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Title: Measurement of gonadotrophin and corticotrophin releasing activity of pituitary stalk blood and brain extracts under various physiological and experimental conditions
Author: Chiappa, Sharon A.
ISNI:       0000 0001 3543 8203
Awarding Body: University of Oxford
Current Institution: University of Oxford
Date of Award: 1976
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The aim of this thesis was to correlate changes in the content of (i) gonadotrophin-releasing hormone (GnRH) in brain extracts, pituitary stalk and peripheral plasma, and (ii) hypothalamic corticotrophin-releasing activity (CR-activity) with the patterns of secretion of luteinizing hormone (LH), follicle stimulating hormone (FSH) and adrenocorticotrophic hormone (ACTH) from the anterior pituitary gland and corticosterone from the adrenal gland, under various physiological and experimental conditions. Most measurements were made on samples obtained from rats of the Wistar strain, but the results of some studies in the human are reported in Chapter 8. Radioimmunoassay techniques were used to measure GnRH, LH, FSH and ACTH concentrations, while a competitive protein binding method was used to determine plasma corticosterone concentrations. CR-activity was assessed using an in-vitro assay involving short-term incubation of hemipituitaries: the amount of ACTH released into the medium was measured by radioimmunoassay. The hypothalamic content of GnRH, pituitary ACTH, and pituitary and plasma LH and FSH were measured in male, female and androgenized female rats from foetal Day 15 to postnatal Day 65. CR-activity was also measured at a few times during the postnatal period. Hypothalamic GnRH was detectable as early as foetal Day 15, increased gradually until postnatal Day 2 and then steeply between Days 5 and 16, a period during which major maturational changes occur in the hypothalamo- hypophysial axis. T|he levels of GnRH were similar in both males and normal females until Day 41, after which the increase which had been occurring from Day 16 continued in the male but not the female. In female animals treated with testosterone propionate (TP) shortly after birth, the hypothalamic content of GnRH at Day 9 was significantly less than that in normal females and males of the same age. Between Days 16 - 22 levels reached those found in the normal females and males. The lower level of GnRH in the TP treated female was associated with pituitary gonadotrophin and plasma FSH levels which were lower than in the normal female until Day.3O, suggesting that TP inhibits the synthesis and release of GnRH and gonadotrophins. In the normal female, vaginal opening was associated with a marked drop in pituitary LH and FSH and hypothalamic GnRH, but in the TP-treated female vaginal opening occurred while hypothalamic GnRH and pituitary LH levels were still rising. The peak in pituitary LH and the rise in plasma LH seen on Days 3O and 41, respectively, in the normal female were each delayed by about 1O days in the TP treated female. Similarly, the peak in pituitary FSH on Day 22 in the normal female occurred on Day 3O in the TP treated female. The profile of plasma FSH concentrations in the latter resembled more closely that of the male. In all three types of animal plasma FSH dropped significantly between Days 5O and 65. This could be attributed to inhibition of FSH release but not synthesis in the male (since pituitary FSH rose markedly during this period). However, in the androgenized female pituitary FSH also fell significantly suggesting that both release and synthesis had been inhibited. The Day 15 foetal pituitary also contained ACTH, and again levels of this hormone rose steeply between Days 5 and 9. In contrast to the gonadotrophins, there was a marked divergence between the pituitary content and concentration of ACTH, content rising while concentration remained relatively steady after Day 9. There was no major difference in pituitary ACTH levels between the three types of animal throughout the study; however, around Days 16 and 50 CR-activity was higher in the males and androgenized females compared with that in the normal females. Hypothalamic CR-activity and GnRH content, and pituitary and plasma LH, FSH and ACTH were measured in adult male rats maintained under a lighting regime in which the lights were on for 14 h. Two experiments were carried out. In the first some of the animals were kept in a room, distant from the animal house, in which the lighting was reversed with respect to the external environment. In animals exposed to the reversed lighting regime, hypothalamic GnRH content and pituitary gonadotrophin concentrations were significantly lower than the values in males kept in the animal house where they were in close proximity to female rats. In the second experiment, which was carried out on animals which had all been kept in the animal house, there were no significant differences between the GnRH contents measured at 3-4 hourly intervals throughout the day. Pituitary LH and FSH contents, but not concentrations, were significantly elevated at 12.OO h. There was little difference between the experiments in CR-activity, plasma ACTH concentrations and profiles of pituitary ACTH content and concentration. As expected there was a diurnal rhythm in plasma corticosterone concentrations; the peak occurred between 15.OO - 18.OO h. The profiles of^plasma and pituitary ACTH were similar to that of plasma corticosterone. CR-activity dropped significantly between 12.OO and 16.OO h, but remained stable at the other times. Hypothalamic GnRH and pituitary and plasma LH and FSH levels were also measured in female rats which had 4-day oestrous cycles. There were no significant differences between hypothalamic GnRH content. During pro-oestrus the mean GnRH content rose to reach a high level at 18.OO h at which time plasma LH concentration had risen sharply to a level consistent with the peak of the pre-ovulatory surge. Plasma FSH concentration also rose significantly between 15.OO and 18.00 h of pro-oestrus. At metoestrus and dioestrus plasma FSH levels were lower in the morning than in the evening. Extracts of cerebral cortex from animals of either sex, both prepubertal and adult, contained no detectable GnRH. However, cortical extracts did exhibit CR-activity This may have been due to the release of substances which cross reacted with the anti-ACTH serum but which were not steroidogenic (assessed in-vivo). These results suggest that (1) there is no diurnal rhythm in hypothalamic GnRH, (2) there may be a diurnal rhythm in pituitary gonadotrophin content in the male and in plasma FSH concentration on the days of metoestrus and dioestrus in the female, (3) if a surge of GnRH does occur on the afternoon of pro-oestrus, the rate of GnRH synthesis exceeds its release, and (4) the mechanism which regulates gonadotrophin secretion in the male may be affected by factors in the environment other than the length of daylight (e.g. presence of female animals). The data also support the view that the diurnal rhythm of corticosterone secretion is under hypothalamo-hypophys ia1 control. The effects of sex steroid hormones on the responsiveness of the neural mechanism responsible for the secretion of GnRH were examined in the female rat. Responsiveness was determined on pro-oestrus by measuring the increments in content (concentration x volume/time) of immunoreactive GnRH of pituitary stalk blood produced by electrical stimulation of the medial preoptic area or median eminence. Ovariectomy on the morning of dioestrus reduced the GnRH response to preoptic stimulation while oestradiol benzoate (OB) or TP administered immediately after ovariectomy augmented the response. The facilitatory effect of TP was possibly due to its conversion'to an aromatized derivative since 5a-dihydrotestosterone monobenzoate was ineffective. Progesterone did not facilitate preoptic responsiveness and, when administered to animals ovariectomized at 12.OO h of pro-oestrus, reduced the GnRH response at 18.OO h of the same day. Stimulation of the median eminence produced a significantly greater increment in GnRH than stimulation of the preoptic area.
Supervisor: Not available Sponsor: Not available
Qualification Name: Thesis (Ph.D.) Qualification Level: Doctoral
EThOS ID:  DOI: Not available