Short communication: Characterizing arterial and venous blood gases over the gas exchange surface, the chorioallantoic membrane, of embryonic American alligators (Alligator mississippiensis) at two points of development.

Assessments of arterial and venous blood gases are required to understand the function of respiratory organs in animals at different stages of development. We measured blood gases in the arteries entering and veins leaving the chorioallantoic membrane (CAM) in embryonic alligators (Alligator mississippiensis). The CAM accounts for virtually all gas exchange in these animals, and we hypothesized that the CAM vasculature would be larger in eggs incubated in hypoxia (10% O2 for 50% or 70% of incubation), which would be reflected in a lower partial pressure of CO2 (PCO2). Contrary to this hypothesis, our measurements revealed no effects of hypoxic incubation on PCO2, and seemingly no increase in vascularization of the CAM in response to incubation in 10% O2. PCO2 was lower on the venous side, but only significantly different from arterial blood at 70% of incubation. The calculated blood flow to the CAM increased with development and was lower in both groups of alligators that had been incubated in hypoxia. Future studies should include measurements of blood parameters taken from embryos held in conditions that mirror incubation O2 levels, in combination with direct measurements of CAM artery blood flow.

Exposure to hypoxia during embryonic development affects blood flow patterns and heart rate in juvenile American alligators during digestion.

The developmental environment can alter an organism's phenotype through epigenetic mechanisms. We incubated eggs from American alligators in 10% O2 (hypoxia) to investigate the functional plasticity of blood flow patterns in response to feeding later in life. Digestion is associated with marked elevations of metabolism, and we therefore used the feeding-induced stimulation of tissue O2 demand to determine whether there are lasting effects of developmental hypoxia on the cardiovascular response to digestion later in life. In all animals studied, digestion elicited tachycardia and an elevation of blood flow in the right aorta, left aorta, and the pulmonary artery, whereas flows in the carotid and subclavian artery did not change. We found that heart rate and systemic blood flow remained elevated for a longer time period in juvenile alligators that had been incubated in hypoxia; we also found that the pulmonary blood flow was elevated at 24, 36, and 48 h. Collectively, our findings demonstrate that exposure to hypoxia during incubation has lasting effects on the hemodynamics of juvenile alligators 4 years after hatching.

Does the left aorta provide proton-rich blood to the gut when crocodilians digest a meal?

Reptiles have the capacity to differentially perfuse the systemic and pulmonary vascular circuits via autonomic regulation of the heart and the vascular trees. While this aptitude is widely recognized, the role of 'shunting' as a homeostatic mechanism to match convective transport with tissue demand remains unknown. In crocodilians, it has been hypothesized that a pulmonary vascular bypass of systemic venous blood - a right-to-left (R-L) shunt - serves to deliver CO2-rich blood with protons needed for gastric acid secretion during digestion. This hypothesis is partially based on the unique crocodilian vascular anatomy where a left aorta (LAo) arises from the right ventricle, and appears to preferentially supply the gastrointestinal system, whereas the right aorta emerges from the left ventricle. Recent theoretical considerations imply that a R-L shunt would have minuscule effects on PCO2 , but direct measurements of blood gases in both the right and left aortae or both the right and left atria in fed animals have not been conducted. For this reason, we measured blood parameters including PO2 , PCO2 , pHe and [HCO3-] in the right and left aortae and atria following ingestion of a gavage-fed standardized meal (5% body mass). Blood samples were taken at 3, 6, 12, 24, 36 and 48 h into the digestive period to encompass the period of maximal gastric acid secretion. At no point did PCO2  or pH differ between the left and right aortae, whereas PO2  was significantly lower in the left aorta at several time points during digestion. Our findings do not support the hypothesis that a R-L shunt serves to deliver CO2 for the gastrointestinal system after feeding in crocodilians.

The electrocardiogram of vertebrates: Evolutionary changes from ectothermy to endothermy.

The electrocardiogram (ECG) reveals that heart chamber activation and repolarization are much faster in mammals and birds compared to ectothermic vertebrates of similar size. Temperature, however, affects electrophysiology of the heart and most data from ectotherms are determined at body temperatures lower than those of mammals and birds. The present manuscript is a review of the effects of temperature on intervals in the ECG of ectothermic and endothermic vertebrates rather than a hypothesis-testing original research article. However, the conclusions are supported by the inclusion of original data (Iguana iguana, N = 4; Python regius, N = 5; Alligator mississippiensis, N = 4). Most comparisons were of animals of approximately 1 kg. Compared to mammals and birds, the reptiles at 35-37 °C had 4 fold lower heart rates, 2 fold slower atrial and ventricular conduction (longer P- and QRS-wave durations), and 4 fold longer PR intervals (atrioventricular delay) and QT intervals (total ventricular repolarization). We conclude that the faster chamber activation in endotherms cannot be explained by temperature alone. Based on histology, we show that endotherms have a more compact myocardial architecture. In mammals, disorganization of the compact wall by fibrosis associates with conduction slowing and we suggest the compact tissue architecture allows for faster chamber activation. The short cardiac cycle that characterizes mammals and birds, however, is predominantly accommodated by shortening of the atrioventricular delay and the QT interval, which is so long in a 1 kg iguana that it compares to that of an elephant.

Acid-base regulation in the air-breathing swamp eel (Monopterus albus) at different temperatures.

