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Summary.

Graafian follicles from mature New Zealand white rabbits were incubated for 4 hr at 37°C with air as the gas phase. The medium was changed every 15 min for 1 hr. Medium containing ovine LH was then added for four consecutive medium changes. Samples of the medium were analysed for 17 β-hydroxyandrogens, oestrogens and progestins by radioimmunoassay procedures. When no LH was present, progestin and 17 β-hydroxyandrogen secretion declined to negligible levels within 2 hr. Addition of 10 μg ovine LH to the medium for 1 hr caused an increase in 17 β-hydroxyandrogen secretion which continued for the duration of the experiment. No significant change was observed in oestrogen secretion but there was a slight increase in progestin secretion. Addition of 5 μg and 1 μg LH for 15 min also caused an elevation of 17 β-hydroxyandrogen secretion within 1 hr whereas progestin secretion took longer to increase and no change in oestrogen secretion was observed. No effect was observed with LH concentrations of less than 500 pg/ml for 1 hr, but 1 and 10 ng LH/ml also caused an increase in 17 β-hydroxyandrogen secretion. Oestrogen secretion was also stimulated at these levels. Control incubations with interstitial tissue showed no oestrogen secretion, slight secretion of 17 β-hydroxyandrogens and greater secretion of progestin which was stimulated by exogenous LH.

Different cell types of the ovarian follicle have been used to study steroid biosynthetic pathways. These investigations involved the use of tissue culture (Channing & Grieves, 1969) and incubations in vitro with specific buffers (Ryan & Short, 1965; Ryan, Petro & Kaiser, 1968). The activity of these cells in biological fluids has not been extensively studied. The aim of the present investigation was, therefore, to compare plasma and follicular fluid as incubating media for the biochemical study of the 3β-hydroxysteroid dehydrogenase and aromatizing activities of granulosa cells from equine ovarian follicles obtained at oestrus.

Radioactive steroids of high specific activity were purchased from the Radiochemical Centre, Amersham, Bucks., and were checked for purity before use. Reproductive tracts were obtained at slaughter from four mares judged to be in oestrus by gross examination of the uterus, cervix and corpus luteum using the criteria outlined by Channing (1969). At the

A number of previous investigations have dealt with steroid levels in follicular fluid of the mare (Short, 1961; van Rensburg & van Niekerk, 1968; YoungLai, 1971), the cow (Short, 1962; Lunaas, 1964) and human (Abraham, Odell, Edwards & Purdy, 1970). With the development of highly sensitive methods for the detection of steroids in small amounts of biological material, it was of interest to determine whether steroids could be detected in follicular fluid of a reflex ovulator such as the rabbit. It was felt that by adopting this approach more direct evidence would be obtained for the synthesis and release of oestrogen by the ovarian follicle of this species.

Sexually mature, virgin, New Zealand white rabbits (3·5 to 5·5 kg) were used. Does were mated to a fertile buck. Anaesthesia was induced with

Summary.

Graafian follicles from New Zealand white rabbits were incubated at 37°C for various periods of time with air as the gas phase. Media were changed every 15 min and stored at -15°C until analysed for progestins, 17β-hydroxyandrogens and oestrogens using established radioimmunoassay procedures. At fixed times after the start of the incubations, media containing various test substances were added with subsequent replacement by medium alone. Addition of 5 μg LH/ml for 1 sec caused a dramatic increase in the synthesis and secretion of androgen with lesser increases in progestin and oestrogen. Puromycin and cycloheximide but not actinomycin D, inhibited LH-induced steroidogenesis. Cyclic AMP, dibutyryl cyclic AMP, cyclic CMP, 5'-AMP, and theophylline also caused an increase in androgen production which rapidly ceased when media without nucleotides were added. Sodium fluoride had no effect on steroidogenesis. From these data it was concluded that (i) the rabbit follicle is the major source of ovarian androgen; (ii) the binding of LH to the follicular cells is a rapid process; (iii) the events following LH binding do not require the presence of LH in the medium; (iv) cyclic nucleotides which may act as second messengers also stimulate steroidogenesis; (v) the effects of LH and cyclic nucleotides on steroidogenesis are different; and (vi) the action of LH on follicular steroidogenesis probably occurs at the translational level.

