Browsing by Author "John Godwin, Committee Member"

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Circadian organization in Japanese quail: Ocular clocks are pacemakers
(2005-12-13) Steele, Christopher Todd; Betty Black, Committee Member; Herbert Underwood, Committee Chair; John Godwin, Committee Member; Tom Siopes, Committee Member
A growing body of evidence supports the idea that circadian clocks in the eye can influence extra-ocular rhythms in both mammalian and non-mammalian vertebrates. Perhaps the most salient example is seen in Japanese quail where ocular clocks are pacemakers in the circadian system since eye removal causes all quail tested to become arrhythmic in constant darkness (DD). Moreover, the eyes play an endocrine role in the quail circadian system since one-third to one-half of the blood melatonin is of ocular origin and rhythmic blood-borne melatonin can affect circadian body temperature and activity rhythms. This investigation was designed to define further the role of the eye as a pacemaker in the quail?s circadian system. Given that quail maintain robust circadian rhythms of body temperature in prolonged DD, the two putative ocular pacemakers in an individual bird must maintain the same phase; otherwise, a consolidated circadian output could not be generated. Furthermore, if ocular clocks are indeed pacemakers, the two putative pacemakers should rapidly regain coupling after being forced out of phase. These predictions were confirmed by demonstrating that 1) the ocular melatonin rhythms of the two eyes maintained phase for at least 57 days in DD and 2) after ocular pacemakers were forced out of phase by alternately patching the eyes (AP) in constant light, two components of body temperature were observed that fused into a consolidated rhythm after 5?6 days in DD, demonstrating pacemaker recoupling. The ability to maintain phase in DD and rapidly recouple after out-of phase entrainment demonstrates that the eyes are strongly coupled pacemakers that work in synchrony to drive circadian rhythmicity in Japanese quail. Since the eyes are indeed coupled pacemakers, the mechanisms of pacemaker coupling were also investigated. Quail were subjected to one of several surgical procedures including optic-nerve section, superior cervical ganglionectomy, ciliary ganglionectomy, and melatonin implantation. Birds were then subjected to AP protocols to determine if these manipulations would affect pacemaker recoupling. Optic-nerve section and melatonin implantation lengthened the recoupling time of ocular pacemakers to 8 and 7 days, respectively, and each were significantly different than control groups. However, neither the superior cervical ganglia nor the ciliary ganglia appear to transmit circadian information between ocular pacemakers. Therefore, both neural (optic nerve) and hormonal (melatonin) coupling mechanisms contribute to ocular pacemaker coupling. It is known that eye-removal abolishes rhythmicity of body temperature and activity rhythms in quail. We hypothesized that the pineal-SCN complex in quail is incapable of maintaining rhythmicity in the absence of ocular pacemakers. This hypothesis was confirmed since birds blinded by complete eye-removal displayed arrhythmic blood melatonin profiles in DD further supporting the idea that ocular clocks are pacemakers in quail. Based on current and previous data, the eye plays a major role within the quail circadian system that can, at least in part, exert its control via the cyclic synthesis and release of melatonin. If this view of the quail circadian system is correct, one must assume that the ocular pacemaker is an autonomous oscillator. In the present study we demonstrate that quail retinal explants possess a light-entrainable circadian clock that can synthesize and secrete melatonin rhythmically in vitro for at least five days in DD. Taken together, the present experiments fully support the hypotheses that 1) the eyes are the loci of tightly coupled circadian pacemakers in Japanese quail, 2) ocular pacemakers are coupled via neural and hormonal mechanisms, 3) in the absence of ocular pacemakers, the pineal-SCN complex is incapable of maintaining rhythmic blood melatonin profiles in DD, and 4) the quail's eye possesses an autonomous, light-entrainable circadian clock capable of rhythmic production and secretion of melatonin.
Coevolution Between Grasshopper mice (Onychomys spp.) and Bark and Striped Scorpions (Centruroides spp.)
