Preeclampsia is a devastating disorder of pregnancy characterized by high blood pressure, proteinuria, headache, renal glomerular endotheliosis, multi-organ system failure, and fetal and maternal demise. As reviewed in Chapter I, not much is known about the pathogenesis of preeclampsia, contributing to a lack of biomarkers and treatments for the disease. In Chapter II, we review arginine vasopressin, a circulating neuropeptide hormone with important fluid balance and cardiovascular actions. Vasopressin binds to numerous receptors throughout the body to elicit its effects and is associated with various disease states. In Chapter III, we discuss evidence for vasopressin as an etiology of preeclampsia. Specifically, that vasopressin secretion is elevated very early in pregnancies affected by preeclampsia and infusion of vasopressin into pregnant C57BL/6J mice causes physiological features similar to those seen in preeclampsia. In Chapter IV, we show that vasopressin administration during mouse pregnancy models specific subtypes of preeclampsia through time- and receptor-specific mechanisms. The role of angiotensin 1a receptors on vasopressin-producing cells in fluid homeostasis is shown in Chapter V and their role in metabolism is depicted in Chapter VI. Overall, we conclude that vasopressin is an important mediator of features of preeclampsia and that angiotensin 1a receptors on vasopressin-producing cells are important for normal fluid homeostasis.

Thesis (M.Sc. (Pharmaceutics))--North-West University, Potchefstroom Campus, 2007.

Aggression is a feature of many clinical disorders including autism, Alzheimer’s disease, bipolar disorder, and schizophrenia. The available treatment options act to prevent impulsive aggression through modulation of GABAergic and dopaminergic pathways which come with metabolic and dyskinetic side effects. The mechanism underlying aggressive motivation, however, has not been elucidated. In addition, previous studies have been heavily biased towards males of various species. Mimicking changes in day length, or photoperiod, in the laboratory is a natural manipulation used to examine seasonal changes in aggressive behavior in many species. In response to the reduction in the duration of light exposure, animals undergo gonadal regression and become reproductively quiescent. During this non-breeding season in male photoperiod-responsive animals, aggressive behavior increases significantly. Although studies have shown offensive aggression remains elevated in female rodents, seasonal regulation of this behavior in females has not been thoroughly studied. The neuropeptide arginine-vasopressin (AVP) has been implicated in the facilitation of aggressive behavior in male rodents and fishes; therefore, it is useful to examine AVP as a modulator of seasonal aggression in females. Because the actions of AVP in female social behavior may be hormonally-dependent, we investigated the hormonal mechanisms that regulate the expression of AVP receptors and the behavioral actions of AVP on aggression. In addition to changes in gonadal steroid hormones during the non-breeding season, we identified photoperiod-dependent alterations in adrenal hormone secretion as AVP plays a role in regulation of hypothalamic-pituitary-adrenal axis (HPA) activity and anxiety-like behaviors in animal models.

In most animal species, reproductive success depends critically on precopulatory or solicitational behaviors that occur prior to mating. The specific sensory systems and behavioral strategies employed in precopulatory behaviors vary across species; in all cases, however, animals must be able to identify potential mating partners and solicit sexual interest. Female Syrian hamsters (Mesocricetus auratus) engage in multiple forms of precopulatory behaviors that are preferentially expressed to males or their odors, including vaginal scent marking and sexual odor preference. Conspecific odors relevant for precopulatory behaviors are processed by a network of forebrain areas that includes the bed nucleus of the stria terminalis (BNST) and the medial preoptic area (MPOA). The precise functional and neurochemical mechanisms whereby these areas regulate the expression of precopulatory behaviors, however, are unknown. Therefore, the aim of this dissertation is to address the following research questions: (1) Is the neuropeptide oxytocin (OT), acting within BNST or MPOA, necessary for the normal expression of odor-­guided precopulatory behaviors? (2) Is BNST or (3) MPOA required for the preferential expression of vaginal marking or investigation towards male odors?, and (4) Does OT interact with social odor processing to regulate vaginal marking? We found that blockade of OT receptors (OTRs) in MPOA and BNST decreased vaginal marking to male odors. There was no effect of OTR blockade on sexual odor preference. Selective lesions of BNST also disrupted preferential vaginal marking responses to male odors, without affecting sexual odor preference. In contrast, lesions of MPOA disrupted odor preference without affecting vaginal marking responses. Finally, central blockade of OTRs eliminated the normal pattern of increased activation of neurons to male vs. female odors in BNST, but not MPOA. Considered together, these results suggest that OT normally acts within BNST to drive preferential vaginal marking responses to male odors via selective facilitation of neural responses to these odors, and further, that there are separate and distinct neural circuits that regulate different forms of odor-guided precopulatory behaviors in females.

