The gulf toadfish (Opsanus beta) is a member of a group of teleosts that have retained their ornithine urea cycle (OUC) allowing them to excrete nitrogenous waste in the form of urea (ureotely). Urea-­N for the entire day is excreted in 1-­2 quick pulsing events (1-­3 h). This study evaluated the hypothesis that urea-­N pulsing events in gulf toadfish can be triggered by social signals from conspecifics via a specific waterborne messenger. Using a crowding protocol, we found that pre-­conditioned seawater induced a secondary urea pulsing event in naïve conspecifics. Furthermore, it was revealed that other factors such as signal concentration and donor body mass relay information to recipients as well. Fractionation of pre-­conditioned seawater was carried out to narrow possible signal candidates and the aqueous portion was found to contain the active molecule. Ammonia was found to be an important factor controlling the response of toadfish to pre-conditioned seawater.

The sonic muscle of toadfish is the fastest vertebrate muscle ever measured, and the rate of transport of Ca2+ and dissociation of cross-bridge are also fatest. Parvalbumins are Ca2+-binding proteins present in vertebrate muscle, and they can aid muscle relaxation. Several isoforms of parvalbumin had been identified and presented in variable proportion in different kinds of muscles (e.g. red muscle, white muscle and pink muscle). Both male and female grunting toadfish (Allenbatrachus grunniens) have intrinsic sonic muscles attached on swim bladders. The morphology of male and female sonic muscle was compared, and no significant differences in both length, width, thickness and weight were found. SDS-PAGE and western blotting were used to determine the total parvalbumin expression and identify the parvalbumins from sonic muscle and body white muscle. There were no significant differences in total parvalbumin expression in sonic muscle and body white muscle. The result indicates that there is no positive correlation between high content of parvalbumins and speed of muscle relaxation. In native-PAGE, two and four parvalbumin isoforms were identified from sonic muscle and body white muscle, respectively. The estimated size of Parv1, Parv2 and Parv3 size in grunting toadfish¡¦s sonic muscle were 10kDa, 10.5kDa and 10.5kDa, respectively, and the isoelectric points of Parv1, Parv2 and Parv3 in grunting toadfish were 4.77, 4.58 and 4.42, respectively. In the sonic muscle, the major parvalbumin isoform was parvalbumin isoform 1 (Parv1), which comprised more than 94% of total parvalbumin, and parvalbumin isoform 2 (Pav2) comprised vi only 5% of total parvalbumin content. In body white muscle, on the other hand, the major isoform was parvalbumin isoform 2 (Parv2) which comprised 58% of toal parvalbumin. Both Parv1 (with Parv1a) and Parv3 comprised about 20%. Parvalbumin isoforms were be discussed. The result supports that Parv1 has a highest effect on the relaxation of the grunting-toadfish¡¦s sonic muscle.

The gulf toadfish (Opsanus beta) is a member of a group of teleosts that have retained their ornithine urea cycle (OUC) allowing them to excrete nitrogenous waste in the form of urea (ureotely). Urea-­N for the entire day is excreted in 1-­2 quick pulsing events (1-­3 h). This study evaluated the hypothesis that urea-­N pulsing events in gulf toadfish can be triggered by social signals from conspecifics via a specific waterborne messenger. Using a crowding protocol, we found that pre-­conditioned seawater induced a secondary urea pulsing event in naïve conspecifics. Furthermore, it was revealed that other factors such as signal concentration and donor body mass relay information to recipients as well. Fractionation of pre-­conditioned seawater was carried out to narrow possible signal candidates and the aqueous portion was found to contain the active molecule. Ammonia was found to be an important factor controlling the response of toadfish to pre-conditioned seawater.

The gulf toadfish (Opsanus beta) is a member of a group of teleosts that have retained their ornithine urea cycle (OUC) allowing them to excrete nitrogenous waste in the form of urea (ureotely). Urea-­N for the entire day is excreted in 1-­2 quick pulsing events (1-­3 h). This study evaluated the hypothesis that urea-­N pulsing events in gulf toadfish can be triggered by social signals from conspecifics via a specific waterborne messenger. Using a crowding protocol, we found that pre-­conditioned seawater induced a secondary urea pulsing event in naïve conspecifics. Furthermore, it was revealed that other factors such as signal concentration and donor body mass relay information to recipients as well. Fractionation of pre-­conditioned seawater was carried out to narrow possible signal candidates and the aqueous portion was found to contain the active molecule. Ammonia was found to be an important factor controlling the response of toadfish to pre-conditioned seawater.

