water. We conclude that chronic exposure to acidic breeding ponds had a profound effect on the microenvironment of developing A. maculatum embryos, which in turn resulted in an elevation of potentially harmful metabolic end products and inhibited growth. Under acidic conditions, the expected benefit provided by the algae to the salamander embryo (i.e., high O(2) and low ammonia microenvironment) is compromised, suggesting that the A. maculatum-algal mutualism is beneficial to salamanders only at higher water pH values.

SUMMARYIn rainbow trout development, a switch occurs from high-affinity embryonic hemoglobin (Hb) and round, embryonic erythrocytes to lower-affinity adult Hb and oval, adult erythrocytes. Our study investigated the early ontogeny of rainbow trout blood properties and the hypoxia response. We hypothesized that hypoxia exposure would delay the ontogenetic turnover of Hb and erythrocytes because retention of high-affinity embryonic Hb would facilitate oxygen loading. To test this hypothesis we developed a method of efficiently extracting blood from individual embryos and larvae and optimized several techniques for measuring hematological parameters on microliter (0.5-2.0 μl) blood samples. In chronic hypoxia (30% of oxygen saturation), stage-matched embryos and larvae possessed half the Hb concentration, erythrocyte counts and hematocrit observed in normoxia. Hypoxia-reared larvae also had threefold to sixfold higher mRNA expression of the embryonic Hb α-1, β-1 and β-2 subunits relative to stage-matched normoxia-reared larvae. Furthermore, in hypoxia, the round embryonic erythrocytic shape persisted into later developmental stages. Despite these differences, Hb-oxygen affinity (P 50 ), cooperativity and the Root effect were unaltered in hypoxia-reared O. mykiss. The data support our hypothesis that chronic hypoxia delays the ontogenetic turnover of Hb and erythrocytes, but without the predicted functional consequences (i.e. higher than expected P 50 ). These results also suggest that the Hb-oxygen affinity is protected during development in chronic hypoxia to favor oxygen unloading at the tissues. We conclude that in early trout development, the blood-oxygen transport system responds very differently to chronic hypoxia relative to adults, possibly because respiration depends relatively more on oxygen diffusion than convection.

Shorebirds depend on staging sites in the Gulf of Mexico that are frequently subject to pollution by oil and its toxic constituents, polycyclic aromatic hydrocarbons (PAHs). It was hypothesized that PAH contamination lowers staging site quality for migratory shorebirds, with consequences for fueling and departure timing. Sediment total PAH concentrations were measured at six staging sites along the Texas and Louisiana Gulf Coast. Sites in Louisiana were expected to have higher total PAH concentrations as they were more heavily impacted by the Deepwater Horizon oil spill. From 2015 to 2017, 165 Sanderling (Calidris alba) and 55 Red knots (C. canutus) were captured at these same sites during their northward migration (late April to mid May). Mass, body morphometrics, and plasma metabolite measurements were taken to determine fuel loads and fueling rates, and a subset of birds (120 Sanderling and 39 Red knots) received a coded radio tag to determine departure dates using the Motus telemetry array. Compared to Texas sites, sediment in Louisiana had higher total PAH concentrations, dominated by heavier 6 ring indeno[1,2,3-cd]pyrene (48%). Plasma metabolite profiles suggested that fueling rates for Sanderling, but not Red knots, tended to be lower in Louisiana, and both species departed later than the study average from Louisiana. However, multiple factors, including migration patterns, food supply, and other contaminants, also likely influenced fueling and departures. PAH contamination in the Gulf of Mexico remains an ongoing issue that may be impacting the staging site quality and migration timing of long-distance migratory birds.