(2004-11-29) Rowe, Ashlee Hedgecock; James Gilliam, Committee Member; John Godwin, Committee Member; Kenneth Pollock, Committee Member; Harold Heatwole, Committee Chair
Asymmetrical selection has been proposed as the strongest argument for rejecting hypotheses of coevolutionary arms races between predators and prey. In many cases there is evidence of increased investment by the prey in response to the predator, but no evidence of increased investment by the predator in response to the prey, thus producing asymmetry in selection. However, selection against a predator may be increased when the interaction is with a "dangerous" prey. Predators are most likely to respond evolutionarily to potentially lethal prey. This study employs grasshopper mice (Onychomys spp.) and bark and striped scorpions (Centruroides spp.) as a model to test the hypothesis that interspecific interaction between a predator and a potentially lethal prey will result in behavioral and physiological adaptations that reciprocally mediate their interaction (i.e., coevolution). Bark scorpions (Centruroides exilicauda) and striped scorpions (Centruroides vittatus) produce a potent venom containing neurotoxins that selectively bind to the ion-channels of vertebrates. Vertebrate-specific neurotoxins may produce lethal effects in mammals, especially small mammals. Southern grasshopper mice (Onychomys torridus) and Mearns' grasshopper mice (O. arenicola) are known to be voracious predators on scorpions. Southern grasshopper mice are broadly sympatric with bark scorpions in the Sonoran Desert, and Mearns' grasshopper mice are broadly sympatric with striped scorpions in the Chihuahuan Desert. The third species in this genus, the northern grasshopper mouse (O. leucogaster) is broadly allopatric with Centruroides spp. In a preliminary study, both southern and Mearns' grasshopper mice demonstrated resistance to bark and striped scorpion neurotoxins. The evolution of toxic venom and resistance to that venom strongly suggests a coevolutionary relationship between Onychomys spp. and Centruroides spp. To test this hypothesis, I evaluated the predator-prey relationship between grasshopper mice and their toxic scorpion prey during staged feeding trials. Additionally, I compared the geographic patterns of venom resistance in all three species of grasshopper mice with geographic patterns of venom toxicity in bark and striped scorpions. Results from the feeding study demonstrated that grasshopper mice do not distinguish between toxic and non-toxic species of scorpions; mice attacked, incapacitated, and consumed bark and striped scorpions without hesitation and as effectively as they attacked crickets and non-toxic scorpions in the genus Vaejovis. The feeding experiments indicate that grasshopper mice have the ability to prey on bark and striped scorpions in habitats where they co-occur. Venom resistance analyses demonstrated that all three species of grasshopper mice have evolved some resistance to the vertebrate-specific neurotoxins produced by Centruroides spp. The assays show that patterns of venom toxicity in Centruroides and venom resistance in Onychomys co-vary geographically, both within and among species; i.e., populations of Onychomys interacting with the most toxic populations of Centruroides were extremely resistant; populations of Onychomys interacting with only moderately toxic populations of Centruroides were only moderately resistant; and populations of Onychomys not sympatric with Centruroides were only weakly resistant. Such systematic covariation between venom toxicity in the scorpions and venom resistance in the mice is consistent with a coevolutionary, arms race hypothesis.
The effect of endocrine disruptors on the monogamous pine vole (Microtus pinetorum).
(2003-04-15) Engell, Miles Dean; John G. Vandenbergh, Committee Chair; John Godwin, Committee Member; Roger Powell, Committee Member; Gerald LeBlanc, Committee Member
Since the discovery in the 1960s that synthetic chemicals in the environment could alter normal hormonal functioning in wildlife, the study of endocrine disruptors has grown rapidly. Most xenobiotic endocrine disrupting chemicals (EDCs) arise from sources such as pesticides, and either mimic estrogens or act as antiandrogens. Previous research has focused largely on physiological and anatomical endpoints. The objective of this research was to examine the effects of EDCs on the behaviors and corresponding neural receptor distribution associated with monogamy in the pine vole. Female pine voles were orally administered an EDC or corn oil control throughout gestation and lactation of pups. As adults, these pre- and neonatally exposed pups were examined for behavioral and physiological alterations. AGD and seminal vesicle weights were masculinized in flutamide exposed males, suggesting flutamide may have acted as androgen rather than antiandrogen. Vinclozolin acted as antiandrogen. Diethylstilbestrol (DES) and methoxychlor are both estrogenic EDCs. Preference for the mate versus a stranger was assessed via a three-chambered apparatus. Flutamide treated males did not show a preference for their mate, but instead spent most of their time alone in the neutral chamber. Their parental behaviors were unaffected. Vinclozolin treated males' preference for their mate was unaltered, but their paternal responsiveness decreased. DES exposed females were more aggressive toward the stranger than any other treatment group. Methoxychlor females showed a strong trend toward spending more time alone in the neutral chamber, like the flutamide males. There were no differences in maternal behavior. Receptor autoradiography was performed to examine any effect on arginine vasopressin receptor binding in males and oxytocin receptor binding in females. The cingulate cortex had decreased binding in flutamide males and methoxychlor females. These findings demonstrate that exposure to endocrine disrupting compounds during early development can alter adult brain receptor distribution and mating system behaviors.