Vertebrates reduce arterial blood pH (pHa) when body temperature increases. In water breathers, this response occurs primarily by reducing plasma HCO3- levels with small changes in the partial pressure of CO2 (PCO2 ). In contrast, air breathers mediate the decrease in pHa by increasing arterial PCO2  (PaCO2 ) at constant plasma HCO3- by reducing lung ventilation relative to metabolic CO2 production. Much less is known about bimodal breathers, which utilize both water and air. Here, we characterized the influence of temperature on arterial acid-base balance and intracellular pH (pHi) in the bimodal-breathing swamp eel, Monopterus albus This teleost uses the buccopharyngeal cavity for gas exchange and has very reduced gills. When exposed to ecologically relevant temperatures (20, 25, 30 and 35°C) for 24 and 48 h, pHa decreased by -0.025 pH units (U) °C-1 in association with an increase in PaCO2 , but without changes in plasma [HCO3-]. pHi was also reduced with increased temperature. The slope of pHi of liver and muscle was -0.014 and -0.019 U °C-1, while the heart muscle showed a smaller reduction (-0.008 U °C-1). When exposed to hypercapnia (7 or 14 mmHg) at either 25 or 35°C, M. albus elevated plasma [HCO3-] and therefore seemed to defend the new pHa set-point, demonstrating an adjusted control of acid-base balance with temperature. Overall, the effects of temperature on acid-base balance in M. albus resemble those in air-breathing amniotes, and we discuss the possibility that this pattern of acid-base balance results from a progressive transition in CO2 excretion from water to air as temperature rises.

Specialized impulse conduction pathway in the alligator heart.

Mammals and birds have a specialized cardiac atrioventricular conduction system enabling rapid activation of both ventricles. This system may have evolved together with high heart rates to support their endothermic state (warm-bloodedness) and is seemingly lacking in ectothermic vertebrates from which first mammals then birds independently evolved. Here, we studied the conduction system in crocodiles (Alligator mississippiensis), the only ectothermic vertebrates with a full ventricular septum. We identified homologues of mammalian conduction system markers (Tbx3-Tbx5, Scn5a, Gja5, Nppa-Nppb) and show the presence of a functional atrioventricular bundle. The ventricular Purkinje network, however, was absent and slow ventricular conduction relied on trabecular myocardium, as it does in other ectothermic vertebrates. We propose the evolution of the atrioventricular bundle followed full ventricular septum formation prior to the development of high heart rates and endothermy. In contrast, the evolution of the ventricular Purkinje network is strongly associated with high heart rates and endothermy.

Embryonic hypoxia programmes postprandial cardiovascular function in adult common snapping turtles (Chelydra serpentina).

Reduced oxygen availability (hypoxia) is a potent stressor during embryonic development, altering the trajectory of trait maturation and organismal phenotype. We previously documented that chronic embryonic hypoxia has a lasting impact on the metabolic response to feeding in juvenile snapping turtles (Chelydra serpentina). Turtles exposed to hypoxia as embryos [10% O2 (H10)] exhibited an earlier and increased peak postprandial oxygen consumption rate, compared with control turtles [21% O2 (N21)]. In the current study, we measured central blood flow patterns to determine whether the elevated postprandial metabolic response in H10 turtles is linked to lasting impacts on convective transport. Five years after hatching, turtles were instrumented to quantify systemic ([Formula: see text]) and pulmonary ([Formula: see text]) blood flows and heart rate (fH) before and after a ∼5% body mass meal. In adult N21 and H10 turtles, fH was increased significantly by feeding. Although total stroke volume (VS,tot) remained at fasted values, this tachycardia contributed to an elevation in total cardiac output ([Formula: see text]). However, there was a postprandial reduction in a net left-right (L-R) shunt in N21 snapping turtles only. Relative to N21 turtles, H10 animals exhibited higher [Formula: see text] due to increased blood flow through the right systemic outflow vessels of the heart. This effect of hypoxic embryonic development, reducing a net L-R cardiac shunt, may support the increased postprandial metabolic rate we previously reported in H10 turtles, and is further demonstration of adult reptile cardiovascular physiology being programmed by embryonic hypoxia.

Metabolic responses to chronic hypoxic incubation in embryonic American alligators (Alligator mississippiensis).

Chronic hypoxic incubation is a common tool used to study developmental changes in reduced O2 conditions, and it has been useful for identifying phenotypically plastic periods during ontogeny in laboratory settings. Reptilian embryos can be subjected to natural hypoxia due to nesting strategy, and recent studies have been important in establishing the phenotypic responses of several species to low developmental oxygen. In particular, the cardiovascular responses of American alligators (Alligator mississippiensis) to low developmental oxygen have been detailed, including a substantial cardiac enlargement that may support a higher mass specific metabolic rate. However, embryo mass-specific metabolic demands of hypoxic incubated alligator embryos have not been measured. In this study, alligator eggs were incubated in 10% O2 (H) or 21% O2 (N) environments for the entire course of embryonic development. Acute metabolic measures in 21% and 10% O2 were taken for both H and N groups. We hypothesized that acute 10% O2 exposure has no impact on metabolic rate of embryonic alligators, and that metabolic rate is unaffected by chronic hypoxic incubation when studied in embryos measured at 21% O2. Our findings suggest phenotypic changes resulting from hypoxic incubation early in incubation, in particular relative cardiac enlargement, enable embryonic alligators to sustain metabolic rate during acute hypoxic exposure.