Blood sugar levels were determined in toadfish and catfish by the Folin reducing procedure and a specific enzymatic glucose procedure. The difference between the two values was considered to be the rest reduction (RR). After alloxan injection, the RR level increased greatly reaching a peak at one hour and then decreasing. The glucose level rose more slowly. An attempt was made to study these changes in blood by paper chromatography using silver nitrate to detect reducing compounds. In normal blood, glucose, an unidentified compound, and several other reducing compounds present in small amounts were separated. The large amount or RR present after alloxan could not be detected by silver nitrate. The significance of the results was discussed and future investigations were suggested. / Thesis / Master of Science (MSc)

Intestinal HCO3- secretion and the excretion of resultant CaCO3 precipitates have become a recognized characteristic of seawater osmoregulation in teleosts; however, this is the first report of this osmoregulatory strategy outside of teleosts and also includes evidence for its use in green turtles, Chelonia mydas. Furthermore, the effects of feeding on intestinal HCO3- secretion were newly investigated in teleosts. Intestinal base secretion via apical Cl-/HCO3- exchange was found to increase following feeding, at a magnitude sufficient to offset the "alkaline tide" commonly associated with digestion. Intestinal HCO3- secretion in marine teleosts draws HCO3- from both endogenous (via hydration of intracellular CO2) and serosal (blood) sources, of which serosal HCO3- was found to contribute a greater proportion to the elevated postprandial intestinal base secretion measured in gulf toadfish, Opsanus beta. The mechanism by which this serosal HCO3- crosses the basolateral membrane for subsequent secretion into the intestinal lumen was confirmed in toadfish to be a basolateral Na+/HCO3- co-transporter, tfNBCe1. Furthermore, the isolated intestinal tissue was found to have a high metabolic rate in both control and postprandial toadfish, with respect to that of the whole animal, and shows a considerable specific dynamic action (SDA) response to feeding. Overall, this dissertation provides evidence for the widespread use of intestinal HCO3- secretion as a strategy of marine osmoregulation across aquatic taxa, and also for its newly recognized involvement in postprandial acid-base balance.

