Recent studies, primarily in males, have shown that postexercise cutaneous vasodilation is attenuated by baroreceptor unloading subsequent to lower body venous pooling. However, gender differences in the control of cutaneous circulation may exist given that females appear to show a reduced tolerance to orthostatic challenge and, an attenuated responsiveness in mechanisms that regulate arterial pressure. We evaluated the hypothesis that females would demonstrate a greater reduction in postexercise mean arterial pressure (MAP) and concurrently a greater increase in the postexercise core temperature at which onset of cutaneous vasodilation occurred as compared to males. Fourteen subjects (7 males and 7 females) of similar age and fitness status remained seated resting for 15 min or cycled for 15 min at 70% of peak oxygen consumption followed by 15 min of seated recovery. (Abstract shortened by UMI.)

Background. Knee braces have been found to provide limited stability to the ACL deficient knee in situations where the knee is loaded during sporting movements. Different adaptation strategies have been found between patients that can cope with the injury and patients that cannot. One of the expected changes can be muscle activation characteristics of the injured knee during strenuous activity with and without a functional knee brace. Methods. Three dimensional (3D) kinematic and electromyography (EMG) data were collected from each participant for ten consecutive gait cycles during running on a treadmill under both braced and unbraced conditions. Participants were administered the "Knee Outcome Survey Activities of Daily Living Scale" to distinguish functional and non-functional candidates. Findings. No significant differences on 3D kinematics and EMG data were noted between functional and non-functional participants, thus data analysis focused on comparisons of bracing conditions for one combined group. Bracing significantly increased total range of motion in the sagittal plane (p<0.05) and reduced total range of motion in the transverse plane (p<0.05). Muscle activity at heel-strike showed a consistent trend to increase for the hamstrings and decrease for the quadriceps under the braced when compared to the unbraced condition. Interpretations. Our findings indicated that bracing the ACL deficient knee altered kinematics of the injured leg while running. Tendencies towards reductions in quadriceps and increases in hamstrings activity at heel-strike provides added stability to the injured knee. These changes to bracing further support the mechanical and the proprioceptive contributions of the functional knee brace to protect the ACLD knee.

Gait initiation is a temporary movement between upright posture and steady-state gait. The activation of several postural muscles has been identified to precede changes observed in vertical reaction force. Previous research examining gait initiation has concentrated on the electromyographic activity of muscles of the lower limbs. Few studies, however, have looked at recruitment patterns of the muscles of the thigh and trunk. This study was conducted to determine the recruitment patterns and the roles of certain muscles of the trail and lead lower limbs and trunk for the duration from quiet stance to trail leg toe-off. Eleven healthy participants initiated gait with their right leg. Electromyographic data were collected bilaterally from the erector spinae, tensor fasciae latae, adductor magnus and tibialis anterior muscles. In addition, force platform data were recorded for the duration of quiet stance to toe-off of the trail limb. (Abstract shortened by UMI.)

Follicle stimulating hormone (FSH) is an important survival factor in the ovarian follicular development. Inhibitor of apoptosis proteins (IAPs) is a family of intracellular anti-apoptosic proteins. X-linked IAP (XIAP) has been shown to be involved in multiple biological activities (e.g. inhibition of caspase activities, promotion of ubiquitin-proteasome-mediated protein degradation, regulation of cell signaling pathways). The present thesis research project examines: (1) the role and gonadotropic regulation of XIAP expression in rat granulosa cells during ovarian follicular development and atresia; (2) the possible involvement of intra-ovarian factors such as transforming growth factor alpha (TGFalpha) in the FSH-induced XIAP expression and follicular development; and (3) the signal pathways involved in the gonadotropic up-regulation of XIAP during follicular development in vitro. A follicle culture system coupled to an adenoviral gene manipulation procedure has been established. FSH significantly increased follicular growth as evident by increases in follicular size, cell number and DNA contents in vitro. While cultured pre-antral or early-antral follicles showed a low XIAP content and evidence of apoptosis in the absence of FSH, gonadotropin addition increased XIAP content and suppressed apoptosis. At low FSH concentration, adenoviral XIAP sense cDNA expression increased follicular cell XIAP and DNA contents, reduced apoptosis, and enhanced follicular growth, while XIAP antisense elicited opposite responses. FSH-induced XIAP up-regulation appeared mediated, in part, by the secretion and action of follicular TGFalpha. In cultured rat follicles, FSH-stimulated estradiol production, TGFalpha secretion, XIAP expression and follicular growth were suppressed by intra-follicular injection of a neutralizing anti-TGFalpha antibody or addition of the estradiol antagonist ICI 182780 to the culture media. These results support my hypothesis that the FSH induces follicular growth by stimulating granulosa cell proliferation via theca TGFalpha secretion and action in response to increased granulosa cell estradiol synthesis. Since the promoter region of XIAP gene has nuclear kappa B (NFkappaB) binding site, it is possible that the transcription of XIAP is mediated via NFkappaB activation. FSH increased rat granulosa cell XIAP mRNA abundance and protein content. While the gonadotropin induced granulosa cell NFkappaB translocation from cytoplasm to nucleus and increased NFkappaB-DNA binding activity, pretreatment with an NFkappaB translocation inhibitor suppressed FSH-stimulated XIAP expression. (Abstract shortened by UMI.)

