Digestion of meals and assimilation of nutrients are energetically-costly endeavors for all animals, and especially for intermittently-feeding ectothermic predators. The costs of digestion are attributable to a suite of aerobically-demanding processes, and interactions of digestive organs with the functionally and anatomically diverse hearts of non-avian reptiles have been hypothesized to support these processes. Specifically, postprandial cardiac hypertrophy is thought to increase convective oxygen transport to meet the aerobic demands of digestion and assimilation, and the right-to-left shunt (R-L) of crocodilians is thought to aid in acid secretion and provision of metabolic substrates.

In Chapter 1, I examined the renowned rapid postprandial cardiac hypertrophy of Burmese pythons, predicting that because it likely exists to augment oxygen delivery during digestion, it should be triggered by an oxygen supply-demand mismatch. To test this prediction, I experimentally manipulated the magnitude of the O2 supply-demand mismatch by rendering animals anemic prior to feeding. I argued, because we found cardiac hypertrophy only in this group of manipulated animals, that an autonomically-mediated response to a “low O2 signal” resulted in altered postprandial hemodynamics and subsequent hypertrophy.

In Chapter 2, I utilized a similar experimental protocol on alligators to determine whether they normally exhibited postprandial cardiac hypertrophy, and if not, whether it could be artificially induced by rendering animals anemic prior to feeding. I found only modest postprandial increases in cardiovascular function, enlargement of only stomach and liver in fed animals, and no other differences in visceral organ mass. I argued that there are neither down-regulations following fasting nor up-regulations following feeding of most visceral organs in alligators.

In Chapter 3, I tested the hypothesis that the R-L shunt of crocodilians facilitates rate of net energy gain in crocodilians. Despite significant changes in ventricular hemodynamics and a near doubling of ventricular mass in animals with surgically-occluded left aortae, I found no consequences of elimination of the shunt. This and other findings support the idea that the R-L shunt is an evolutionary remnant which has never been selected against.

Together these results illustrate the complex and plastic interactions between the digestive and circulatory systems.

Physiologists have traditionally studied elevated metabolic states in isolation (e.g., the physiological response to physical activity). However, under natural conditions animals may perform more than one elevated metabolic state simultaneously (e.g., predator avoidance while processing a recently ingested meal) and the integrated response to such a condition is not well understood.

For my dissertation work, I used oviparous (Lamprophis fuliginosus) and viviparous (Thamnophis marcianus) snakes to test whether the oxygen delivery capacity of these species' cardiopulmonary systems would be sufficient to meet the oxygen demands from reproduction, physical activity, and digestion simultaneously (i.e., additivity). Flow-through respirometry was used to measure levels of gas exchange (i.e., oxygen consumption and carbon dioxide production) during standard conditions (i.e., standard metabolic rate, SMR), and then during reproduction, activity (while fasting), digestion of a large (i.e., 10 to 20% body mass) meal, and post-prandial activity.

Results indicate that reproduction causes significant increments in oxygen consumption for females of both species. Levels ranged from 1.4 to 3.2 fold above SMR. During physical activity (while fasting), males and females of both species exhibited peak factorial increments in oxygen consumption that ranged from ~6 to 9.6 fold above SMR; digestion resulted in peak factorial increments of ~3 to 6 fold above SMR. Contrary to predictions, males and non-reproductive females of both species exhibited a prioritization pattern of oxygen delivery during post-prandial activity (i.e., factorial increments in oxygen consumption were similar to or lower than levels attained during activity while fasting). When reproduction and digestion were combined, female Lamprophis fuliginosus exhibited a prioritization pattern of oxygen delivery, while Thamnophis marcianus exhibited a more-than-additive pattern of oxygen delivery (i.e., there was a 30-50% increase in oxygen consumption above non-reproductive levels). Contrary to predictions, during the combination of reproduction and post-prandial activity females of both species exhibited a prioritization pattern of oxygen delivery.

Overall, results from my dissertation work indicate that these species' cardiopulmonary systems' capacity for oxygen delivery is limited to increments of approximately 9 to 10 fold above SMR. In most cases, activity performance (i.e., time to exhaustion, TTE) was maintained, possibly at the expense of digestive and reproductive processes.

Over the past two decades, global aquaculture production has seen significant growth, particularly in Vietnam with the use of air-breathing fishes. However, the sudden economic importance of air-breathing species, such as Pangasianodon hypophthalmus, has resulted in a significant knowledge gap surrounding their basic biology. A fundamental understanding of how these animals work and how they interact with their environment is vital for the establishment of data-driven policies and best practice and essential for optimizing their growth and production. The aim of this work is to provide the aquaculture industry with the knowledge base necessary to fulfill it’s potential in improving food security and meeting the sustainable development goals of our planet in the 21st century and beyond.

The content of this dissertation consists of several studies characterizing different aspects of adapting, both behaviorally and physiologically, to aquatic hypercapnia and are as follows: Chapter 1 explores the effect on pulmonary respiration when brachial CO2 exchange is disrupted either though aquatic hypercapnia or forced emersion in Polypterus lapradei. Chapter 2 compares how air-breathing is used differently in three Vietnamese aquaculture species when encountered with aquatic hypoxia and hypercapnia. Chapter 3 investigates the role of ecophysiology on the use of air-breathing in the avoidance behaviors to either hypoxia and hypercapnia in Po. lapradei and Pa. hypophthalmus. Chapter 4 uses blood stress bioindicators to assess if hypercapnia levels in Pa. hypophthalmus aquaculture ponds represent a significant physiological challenge. We found Pa. hypophthalmus does not respond from a respiratory, behavioral, or blood biomarker perspective when exposed to levels of hypercapnia typically found in aquaculture (34 mmHg CO2). This demonstrates the extreme tolerance of Pa. hypophthalmus to aquatic hypercapnia acquired though millions of years of evolution to its natural environment, as well as suggest a lack of potential benefit for regulating CO2 in aquaculture ponds.

Much of this research focuses on P. hypophthalmus due to its central role in aquaculture, yet several other air-breathing fish species were used as a comparative approach to investigate how uniquely evolved respiratory systems respond differently. My hope is that that is work can provide scientific data informing the impact of hypercapnia in aquaculture practices, and at the same time, demonstrate the diversity of evolutionary solutions to an environmental challenge that exist among a fascinating group of organisms, the air-breathing fishes.

Objectives

Design

Methods

Results

Conclusions

Herpes simplex virus infects both epithelial cells and neuronal cells of the human host. Although HSV assembly has been studied extensively in cultured epithelial and neuronal cells, cultured neurons are biochemically, physiologically and anatomically significantly different than mature neurons in vivo. Therefore, it is imperative that viral maturation and assembly be studied in vivo. To study viral assembly in vivo, we have inoculated wild-type and replication-defective viruses into the posterior chamber of mouse eyes and followed infection in retinal ganglion cell bodies and axons. We used PCR techniques to detect viral DNA and RNA and EM immunohistochemistry and western blotting to detect viral proteins in specific portions of the optic tract. This approach has shown that viral DNA replication is necessary for viral DNA movement into axons. Movement of viral DNA along ganglion cell axons occurs within capsid-like structures at the speed of fast axonal transport. These studies show the combined use of intravitreal injections of replication-defective viruses and molecular probes allow the genetic analysis of essential viral replication and maturation processes in neurons in vivo. They also provide novel direct evidence for the axonal transport of viral DNA and support for the subassembly hypothesis of viral maturation in situ.