Fish sound production has been associated with courtship and spawning behavior. Acoustic recordings of fish sounds can be used to identify distribution and behavior. Passive acoustic monitoring (PAM) can record large amounts of acoustic data in a specific area for days to years. These data can be collected in remote locations under potentially unsafe seas throughout a 24-hour period providing datasets unattainable using observer-based methods. However, the instruments must withstand the caustic ocean environment and be retrieved to obtain the recorded data. This can prove difficult due to the risk of PAMs being lost, stolen or damaged, especially in highly active areas. In addition, point-source sound recordings are only one aspect of fish biogeography. Passive acoustic platforms that produce low self-generated noise, have high retrieval rates, and are equipped with a suite of environmental sensors are needed to relate patterns in fish sound production to concurrently collected oceanographic conditions on large, synoptic scales. The association of sound with reproduction further invokes the need for such non-invasive, near-real time datasets that can be used to enhance current management methods limited by survey bias, inaccurate fisher reports, and extensive delays between fisheries data collection and population assessment. Red grouper (Epinephelus morio) exhibit the distinctive behavior of digging holes and producing a unique sound during courtship. These behaviors can be used to identify red grouper distribution and potential spawning habitat over large spatial scales. The goal of this research was to provide a greater understanding of the temporal and spatial distribution of red grouper sound production and holes on the central West Florida Shelf (WFS) using active sonar and passive acoustic recorders. The technology demonstrated here establishes the necessary methods to map shelf-scale fish sound production. The results of this work could aid resource managers in determining critical spawning times and areas. Over 403,000 acoustic recordings were made across an approximately 39,000 km2 area on the WFS during periods throughout 2008 to 2011 using stationary passive acoustic recorders and hydrophone-integrated gliders. A custom MySQL database with a portal to MATLAB was developed to catalogue and process the large acoustic dataset stored on a server. Analyses of these data determined the daily, seasonal and spatial patterns of red grouper as well as toadfish and several unconfirmed fish species termed: 100 Hz Pulsing, 6 kHz Sound, 300 Hz FM Harmonic, and 365 Hz Harmonic. Red grouper sound production was correlated to sunrise and sunset, and was primarily recorded in water 15 to 93 m deep, with increased calling within known hard bottom areas and in Steamboat Lumps Marine Reserve. Analyses of high-resolution multibeam bathymetry collected in a portion of the reserve in 2006 and 2009 allowed detailed documentation and characterization of holes excavated by red grouper. Comparisons of the spatially overlapping datasets suggested holes are constructed and maintained over time, and provided evidence towards an increase in spawning habitat usage. High rates of sound production recorded from stationary recorders and a glider deployment were correlated to high hole density in Steamboat Lumps. This research demonstrates the utility of coupling passive acoustic data with high-resolution bathymetric data to verify the occupation of suspected male territory (holes) and to provide a more complete understanding of effective spawning habitat. Annual peaks in calling (July and August, and November and December) did not correspond to spawning peaks (March - May); however, passive acoustic monitoring was established as an effective tool to identify areas of potential spawning activity by recording the presence of red grouper. Sounds produced by other species of fish were recorded in the passive acoustic dataset. The distribution of toadfish calls suggests two species (Opsanus beta and O. pardus) were recorded; the latter had not been previously described. The call characteristics and spatial distribution of the four unknown fish-related sounds can be used to help confirm the sources. Long-term PAM studies that provide systematic monitoring can be a valuable assessment tool for all soniferous species. Glider technology, due to a high rate of successful retrieval and low self-generated noise, was proven to be a reliable and relatively inexpensive method to collect fisheries acoustic data in the field. The implementation of regular deployments of hydrophone-integrated gliders and fixed location passive acoustic monitoring stations is suggested to enhance fisheries management.

Sperm competition is a widely-recognized and powerful selective force. Male accessory glands are organs found across animal taxa that can influence sperm performance, and thus may be selected for in competitive contexts. In fishes, these organs are in fact rare, but display great diversity in form and function across species. Although the accessory gland is known to play a role in mate attraction, parental care, fertilization, or post-copulatory competition in a few select species, the role of this organ remains a mystery in most species. Many fishes with accessory glands also exhibit alternative reproductive tactics (ARTs), which add an extra layer of complexity to how species respond to sperm competition. Because males of different ARTs typically experience different levels of sperm competition risk, it’s possible they may differentially invest in accessory glands to overcome this competition. In this thesis, I used the plainfin midshipman fish (Porichthys notatus), a species with both ARTs and an accessory gland, to experimentally investigate the role of the accessory gland in sperm competition and uncover how this organ may differ between ARTs. Over a two-year period, I studied tactic-specific investment in the accessory gland in fish from the beaches of British Columbia. I also examined the effects of seminal fluid, produced in part by the accessory gland, on sperm performance and morphology. I found that males adopting the ‘guarder’ male tactic invested more in one region (the lobules) of this organ, while males adopting the smaller ‘sneaker’ male tactic invested more the other region of the gland (the nodes). Using data collected over five years, I found that guarder males also invested more in their whole accessory glands. Additionally, I report that sperm swam faster in the presence of seminal fluid, and seminal fluid increased sperm head size in both male tactics and increased midpiece size in guarder males. These results suggest that the plainfin midshipman accessory gland may have dual functions, one of which may be to aid sperm competitive ability through enhancements in swimming speed and potentially more successful sperm morphology. Taken together, the results of my thesis improve our knowledge of the role of non-sperm components like seminal fluid and the accessory gland in sperm competition, and demonstrate how species with ARTs can have varying physiological responses to such competition. Only a handful of studies have considered the effects of seminal fluid on sperm performance. By examining sperm competition in a more biologically relevant way and incorporating the effects of a little-studied organ that impacts sperm competition, we should be able to more generally and accurately appreciate the dynamics of post-copulatory competition and fertilization. / Thesis / Master of Science (MSc)