Xenopus laevis oocytes are physiologically arrested at the first meiotic prophase. Progesterone reinitiates meiosis (maturation) through inhibition of an oocyte adenylyl cyclase (AC) and reduction of intracellular cAMP. However, the mechanism by which progesterone regulates AC activity and cAMP level still remains unclear. In this thesis, I summarize work I conducted that collectively helps elucidate how high levels of cAMP might be achieved in G2 arrested oocytes. In Chapter 2, I describe our finding that inhibiting endogenous G-protein betagamma subunits, through the use of two structurally distinct Gbetagamma scavengers, causes hormone-independent oocyte maturation. In contrast, overexpression of Xenopus Gbeta1, alone or together with bovine Ggamma2, inhibits progesterone-induced oocyte maturation. These results for the first time implicate that an endogenous G protein coupled receptor system releases a Gbetagamma complex as the dominant meiosis inhibitor. Chapter 3 describes my research aiming to reveal the identity of the oocyte AC responsible for generating meiosis-inhibiting cAMP. I provide further evidence here that the ability of Gbetagamma to inhibit meiosis is attributed to the activation of an endogenous AC, rather than other possible Gbetagamma effectors. Through molecular cloning and biochemical characterization, I discovered that the likely AC candidate is Xenopus AC7, an isoform that is activated by Gbetagamma, but only in the presence of GTP-bound Gsalpha. The identification of xAC7 suggests that the maintenance of high levels of cAMP may require the cooperation of Gsalpha and Gbetagamma. Finally, in Chapter 4, I describe our efforts in identifying the GPCR(s) responsible for activating the cAMP signaling in prophase-arrested oocytes. A screening of known antagonists of GPCR(s) led to the identification of ritanserin, a potent antagonist of serotonin receptors, as a potent maturation inducer in Xenopus oocytes. Pharmacological and molecular studies, however, have ruled out the involvement of a known serotonin receptor in meiosis arrest. Instead, the most likely candidate is a "constitutively activated" GPCR that bears structural similarities to Xenopus serotonin receptor 7.

Intercellular communication via secreted signaling molecules is essential for the generation of a normally patterned central nervous system (CNS). The mammalian retina, which is neuroectoderm derived, is an excellent model system in which to study signaling events in patterning, cell proliferation and diversification in the CNS. Although the identity and function of some signaling molecules that regulate retinal development are known, the function of many others, especially members of the Wnt family, has yet to be well characterized. Wnt ligands have been established as critical regulators of multiple developmental processes in a variety of organs and tissues during embryogenesis. To ascertain the function of Wnt signaling in the context of retinal development, I examined the expression of Wnt pathway components in the mouse neural retina. I showed that Wnt2b (formerly known as Wnt13), Wnt receptors and Wnt antagonists are expressed within and adjacent to the distal part of the eyecup, the ciliary margin (CM). beta-gal staining in the eyes of TCF/Lef-LacZ mice, a canonical Wnt pathway reporter mouse strain, revealed the highest level of reporter activity in the CM throughout retinal development. Thus, I hypothesized that canonical Wnt signaling plays a role in the specification and formation of the CM and its derivatives. To test the above hypothesis, I used several approaches to activate Wnt signaling in retinal explants from TCF/Lef-LacZ reporter mice. Ectopic expression of Wnt2b did not activate Wnt reporter activity, while over expression of Wnt3a, a well-characterized canonical Wnt pathway activator, induced reporter gene expression in the CM, but after a considerable delay. Treatment with lithium, a well-known Wnt pathway agonist, resulted in rapid upregulation of Wnt reporter activity, and alterations in morphology and gene expression that were consistent with induction of a CM identity. The expression of stabilized beta-catenin, the key mediator of canonical Wnt pathway activation, was then targeted to the peripheral retina by using a Cre-loxP approach. These transgenic mice exhibited a dramatic increase in beta-catenin-dependent signaling, as well as an expansion of the CM/ciliary epithelium in Cre-expressing region of the retina. Taken together, the findings presented in this thesis indicate that beta-catenin-mediated signaling can promote the development of the CM and its derivatives.

