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The synthesis and release of prolactin in rats were significantly higher at oestrus than at any other stage of the cycle, when measurements were made between 10.00 and 11.00 h (Ieiri, Akikusa & Yamamoto, 1971). Since considerable evidence indicates that stimulation of prolactin release begins on the afternoon of the day of pro-oestrus (Ieiri et al. 1971), synthesis and release were measured in the afternoon of the day of each stage of the oestrous cycle to determine the fluctuations of these parameters in the course of the cycle.

Adult female rats of the Wistar strain showing regular 4-day cycles were used. Animals were killed by decapitation between 16.00 and 17.00 h. Each isolated anterior pituitary was incubated with [¹⁴C]leucine. All the experimental conditions were the same as described previously (Ieiri et al. 1971).

The anterior pituitary functions were estimated by counting the radioactivity of [¹⁴C]leucine incorporated into prolactin

ABSTRACT

This study was carried out (1) to compare the time-course of the change in bone metabolism in rats administered gonadotrophin-releasing hormone agonist (GnRHa) and bilaterally ovariectomized (OVX) rats and (2) to investigate the changes in bone metabolism after discontinuance of GnRHa.

Seventy female Sprague–Dawley rats, aged 90 days, were divided into four groups. Group 1 underwent a sham operation, group 2 was surgically ovariectomized and group 3 was given a GnRHa (leuprorelin acetate for depot suspension) s.c. injection every 30 days. Group 3 was further divided into three subgroups: rats were administered GnRHa for 12 months (GnRHa 12M), 6 months (GnRHa 6M) or 3 months (GnRHa 3M). Group 4 served as a basal control. The bone mineral density (BMD) of lumbar vertebrae and femoral bone, measured by dual-energy X-ray absorptiometry, and the serum bone metabolic parameters were determined every 45–90 days. The bone histomorphometry of lumbar vertebra was measured on days 180 and 360 after surgery.

GnRHa 12M rats showed significantly lower BMD of vertebrae and femoral bone, lower bone volume and higher bone turnover compared with sham-operated rats and those with secondary hyperparathyroidism on days 180 and 360. Their time-course for changes in bone metabolism was almost the same as that of OVX rats. GnRHa-discontinued rats showed a recovery of bone turnover. The recovery of BMD in GnRHa 6M rats was smaller than that of GnRHa 3M rats after GnRHa discontinuance. The bone volume for GnRHa 6M rats was significantly lower than that for GnRHa 3M on day 360.

Journal of Endocrinology (1993) 138, 115–125

ABSTRACT

While prostaglandin F_2α (PGF_2α) has been thought to be a natural luteolysin in non-primates, a luteolytic effect in the human corpus luteum is less evident. We therefore investigated the action of PGF_2α on monolayer cultures of human luteal cells obtained from mid-luteal phase corpora lutea.

PGF_2α increased basal and human chorionic gonadotrophin (hCG)-stimulated progesterone production by human cultured luteal cells. A potent tumour-promoting phorbol ester, phorbol 12-myristate-13-acetate (PMA), also stimulated progesterone production by cultured human luteal cells.

Although human luteal cells were incubated for 24 h with PMA, hCG was still able to stimulate the production of progesterone by PMA-pretreated cells. However, PMA pretreatment blocked the ability of PGF_2α to stimulate progesterone production. It is possible that the luteotrophic effect of PGF_2α may be mediated, in part, by the activation of protein kinase C.

Addition of PGF_2α to suspensions of human luteal cells preincubated with myo-[2-³H]inositol promoted an increase in labelled inositol phosphates. PGF_2α also rapidly increased intracellular free Ca²⁺ in human luteal cells loaded with the fluorescent Ca²⁺ probe, fura-2.

We conclude that PGF_2α and PMA stimulate progesterone production and that PGF_2α increases the intracellular free calcium and inositol phosphates of human cultured luteal cells in the mid-luteal phase.