Genetic and Temperature Manipulation of Southern Flounder (Paralichthys lethostigma) for the Production of Monosex Populations
(2005-07-11) Morgan, Andrew Jackson; John Godwin, Committee Member; Russell Borski, Committee Member; Harry V. Daniels, Committee Chair
Experiments were conducted to establish methods to produce an all female population of southern flounder (Paralichthys lethostigma) gynogens. The first series of experiments was to optimize the protocol for using UV irradiated black sea bass (Centropristis striata) sperm for the induction of meiogynogenesis. Milt was collected from multiple black sea bass, pooled, and diluted 1:10 using Ringer's solution. The diluted milt was irradiated using an UV Crosslinker with doses ranging from 0-130 J/cm2. The optimal UV dose of 70 J/cm2 was determined by observing a 10 % motility, motility duration of approximately 1 minute, and giving the highest percent hatch (7.73+1.57 %) which was statistically different (P=0.05) than the control. Pressure shock was used to retain the 2nd polar body in southern flounder eggs. Application of 8500 psi for 6 minutes was initiated at varying times 1, 2, and 3 minutes post fertilization with no pressure acting as the negative control. Based upon percentage hatch and survival, the optimal time to apply pressure was either 1 or 2 minutes post fertilization, which was temperature dependent (1 minute at 18oC and 2 minutes at 16oC.) The overall average hatch varied with initiation times and ranged from 1.48 + 0.52% (1 min) to 0.61 + 0.11% (3 min) of the fertilized eggs (average 70.9 + 12.8% fertility.) The development of these techniques can allow aquaculturists to take advantage of the better growth rates of female southern flounder by producing all female offspring. Three preliminary experiments were also conducted to establish methods to produce cloned, monosex populations of southern flounder through mitotic gynogenesis. The first experiment determined the timing of the first mitotic division of the southern flounder egg. Eggs were fertilized with UV irradiated (70 J/cm2) black sea bass sperm and allowed to normally divide between 17-20oC. There was no correlation between spawning temperature, the timing of the first mitotic division, or the rate of mitosis. The timing of first cleavage varied from 63-88 minutes post fertilization. Once eggs began the first cellular division, 90 % of the eggs had completed this stage within 40 minutes. Using these data, southern flounder eggs were fertilized with UV irradiated (70 J/cm2) black sea bass sperm to create mitogynogens. Percent fertilization was 44.7% and percent hatch was 0%, although some eggs advanced to the blastula stage. Triploidy hatch percentages were 18.0-42.7 % of the normal diploid control percent hatch. A third series of experiments were designed to test for sex differences among populations from North Carolina and Texas southern flounder raised under the same conditions and shifted to higher temperatures. At the end of the study, there was no significant difference in lengths or weights between the treatments. The flounder that remained in 23oC indicated a trend for larger growth for the Texas flounder while the 28oC tended to grow larger for the North Carolina flounder (not significant.) Final sex ratios were 100% male for all treatments. An additional experiment was conducted to determine the window of temperature dependent sex determination in North Carolina southern flounder. A sample was shifted to 28oC at 60 mm and 90 mm in duplicate to determine when sex differentiation occurred. The North Carolina southern flounder in this study grew better in 23oC until 60 mm and then their growth rate increased in 28oC until approximately 125 mm.
Host Location by Adults and Larvae of Specialist Herbivore Heliothis Subflexa G. (Lepidoptera: Noctuidae)
(2007-03-26) Benda, Nicole Denise; Coby Schal, Committee Co-Chair; John Godwin, Committee Member; George Kennedy, Committee Member; Fred Gould, Committee Co-Chair
Heliothis subflexa is a specialist herbivore, whose larvae feed on fruits of Physalis species (Solanaceae). Although Physalis surface extracts have been shown to elicit attraction and oviposition by adult ovipositing females, little else was known as to how adults locate their host plants for oviposition. Also, larvae are sometimes dislodged from the plant when the plant abscises the fruit they are feeding on. The ability of larvae to re-locate their host plant from the ground had not been evaluated. The purpose of this research was to evaluate the host location and oviposition behavior of H. subflexa adult females and larvae using common garden studies, behavioral assays in the lab and in the field, and direct field observations. H. subflexa showed a clear preference for some Physalis species over others in the common garden. Plant age and date also affected incidence of eggs and fruit damage on the Physalis plants. In laboratory two-choice simultaneous and sequential behavioral assays, H. subflexa showed a preference for P. philadelphica over tobacco. However, preference for Physalis was not to the complete exclusion of tobacco and a portion of eggs were placed on tobacco (˜20% on average). Field observations found that this was not a lab artifact. Females laid an average of 20% of eggs on non-hosts in the field, though most were placed very close to the plant. An evaluation of the host location ability of neonates found a high fitness cost to ovipositing on non-hosts. The ability of larger larvae (third and fifth instar) to re-locate the host plant after being dislodged from the plant via fruit abscission was affected by larval size, plant architecture, and host-specific behavior as compared to a close relative, H. virescens.