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K. YAMANOUCHI and Y. ARAI

Department of Anatomy, Juntendo University School of Medicine, Hongo, Tokyo 113, Japan

(Received 18 October 1977)

Differentiation of sexual behaviour patterns in male rats is dependent on the internal secretion of the testes during neonatal life. Removal of the testes at this time causes feminization and results in male rats which display female patterns of sexual behaviour (Gorski, 1971). Female patterns of behaviour are usually rare in normal male rats but recently we found that transection of the dorsal afferent neurones to the preoptic and anterior hypothalamic areas potentiated the display of lordosis in hormonally primed male rats (Yamanouchi & Arai, 1975). In the present study, further neuroanatomical analysis was carried out to clarify the localization of the afferent pathway involved in the regulation of lordosis behaviour.

Anterior or posterior roof deafferentation (ARD or PRD) was performed by lowering an L-shaped Halasz knife (2-5 mm horizontal blade) to the level

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K Ohshima, H Kishi, M Itoh, KY Arai, G Watanabe, K Arai, K Uehara, NP Groome and K Taya

The changes in plasma concentrations of inhibins A, B and pro-alpha C were determined in the cyclic golden hamster during follicular atresia induced with antiserum against luteinizing hormone releasing hormone (LHRH-AS) at 1100 h on day 4 (day 1=day of ovulation). Follicular status in the ovary was also studied by determining the number of follicles ovulating in response to human chorionic gonadotrophin (hCG) injection. The time-courses of changes in plasma concentrations of inhibins A, B and pro-alpha C were different from each other during induced follicular atresia and subsequent follicular development. Plasma concentrations of inhibin A decreased to 58.6% of initial values by 24 h after LHRH-AS treatment, and then remained relatively low until at least 60 h later. Plasma concentrations of inhibin B decreased to 64.2% of the initial values by 18 h after LHRH-AS treatment and remained at basal values for 36 h, but increased abruptly to greater than initial values at 42 h after the treatment. Plasma concentrations of inhibin pro-alpha C increased at 6 and 12 h, decreased suddenly to 21.9% of the initial values by 24 h after LHRH-AS treatment, and then gradually increased until 60 h after LHRH-AS. The number of follicles responding to hCG decreased gradually between 0 and 30 h after LHRH-AS, when no ovulations were observed, and then gradually increased until 60 h. The changes in follicular ovulatory responses to hCG correlated with the plasma profile of inhibin A throughout the experiment. These results suggest that inhibin A is mainly secreted by large antral follicles. In contrast, during the subsequent follicular development, the plasma concentration of inhibin B increased earlier than that of inhibin A. These results suggest that inhibin B is secreted by small and large antral follicles. Plasma concentrations of inhibin pro-alpha C were high at a time when plasma concentrations of oestradiol-17 beta had already decreased, indicating that inhibin pro-alpha C is secreted not only from healthy follicles but also from early atretic antral follicles.

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K Ohshima, H Kishi, M Itoh, G Watanabe, K Arai, K Uehara, NP Groome and K Taya

Plasma concentrations of inhibin pro-alphaC, inhibin A and inhibin B were determined by enzyme-linked immunosorbent assay at 6 h intervals throughout the 4-day oestrous cycle of the golden hamster. Plasma concentrations of follicle-stimulating hormone (FSH) and oestradiol-17beta were also measured by radioimmunoassay during the oestrous cycle. Plasma concentrations of inhibin A increased from the early morning of day 1 (day 1=day of ovulation) and reached plateau levels at 0500 h on day 2. An abrupt increase in plasma concentrations of inhibin A was found at 1700 h on day 4, when the preovulatory FSH surge was observed. An increase in plasma concentrations of inhibin B occurred on day 1 and reached plateau levels at 1700 h on day 1. The levels remained elevated until 0500 h on day 4 and declined gradually by 2300 h on day 4. Plasma concentrations of inhibin pro-alphaC gradually increased with some fluctuation from day 1 to 1700 h on day 4 and then declined. Significant negative relationships were noted between plasma FSH and both dimeric forms of inhibin from day 1 to day 3. Significant positive relationships were found between plasma oestradiol-17beta and inhibin A or inhibin pro-alphaC throughout the oestrous cycle. In contrast, no significant relationship was found between plasma oestradiol-17beta and inhibin B. These findings suggest that both dimeric forms of inhibin play a role in the regulation of FSH secretion during follicular development. These findings also suggest that inhibin pro-alphaC could be secreted primarily by large follicles, and early atretic follicles could also be responsible for inhibin pro-alphaC secretion. On the other hand, the secretory pattern of dimeric inhibins might shift from inhibin B to inhibin A with follicular development.