Summary.

Seasonal, diurnal and episodic patterns of LH and testssterone secretion in sexually mature male New Zealand rabbits were studied. Blood samples were obtained from the central ear artery by puncture or through an indwelling catheter, and were assayed for hormones using radioimmunoassay. Testosterone values appeared to be lower in the summer months while LH showed no seasonal cyclicity. There were no significant fluctuations when samples were taken at 10min intervals, but specimens taken every hour for 24 or 36 hr revealed an episodic pattern of release. Peaks of both hormones occurred every 4 to 5 hr in most animals. Testosterone levels ranged from 0·5 to 10 ng/ml and LH from 15 to 200 ng/ml of WP360A standard. In general, a rise in LH preceded or coincided with an increase in testosterone. No specific diurnal rhythm could be demonstrated and the patterns appeared to be unrelated to external stimuli.

Summary. Monthly collections of hibernating little brown bats contained (1) nulliparous females with small uteri and no antral follicles, (2) nulliparous females with swollen uteri and mature follicles, and (3) parous females, which, despite obvious differences in reproductive status, had equivalent plasma progesterone values. During the principal study season, mean monthly progesterone concentrations (measured by radioimmunoassay) showed recurrent increases with an apparent periodicity of about 60 days, but limited data obtained in the subsequent season did not. However, comparison of activity patterns in the two seasons with monthly progesterone concentrations suggests that ovarian activity during hibernation is affected by variations in metabolic level.

We saw no evidence that nulliparous bats with small uteri developed antral follicles during hibernation. Despite their apparent immaturity, however, they had cornified vaginae and most were demonstrably inseminated. These indications of oestrus and the lack of differences between their plasma progesterone concentrations and those of patently mature females suggest that they were physiologically post-pubertal but failed to complete folliculogenesis before entering hibernation.

Keywords: progesterone; bat; hibernation

Summary. Metabolism of [U-¹⁴C]glucose was studied in the prenatal and neonatal rabbit ovary. Control tissues included the testis and female liver. No significant changes in glucose metabolism were observed in liver tissue. Mitosis and glucose oxidation were maximal in ovary and testis at 30 days post coitum and then declined dramatically by Day 8 after birth. Since mitosis is the primary physiological event in the gonad during the perinatal period these data suggest that glucose may be an important carbohydrate source for energy at this time.

Summary. Developing female rabbits were studied weekly from Day 22 of life to Day 100. At all ages GnRH 1·5 μg/kg) induced a large increase in LH release 15 min later. By contrast, FSH was significantly increased only on Days 22, 29 and 72 and no significant increase was detected up to 2 h after GnRH administration at other ages. Functional corpora lutea were absent at the start of all treatments as indicated by circulating concentrations of progesterone < 2 ng/ml. It is concluded that the immature rabbit pituitary is functionally capable of responding to GnRH with an increase in LH secretion, whereas the control of FSH secretion may be regulated by other factors.

Keywords: LH; FSH; GnRH; age-dependent changes

Summary.

The involvement of central catecholaminergic neurons in tonic gonadotrophin release was investigated. Male rats were given a single cerebral intraventricular injection of 6-hydroxydopamine hydrochloride (6-OHDA; 170 μg free base) dissolved in 0·001 n-HCL. Plasma LH and FSH were measured by double antibody radioimmunoassay. In the 6-OHDA-treated animals, LH was significantly reduced (P<0·001 after 1 hr and remained consistently lower for 8 hr. After a transient elevation, the controls showed no significant change (P>0·01). No difference in LH concentrations was found between experimental groups sampled at 2 days or later (P>0·01). Levels of FSH were less consistently different. Administration of 6-OHDA dissolved in different vehicles, with varied pH and osmolarity, gave similar results which suggests that the quinone derivative of 6-OHDA is effective in producing the catecholaminergic impairment. These findings indicate that LH release in the male rat may be controlled (or at least modulated) by a central adrenergic mechanism.

Summary.

The testosterone concentration in allantoic fluid between 90 and 150 days of gestation in cattle can be used to determine the fetal sex; values were 442±20·3 (S.E.M.) pg testosterone/ml for male fetuses and 215±8·2 pg/ml for female fetuses.