Journal of Endocrinology (1992) 133, 451–458

ABSTRACT

Immunohistochemical localization of 17α-hydroxylase/C17-20 lyase (P-450_17α,lyase) and aromatase cytochrome P-450 (P-450_arom) in polycystic ovary (PCO) syndrome was studied using specific polyclonal antibodies which had been raised against the corresponding enzymes. In the majority of follicles that were atretic and smaller than 7 mm in diameter, theca interna cells showed high P-450_17α,lyase immunoreaction, while small numbers of granulosa cells showed little P-450_arom immunoreaction. In some atretic follicles that were larger than 11 mm in diameter, the hyperplastic theca interna cell layer showed high immunoreaction to P-450_17α,lyase, while the poorly proliferated granulosa cell layer showed a mixture of weak and negative immunoreaction to P-450_arom. No immunoreaction to P-450_17α,lyase or P-450_arom was recognized in PCO stroma. These findings suggest that the theca interna cells and the granulosa cells from PCOs show abnormal steroidogenic function, while the localization of P-450_17α,lyase and P-450_arom in PCOs was essentially identical to that in the normal ovary. Theca interna cells in PCO atretic follicles are the main site of excess androgen production.

Journal of Endocrinology (1993) 139, 503–509

ABSTRACT

Immunohistochemical localization of 17α-hydroxylase/C17–20 lyase (P-450_17α,lyase) and aromatase cytochrome P-450 (P-450_arom) in normal human ovaries during the menstrual cycle was studied using specific polyclonal antibodies which were raised against corresponding enzymes. In the follicular phase of matured follicles, P-450_17α,lyase was localized in theca interna cells and P-450_arom in granulosa cells. P-450_17α,lyase was expressed in theca interna cells before P-450_arom was expressed in granulosa cells. The corpus luteum showed immunoreactivity to both enzymes and, after menstruation, immunoreactivity decreased gradually until it could not be detected in the corpus albicans. In corpus luteum graviditatis the immunoreactivity continued to be expressed strongly. In some atretic follicles, P-450_17α,lyase and/or P-450_arom continued to be expressed. In the stromal layer, P-450_17α,lyase was detected in secondary interstitial cells, which originated from the theca interna of atretic follicles, and P-450_arom was detected in hilar cells. Immunoreactivity to both enzymes was also detected in oocytes of developing follicles. These results are consistent with the two cell theory in the human ovary. They also suggest that androgens and oestrogens are produced not only by follicles and corpora lutea but also by stroma and oocytes.

Journal of Endocrinology (1992) 135, 589–595

Abstract

Arachidonate 12-lipoxygenase, which oxygenates positions 12 and 13 of arachidonic and linoleic acids, is present in porcine anterior pituitary cells. Colocalization of the 12-lipoxygenase with various pituitary hormones was examined by immunohistochemical double-staining using antibodies against 12-lipoxygenase and various anterior pituitary hormones. Under light microscopy, approximately 7% of the cells producing luteinizing hormone (LH) and follicle-stimulating hormone (FSH) were positive for 12-lipoxygenase, whereas the enzyme was detected in less than 2% of the cells producing thyrotrophin, prolactin, growth hormone (GH), and adrenocorticotrophin. In an attempt to examine the participation of 12-lipoxygenase metabolites in pituitary hormone release, we incubated the primary culture of porcine anterior pituitary cells with 12-hydroperoxy-arachidonic acid or 13-hydroperoxy-linoleic acid. Significant stimulation of LH and FSH release by these hydroperoxides was observed at 10 μm in a time-dependent manner. At doses around 10 μm these compounds produced responses of similar magnitude to 1 nm gonadotrophin-releasing hormone (GnRH), but higher concentrations (30 μm) of the compounds were required for GH release. In contrast, 12-hydroxy-arachidonic and 13-hydroxy-linoleic acids were almost ineffective. Furthermore, the gonadotrophin release by 1 nm GnRH was inhibited by nordihydroguaiaretic acid (a lipoxygenase inhibitor) with an IC₅₀ of about 5 μm. Thus, the hydroperoxy (but not hydroxy) products of 12-lipoxygenase may be involved in the release of pituitary hormones especially LH and FSH.

Journal of Endocrinology (1996) 148, 33–41