Janna Crossley

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

ABSTRACT 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 () and pulmonary () 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 (). 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 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. Summary: During embryonic development in underground nests, reptiles routinely experience hypoxia, which programmes cardiovascular physiology into adulthood, dictating convective transport during periods of elevated oxygen demand.

Developmental plasticity of mitochondrial function in American alligators, Alligator mississippiensis

The effect of hypoxia on cellular metabolism is well documented in adult vertebrates, but information is entirely lacking for embryonic organisms. The effect of hypoxia on embryonic physiology is particularly interesting, as metabolic responses during development may have life-long consequences, due to developmental plasticity. To this end, we investigated the effects of chronic developmental hypoxia on cardiac mitochondrial function in embryonic and juvenile American alligators (Alligator mississippiensis). Alligator eggs were incubated in 21% or 10% oxygen from 20 to 90% of embryonic development. Embryos were either harvested at 90% development or allowed to hatch and then reared in 21% oxygen for 3 yr. Ventricular mitochondria were isolated from embryonic/juvenile alligator hearts. Mitochondrial respiration and enzymatic activities of electron transport chain complexes were measured with a microrespirometer and spectrophotometer, respectively. Developmental hypoxia induced growth restriction and increased relative heart mass, and this phenotype persisted into juvenile life. Embryonic mitochondrial function was not affected by developmental hypoxia, but at the juvenile life stage, animals from hypoxic incubations had lower levels of Leak respiration and higher respiratory control ratios, which is indicative of enhanced mitochondrial efficiency. Our results suggest developmental hypoxia can have life-long consequences for alligator morphology and metabolic function. Further investigations are necessary to reveal the adaptive significance of the enhanced mitochondrial efficiency in the hypoxic phenotype.

Development and application of sub-2-μm particle CO2-based chromatography coupled to mass spectrometry for comprehensive analysis of lipids in cottonseed extracts: UPC2analysis of cottonseed lipids

RATIONALE Refined cottonseed oil has widespread applications in the food and chemical industries. Although the major lipids comprising cottonseed oil (triacylglycerols) are well known, there are many diverse lipid species in cotton seeds that occur at much lower levels and have important nutritional or anti-nutritional properties. METHODS The lipid technical samples were prepared in chloroform. The biological samples were extracted using a mixture of isopropanol/chloroform/H2 O (2:1:0.45). The data were collected using high and low collision energy with simultaneous data collection on a time-of-flight (TOF) mass spectrometer which allowed the characterization of lipids by precursor and product ion alignment. The supercritical fluid chromatography methodology is flexible and can be altered to provide greater retention and separation. The comprehensive method was used to screen seed lipid extracts from several cotton genotypes using multivariate statistical analysis. RESULTS Method variables influencing the peak integrity and chromatographic separation for a mixture of lipids with different degrees of polarity were explored. The experiments were designed to understand the chromatographic behavior of lipids in a controlled setting using a variety of lipid extracts. Influences of acyl chain length and numbers of double bonds were investigated using single moiety standards. CONCLUSIONS The methodology parameters were examined using single moiety lipid standards and standard mixtures. The method conditions were applied to biological lipid extracts, and adjustments were investigated to manipulate the chromatography. Insights from these method variable manipulations will help to frame the development of targeted lipid profiling and screening protocols. Copyright

Contribution of active atrial contraction to cardiac output in anesthetized American alligators (Alligator mississippiensis)

ABSTRACT Ventricular filling may occur directly from the venous circulation during early diastole or via atrial contraction in late diastole. The contribution of atrial contraction to ventricular filling is typically small in mammals (10–40%), but has been suggested to predominate in reptiles. We investigated the importance of atrial contraction in filling of the ventricle in American alligators (Alligator mississippiensis) by bypassing both atria (with the use of ligatures to prevent atrial filling) and measuring the resultant effects on cardiac output in anesthetized animals. Atrial ligation had no significant effects on total systemic blood flow before or after adrenaline injection. Unexpectedly, pulmonary flow was increased following atrial ligation prior to adrenaline treatment, but was unaffected after it. These findings suggest that the atria are non-essential (i.e. redundant) for ventricular filling in alligators, at least under anesthesia, but may serve as important volume reservoirs. Summary: Ligation of both atria had little effect on cardiac output in anesthetized American alligators and, surprisingly, increased pulmonary blood flow. This suggests atria have a non-essential role in ventricular filling.

Developmental cardiovascular physiology of the olive ridley sea turtle (Lepidochelys olivacea)

Our understanding of reptilian cardiovascular development and regulation has increased substantially for two species the American alligator (Alligator mississippiensis) and the common snapping turtle (Chelydra serpentina) during the past two decades. However, what we know about cardiovascular maturation in many other species remains poorly understood or unknown. Embryonic sea turtles have been studied to understand the maturation of metabolic function, but these studies have not addressed the cardiovascular system. Although prior studies have been pivotal in characterizing development, and factors that influence it, the development of cardiovascular function, which supplies metabolic function, is unknown in sea turtles. During our investigation we focused on quantifying how cardiovascular morphological and functional parameters change, to provide basic knowledge of development in the olive ridley sea turtle (Lepidochelys olivacea). Embryonic mass, as well as mass of the heart, lungs, liver, kidney, and brain increased during turtle embryo development. Although heart rate was constant during this developmental period, arterial pressure approximately doubled. Further, while embryonic olive ridley sea turtles lacked cholinergic tone on heart rate, there was a pronounced beta adrenergic tone on heart rate that decreased in strength at 90% of incubation. This beta adrenergic tone may be partially originating from the sympathetic nervous system at 90% of incubation, with the majority originating from circulating catecholamines. Data indicates that olive ridley sea turtles share traits of embryonic functional cardiovascular maturation with the American alligator (Alligator mississippiensis) but not the common snapping turtle (Chelydra serpentina).

Thyroid hormone manipulation influences development of cardiovascular regulation in embryonic Pekin duck, Anas platyrhynchos domestica

Thyroid hormones are key regulators of avian metabolism and may play a significant role in development at hatching. To better understand the role of thyroid hormones in avian development, we examined autonomic control of heart rate and blood pressure while manipulating thyroid hormone levels in the late stage embryonic Pekin duck (Anas platyrhynchos domestica). Thyroid hormone levels were manipulated on day 24 of a 28-day incubation period with the thyroperoxidase inhibitor methimazole (MMI), triiodothyronine (T3), or saline. On day 25 of incubation, autonomic tone on cardiovascular function was studied by injections of cholinergic and adrenergic receptor antagonists. Embryos from all treatment groups expressed a cholinergic and β-adrenergic tone on heart rate at this age. Cholinergic blockade with atropine produced a larger change in heart rate in the hyperthyroid animals compared with euthyroid animals. In response to β-adrenergic blockade, hyperthyroid conditions produced a larger decrease in heart rate compared with euthyroid animals, with no change in mean arterial blood pressure. In response to α-adrenergic blockade, mean arterial blood pressure decreased in the euthyroid animals and more developed hyperthyroid animals. Collectively, the data indicate that elevated levels of T3 can influence maturation of cholinergic and adrenergic receptor-mediated cardiovascular regulation in developing Pekin ducks near the end of incubation.