High salt diet cardiac fibrosis by activating cardiac aldosterone . We hypothesized that enhanced cardiac aldosterone production by high salt intake may increase left ventricle (LV) weight and cardiomyocyte size and induce fibrosis in both ventricles. Regular salt (0.6%) or high salt (8%) diet, either without or with spironolactone (20 or 80 mg/kg/day) were given to Wistar rats for 4 and 8 weeks. Both LV weight and cardiomyocyte cross-sectional diameter were increased significantly by high salt diet after 4 and 8 weeks. Both LV and right ventricle (RV) collagen and fibrosis as well as mean arterial pressure (MAP) remained unchanged after 4 weeks, but increased significantly after 8 weeks on high salt diet. Spironolactone (80 mg/kg) prevented the high salt-induced increases in LV weight and cardiomyocyte cross-sectional diameter as well as LV and RV collagen and fibrosis, and attenuated the increase in MAP. Spironolactone (20 mg/kg) was somewhat less effective. The high salt induced changes in cardiac and cardiomyocyte hypertrophy, and cardiac fibrosis were prevented by spironolactone, which is consistent with the concept that cardiac aldosterone mediates these cardiac effects of high salt diet. Role of brain renin-angiotensin-aldosterone-system (RAAS) in cardiac remodeling post-MI. To assess the contribution of the brain RAAS in the activation of the cardiac RAAS and remodeling post-MI, Wistar rats with intracerebroventricular (icv) infusion of spironolactone and transgenic (TG) rats deficient in brain angiotensinogen were studied. An MI was induced by acute coronary artery ligation. Spironolactone was administered by icv infusion (100 ng/h) or orally (80 mg/kg/day) for 6 weeks post-MI in Wistar rats. TG rats and control Sprague-Dawley (SD) rats were followed for 8 weeks. MI decreased LV peak systolic pressure (LVPSP) and LV dP/dt max and increased LV end diastolic pressure (LVEDP) and plasma catecholamines and serum aldosterone, which were prevented or attenuated by both icv and oral spironolactone at 6 weeks post-MI. MI increased internal circumferences, cardiomyocyte diameter, fibrosis, laminin and fibronectin, and aldosterone in the LV or and RV, which were also significantly prevented/inhibited by both icv and oral spironolactone at 6 weeks post-MI in Wistar rats as well as at 8 weeks post-MI in TG rats. Magnitude of beneficial effects of icv spironolactone at low doses was largely equal to that achieved with its oral administration at much higher doses, indicating that in addition to other sites of actions, aldosterone appears to activate central nervous system (CNS) pathways and thereby influences peripheral mechanisms involved in cardiac remodeling. The findings in TG rats support the pivotal role of locally produced angiotensin II in the brain in cardiac remodeling post-MI. The brain RAAS appears to activate a cascade of events, among others an increase in cardiac aldosterone, which play a major role in cardiac remodeling post-MI.

The eggs of the dengue fever vector Aedes aegypti possess the ability to undergo an extended quiescence period hosting a fully developed first instar larvae within its chorion. As a result of this life history stage, pharate larvae can withstand months of dormancy inside the egg where they depend on stored reserves of maternal origin. This adaptation known as pharate first instar quiescence, allows A. aegypti to cope with fluctuations in water availability. An examination of this fundamental adaptation has shown that there are trade-offs associated with it. Aedes aegypti mosquitoes are frequently associated with urban habitats that may contain metal pollution. My research has demonstrated that the duration of this quiescence and the extent of nutritional depletion associated with it affects the physiology and survival of larvae that hatch in a suboptimal habitat; nutrient reserves decrease during pharate first instar quiescence and alter subsequent larval and adult fitness. The duration of quiescence compromises metal tolerance physiology and is coupled to a decrease in metallothionein mRNA levels. My findings also indicate that even low levels of environmentally relevant larval metal stress alter the parameters that determine vector capacity. My research has also demonstrated that extended pharate first instar quiescence can elicit a plastic response resulting in an adult phenotype distinct from adults reared from short quiescence eggs. Extended pharate first instar quiescence affects the performance and reproductive fitness of the adult female mosquito as well as the nutritional status of its progeny via maternal effects in an adaptive manner, i.e., anticipatory phenotypic plasticity results as a consequence of the duration of pharate first instar quiescence and alternative phenotypes may exist for this mosquito with quiescence serving as a cue possibly signaling the environmental conditions that follow a dry period. M findings may explain, in part, A. aegypti’s success as a vector and its geographic distribution and have implications for its vector capacity and control.

acase@tulane.edu

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