Thesis (M. Med. Sc.)--University of Hong Kong, 1991.

Thesis (M.Med.Sc.)--University of Hong Kong, 1991. / Also available in print.

Thesis (Ph. D.)--Michigan State University, 2004. / Includes bibliographical references (leaves 111-121). Also available online (PDF file) by a subscription to the set or by purchasing the individual file.

Thesis (Ph. D.)--Michigan State University, 2004. / Includes bibliographical references (leaves 111-121).

Corticotropin-releasing factor (CRF) is the primary hormone of the hypothalamo-pituitary adrenal axis (HPA-axis). In addition to its endocrine function, it has been proposed that CRF acts as a neurotransmitter. The widespread distribution of CRF immunoreactivity and CRF receptors in the rat central nervous system (CNS) supports this theory. Immunohistochemical studies have demonstrated high levels of CRF immunoreactivity the rat hypothalamus, a brain region involved in the regulation and integration of a variety of endocrine and autonomic homeostatic mechanisms. CRF has been shown to be involved in a number of these activities such as blood pressure control, food and water intake, behaviour and emotional integration. Many of these activities demonstrate progressive dysfunction as ageing proceeds. The aim of this thesis was to investigate the regulation of CRF in the rat CNS, particularly over the period of maturation and ageing. Tissue extraction and peptide radioimmunoassay (RIA) techniques were developed in order to measure regional CRF concentrations as a function of age in the rat CNS. Seven brain regions were examined including the hypothalamus, pituitary, medulla oblongata, pons, cerebral cortex, cerebellum and midbrain. Three age ranges were investigated: 3 – 4 weeks, 4 – 5 months and 14 – 18 months, representing young, mature and old age groups. Data for the tissues of individual rats from each age group were analysed using one-way analysis of variance (ANOVA) with post-hoc Scheffé tests (SPSS Release 6 for Windows, 1989 – 1993). CRF were detected in measurable quantities in all brain regions examined. Different age-related patterns of change were observed in each brain region. CRF concentrations (ng/g tissue) were highest in the pituitaries of young rats and were significantly reduced over the period of maturation (P< 0.05). However, the high CRF concentration of the young rat pituitary was likely to be a factor of the smaller tissue mass. Although the absolute CRF content (ng/tissue) of this tissue appeared to decline with maturation and ageing, the reduction was not significant (P>0.05). Therefore the pituitary of the young rat was relatively enriched with CRF per gram tissue. The highest CRF concentration in mature and aged rats was measured in the hypothalamus, in accordance with previous immunohistochemical studies. Hypothalamic CRF concentrations (ng/g tissue) demonstrated no significant alterations with maturation and ageing. The absolute CRF content (ng/tissue) of the hypothalamus was significantly less in the young rat compared to mature and aged animals, however this was accompanied by a smaller tissue mass (P<0.05). The CRF concentrations (ng/g tissue) of the rat cerebral cortex and medulla oblongata demonstrated significant reduction with advancing age (P<0.05), however in both cases this appeared to be due to significant increases in mean tissue mass. The absolute CRF content of these tissues (ng/tissue) were not significantly different over the period of maturation and ageing (P>0.05). CRF concentration (ng/g tissue) and absolute content (ng/tissue) of the pons demonstrated a trend to increase with advanced age in the rat, however this was not significant in both cases (P>0.05). Of interest were the significant increases observed in the CRF concentrations of the cerebellum and midbrain (ng/g tissue with advanced ageing (P<0.05). Significant increases were also observed in the mean tissue mass and absolute CRF content (ng/tissue) of these regions in aged rats (P<0.05). These findings perhaps indicate increased CRF synthesis and or decreased CRF turnover in these tissues with advancing age. The second stage of these studies examined age-related alterations in basal and potassium-stimulated hypothalamic CRF and overflow over the period of maturation and ageing in the rat, and required the preliminary development of an in vitro tissue superfusion system. The concomitant release of the co-modulatory compound, neuropeptide Y (NPY) was also measured. NPY has been shown to positively regulate CRF release and gene expression in the hypothalamus. In addition, NPY has been demonstrated to be involved in a number of hypothalamic activities, including blood pressure control and food intake regulation. Hypothalamic superfusion data were analysed using one factor repeated measures ANOVA (SPSS Release 6 for Windows, 1989-1993) followed by least significant difference tests ( Snedecor and Cochran, 1967) to enable both time and age comparisons. Basal hypothalamic CRF overflow was unaltered with maturation and ageing in the rat. Potassium stimulation (56 mM) elicted a significant 2 – 3 fold increase in hypothalamic CRF overflow across age groups (P<0.05). Stimulated hypothalamic CRF overflow was significantly greater in the young rat compared to the mature and aged animals (P<0.05). The enhanced response to depolarizing stimulus was observed at an age when the absolute CRF content of the hypothalamus was significantly less that of other age groups. It is possible that the enhanced responsiveness of the young rat may be of survival advantage in life threatening situations. Basal hypothalamic NPY overflow was much less than that of CRF, and potassium stimulation resulted in a very different age-related profile. The hypothalamic NPY response to depolarization was significantly reduced in the young rat and declined significantly with advanced ageing (P<0.05). The contrasting profiles of stimulated CRF and NPY overflow may indicate the activity of alternative regulatory factors present in the hypothalamus, whose activity may also be affected in an age-related manner. The final stage of these studies examined the nature of NPY modulation of hypothalamic CRF overflow in the mature rat. The facilitatory effect of NPY on hypothalamic CRF overflow was confirmed. The application of NPY (0.1 µM) significantly increased CRF overflow approximately 4 fold of basal (P<0.05). In addition, the role of the NPY-Y1 receptor was investigated by the prior application of Y1 receptor antagonists, GW1229 (0.05 µM). At this concentration GW1229 significantly reduced hypothalamic CRF overflow induced by perfusion with NPY (0.1 µm), P<0.05. It was concluded the Y1 receptor does have a role in the regulation of hypothalamic CRF overflow by NPY.