published_or_final_version / abstract / toc / Anatomy / Doctoral / Doctor of Philosophy

Stress impacts upon all organisms and a robust stress response is required for adaptive interactions of the organism with the environment. In most higher organisms, an individual’s response to stress is mediated by the hypothalamic pituitary adrenal (HPA) axis. Inappropriate regulation of this axis can cause debilitating mental health disorders including depression and anxiety. These disorders can affect an individual’s ability to interact and respond appropriately as different situations arise. An important component of this axis is the vasopressin V1b receptor (V1bR), which mediates adrenocorticotropin (ACTH) secretion from the anterior pituitary in response to stimulation by arginine vasopressin (AVP). AVP also potentiates the ACTH secretion mediated by corticotropin-releasing hormone type 1 receptor (CRH-R1) in response to corticotropin- releasing hormone (CRH) stimulation. Both the V1bR and CRH-R1 are G protein coupled receptors (GPCRs). A common feature of GPCR signalling is desensitisation of the response following prolonged or repeated exposure to an agonist. Phosphorylation of the receptor is one of the mechanisms of desensitisation. This directly, or indirectly, results in rapid and reversible uncoupling of the receptor from its heterotrimeric guanine nucleotide binding protein (G-protein). Previous research has shown that G protein coupled receptor kinases (GRKs) are key phosphorylators involved in the molecular mechanism of GPCR desensitisation. One of the mains goals of the research carried out in the Mason laboratory is to examine the molecular mechanisms of V1bR desensitisation. The current short term aim is to examine the potential role for GRKs in this mechanism. It is difficult to study a single receptor type and the molecular mechanisms involved in its regulation in a system larger than a cell based assay. As the proposed method of assessing the involvement of GRKs in desensitisation of the V1bR is to use RNA interference (RNAi) to knock down the expression of the GRKs, primary cell cultures of pituitary corticotrophs are an inappropriate choice. This is due to a number of factors, including the difficulty involved in transfecting primary cells, and the difficulty involved in interpreting the results from primary cell culture experiments as these cultures are composed heterogenous population of cells. Therefore, the main aim of this research was to develop a model cell system from an immortalised cell line, stably-transfected with the V1bR, in which the involvement of GRKs in the molecular mechanism of V1bR desensitisation could be studied. Development of stably-transfected cell lines requires substantial preliminary work and planning in order to produce a successful outcome. Once developed, characterisation of the clonal cell lines is required. The preliminary work involved determining the cell proliferation rate of the parental cell line, plasmid sub-cloning and production of a large quantity of plasmid DNA, optimisation of the antibiotic selection conditions, and optimisation of the transfection protocol, as well as modification of the inositol phosphate (IP) assay protocol. The V1bR activates the phospholipase Cβ (PLCβ) second messenger signalling pathway in response to stimulation with AVP. This results in the production of IPs and therefore, measurement of IPs in response to AVP stimulation of cells labelled with myo-[³H]inositol can be used as an indicator of functional V1bR expression. In this research a total of nine clonal cell lines resistant to the antibiotic G418 were generated. Initial testing of these lines indicated that four probably expressed the V1bR and these were selected for characterisation in greater detail. All of these four lines showed significantly increased IP production in response to AVP stimulation (P<0.05; t-test). A significant decrease in IP production in response to AVP stimulation following an AVP pre-treatment was also seen with all four lines (P<0.05; t-test). Current evidence therefore suggests that the V1bR in these clonal cell lines signals and desensitises in the normal way. Although further characterisation of the clonal cell line is desirable, the data to date indicate that these lines should be considered to provide an appropriate model system for examining the molecular mechanisms involved in the regulation of the V1bR. It appears that there are some minor differences in signalling between the clonal cell lines and therefore this should be a consideration when deciding which line is most appropriate to use for investigating a particular question.