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K Ohshima, K Ohshima, KY Arai, H Kishi, M Itoh, G Watanabe, PF Terranova, K Arai, K Uehara, NP Groome and K Taya

Numerous antral follicles develop during the second half of pregnancy in the golden hamster. However, mechanisms regulating follicular development during this period are unknown. Because inhibin and activin are related to follicular development, these hormones were studied to gain insight into any potential roles in follicular development. Plasma inhibin A and B suddenly increased from day 8 of pregnancy, reached peak levels on day 10 and gradually declined to term. Plasma activin A gradually increased from day 8 to day 15 of pregnancy, and this was followed by an abrupt decrease at day one of lactation. Ovariectomy on day 12 of pregnancy rapidly reduced plasma inhibin A and B, but not activin A levels. Hysterectomy or placentectomy on day 12 of pregnancy caused an abrupt decrease in the levels of plasma activin A and FSH, but not inhibin A and B at 6 h after surgery. Hysterectomy also induced atresia of large antral follicles at 24 h after surgery. These results indicate that antral follicles are the main source of circulating inhibin A and B, whereas uteri and placentae are the main source of circulating activin A. These results suggest that increased levels of activin A may be involved in folliculogenesis in the ovary during the second half of pregnancy in the golden hamster.

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T Tsushima, M Arai, O Isozaki, Y Nozoe, K Shizume, H Murakami, N Emoto, M Miyakawa and H Demura

Abstract

Although endothelins were originally discovered as peptides with vasoconstrictor activity, recent studies have indicated a number of endothelin (ET)-induced hormonal functions in various tissues. We have studied the interaction of endothelins with porcine thyroid cells in culture. Specific binding of 125I-labelled ET-1 was demonstrated in porcine thyroid cells. The binding was displaced equally by unlabelled ET-1 and ET-2, but receptor affinity for ET-3 was lower than that for ET-1 and -2. Scatchard analysis of the data revealed a single class of high-affinity ET-1 receptors with a K d of 0·45 nmol/l and a binding capacity of 2100 sites/cell. SDS-PAGE and autoradiography of 125I-labelled ET-1 cross-linked with thyroid cell membranes demonstrated ET-1 binding sites with an apparent molecular weight of 50 kDa. These results indicated that ET-1 receptors in thyroid cells are type A ET receptors. In association with the presence of ET-1 receptors, porcine thyroid cells responded to ET-1 and ET-2 with an increase in c-fos mRNA expression. Although ET-1 did not affect DNA synthesis stimulated by either EGF or IGF-I, it dose-dependently inhibited TSH-induced iodide uptake and also inhibited iodide uptake stimulated by forskolin and 8-bromo-cAMP. ET-1 had no effect on TSH-stimulated cAMP production. Thus, ET-1 inhibited TSH-induced iodine metabolism by acting at the steps distal to cAMP production. In agreement with a recent report, immunoreactive ET-1 was detected in medium conditioned by porcine thyroid cells. Antibody to ET-1 was found to increase TSH-induced iodide uptake. These results are compatible with the notion that ET-1 negatively regulates TSH-induced iodide uptake in an autocrine manner.

Journal of Endocrinology (1994) 142, 463–470

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Koji Y Arai, Hisashi Kishi, Satoshi Onodera, Wanzhu Jin, Gen Watanabe, Akira K Suzuki, Shinji Takahashi, Toshihiko Kamada, Toshio Nishiyama and Kazuyoshi Taya

To elucidate changing patterns of inhibin/activin subunit mRNAs in the ovary of the golden hamster (Mesocricetus auratus) during the oestrous cycle, inhibin/activin subunit cDNAs of this species were cloned and ribonuclease protection assay and in situ hybridization were carried out. Inhibin α-subunit mRNA was localized in granulosa cells of primary, secondary, tertiary and atretic follicles throughout the 4-day oestrous cycle. It was also expressed in luteal cells on days 1 (oestrus), 2 (metoestrus) and 3 (dioestrus). βA-subunit mRNA was localized in granulosa cells of large secondary (>200 μm) and tertiary follicles throughout the oestrous cycle. βB-subunit mRNA was confined to granulosa cells of large secondary and tertiary follicles. Both α- and βA-subunit mRNAs were also found in ovarian interstitial cells and theca interna cells of tertiary and atretic follicles in the evening of day 4 (pro-oestrus). A striking increase in βA-subunit mRNA levels was also observed during the preovulatory period. The expression pattern of βA-subunit mRNA during the preovulatory period is unique and not found in other species. An i.v. injection of anti-luteinizing hormone-releasing hormone (LHRH) serum before the LH surge abolished the expression of α- and βA-subunit mRNAs in ovarian interstitial cells and theca interna cells. The treatment also abolished the preovulatory increase in βA-subunit mRNA. Furthermore, administration of human chorionic gonadotrophin (hCG), which followed the injection of anti-LHRH serum, restored the expression patterns of α- and βA-subunit mRNAs. The present study revealed that the golden hamster showed a unique expression pattern of βA-subunit mRNA in response to the LH surge.