The aim of this study was to investigate in vitro transdermal diffusion of a small peptide namely arginine vasopressin (AVP) with the aid of the novel PheroidTM drug delivery system. Generally, peptides seem unfit for transdermal permeation, but it was thought prudent to explore the suitability of this lipid-based system after success was achieved with entrapment of tuberculostatics, bacteria and viruses. Bestatin (a selective aminopeptidase inhibitor) was employed to circumvent any skin-related degradation of the active. Therefore, the effect of bestatin on the preservation of AVP during diffusion was investigated. Vertical Franz cell diffusion studies were conducted with female abdominal skin, with AVP at a concentration of 150 pglml in the donor phase and Hepes buffer as the receptor phase over a twelve-hour period. To prove entrapment of AVP within the lipid structures of the PheroidsTM, fluorescentlylabelled samples were monitored by means of confocal laser scanning microscopy (CLSM), which revealed definite entrapment. In vitro permeation profiles for AVP exhibited a biphasic character, with the majority of permeation occurring during the first two hours. The PheroidTM delivery system proved to be advantageous when applied as delivery medium. The inclusion of bestatin has an enhancing effect on permeation probably due to its protection of AVP. / Thesis (M.Sc. (Pharmaceutics))--North-West University, Potchefstroom Campus, 2007.

No description available.

The purpose of these studies was to examine mediation and moderation processes for the influence of early adversity and current stressful circumstances on maternal sensitivity. Evidence of mediation was found in Study 1 where maternal depression and mothers’ negative appraisal of their infant mediated the influence of early adversity and low family income on maternal sensitivity. Study 1 also examined the influence of the neighborhood. A moderate-mediation model was tested where the mediating influence afore-stated was hypothesized to vary across levels of neighborhood quality. Partial evidence of moderation was found. In the context of a high quality neighborhood, mothers’ early adversity was not associated with maternal depression. Across levels of neighborhood quality, complex relationships emerged between the variables low family income, maternal depression and mothers’ appraisal of infant temperament. In a context of low neighborhood quality, there was no evidence of a direct association between low family income and maternal sensitivity, rather, low family income operated indirectly through maternal depression. In a context of high neighborhood quality, there was evidence for a direct and indirect association between low family income and maternal sensitivity. Study 2 examined associations between variation in the Arginine Vasopressin 1a receptor gene (AVPR1a) and maternal sensitivity, and whether variation in this gene moderated the influence of mothers’ early adversity on sensitivity. Mothers homozygous for the long alleles of the RS3 microsatellite were significantly less sensitive than mothers heterozygous for the long alleles and those homozygous for the short alleles. Homozygosity for the RS3 long alleles moderated the influence of mothers’ early adversity on their sensitivity; the influence of early adversity on maternal sensitivity was most pronounced for mothers with the RS3 long/long genotype. These results suggest that variation in the AVPR1a gene may be important not only for human maternal behavior, but also for stress reactivity.

The purpose of these studies was to examine mediation and moderation processes for the influence of early adversity and current stressful circumstances on maternal sensitivity. Evidence of mediation was found in Study 1 where maternal depression and mothers’ negative appraisal of their infant mediated the influence of early adversity and low family income on maternal sensitivity. Study 1 also examined the influence of the neighborhood. A moderate-mediation model was tested where the mediating influence afore-stated was hypothesized to vary across levels of neighborhood quality. Partial evidence of moderation was found. In the context of a high quality neighborhood, mothers’ early adversity was not associated with maternal depression. Across levels of neighborhood quality, complex relationships emerged between the variables low family income, maternal depression and mothers’ appraisal of infant temperament. In a context of low neighborhood quality, there was no evidence of a direct association between low family income and maternal sensitivity, rather, low family income operated indirectly through maternal depression. In a context of high neighborhood quality, there was evidence for a direct and indirect association between low family income and maternal sensitivity. Study 2 examined associations between variation in the Arginine Vasopressin 1a receptor gene (AVPR1a) and maternal sensitivity, and whether variation in this gene moderated the influence of mothers’ early adversity on sensitivity. Mothers homozygous for the long alleles of the RS3 microsatellite were significantly less sensitive than mothers heterozygous for the long alleles and those homozygous for the short alleles. Homozygosity for the RS3 long alleles moderated the influence of mothers’ early adversity on their sensitivity; the influence of early adversity on maternal sensitivity was most pronounced for mothers with the RS3 long/long genotype. These results suggest that variation in the AVPR1a gene may be important not only for human maternal behavior, but also for stress reactivity.