Biomedical subjects

M Tetsuka

Publications and source records attributed to M Tetsuka.

14 recordsLinked to original sources

PubMed1999

GABA(B) receptors in the nucleus tractus solitarii modulate the carotid chemoreceptor reflex in rats.

In urethane-chloralose anesthetized rats, the role of GABA(B) receptor in the commissural subnucleus of the nucleus tractus solitarii (commNTS) on the carotid chemoreceptor reflex was investigated. Microinjection of a GABA(B) agonist baclofen into the commNTS did not have any effects on arterial blood pressure (BP) or respiration (RP), while it attenuated the increases in BP and RP elicited by carotid chemoreceptor stimulation. These effects were blocked by microinjection of a GABA(B) antagonist 2-OH-saclofen into the same site. Prior microinjection of 2-OH-saclofen did not have any effects on the chemoreflex or on resting BP or RP, while the effects of baclofen on the chemoreflex were completely blocked. These results suggest that GABAB receptors are present in the carotid chemoreceptor reflex pathway in commNTS and modulate the chemoreceptor reflex.

Animals↗

PubMed1999

Expression of 11beta-hydroxysteroid dehydrogenase, glucocorticoid receptor, and mineralocorticoid receptor genes in rat ovary.

A new concept in reproductive endocrinology is that the status of the ovary as a glucocorticoid target organ alters with follicular development. Evidence for a physiological role of glucocorticoids in the regulation of ovarian folliculogenesis has been strengthened by the discovery that 11beta-hydroxysteroid dehydrogenase (11betaHSD) mRNA expression in human granulosa cells is developmentally regulated. In this study, we quantified the pattern of expression and investigated the cellular location of 11betaHSD type 1 (11betaHSD1), 11betaHSD type 2 (11betaHSD2), glucocorticoid receptor (GR), and mineralocorticoid receptor (MR) mRNAs during follicular maturation in rat ovary. Immature female rats received treatment with eCG to induce preovulatory follicular development or eCG followed by hCG to induce luteinization. 11betaHSD1, 11betaHSD2, GR, and MR mRNAs were all detectable by ribonuclease protection assay in ovarian total RNA. Treatment with eCG alone caused an approximately 8-fold increase in the ovarian level of 11betaHSD1 mRNA, which rose to approximately 30-fold after additional treatment with hCG. Equine CG alone did not measurably affect the ovarian 11betaHSD2 mRNA level, but additional treatment with hCG reduced it to 34% of the control level. Expression of GR mRNA was unchanged by any gonadotropin treatment, while MR mRNA was down-regulated. A similar pattern of 11betaHSD1, 11betaHSD2, GR, and MR mRNA expression was observed in isolated granulosa cells. These results provide direct experimental evidence that 11betaHSD genes are gonadotropically regulated in the rat ovary, including granulosa cells, and are consistent with a shift in glucocorticoid metabolism from inactivation (due to oxidation by 11betaHSD2) to activation (reduction by 11betaHSD1) during hCG-induced granulosa cell luteinization.

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed1998

Expression of oestrogen receptor isoforms in relation to enzymes of oestrogen synthesis in rat ovary.

Oestrogen acts as a local regulator of follicular development, mediated by specific nuclear oestrogen receptors (ER). The aim of this study was to examine the gene expression of two ER isoforms, ERalpha and beta, in relation to oestrogenic enzymes, P450aromatase (P450arom) and 17betaHSD type1 (17betaHSD1) during follicular maturation and luteinization. Ovaries were obtained from immature rats treated with PMSG (10 iu for 48 h) followed by hCG (10 iu). Expression of ERalpha and beta was down-regulated by the treatment with PMSG. Following the hCG injection, further down-regulation of both ERs occurred. Conversely, expression of P450arom and 17betaHSD1 was initially up-regulated by PMSG and then rapidly down-regulated following injection of hCG. In isolated granulosa cell, ERbeta was the predominant ER while ERalpha was mainly expressed in residual ovary and corpora lutea (CL). These results indicate that ERalpha and ERbeta mRNAs are expressed in different cell types and are both down-regulated during gonadotrophin stimulated follicular maturation and luteinization in the rat ovary. Down-regulation of ER is accompanied by down-regulation of oestrogenic enzymes during luteinization. Thus both oestrogen production and reception are shut down as ovulation approaches.

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed1998

An anti-inflammatory role for glucocorticoids in the ovaries?

Roles of glucocorticoids in the direct regulation of ovarian function are poorly understood. This opinion paper highlights: (1) the inflammatory nature of the ovulatory process; (2) the contributions of cytokines and prostaglandins (hence inflammation) to ovulation; (3) the development-related pattern of 11beta-hydroxysteroid (11beta HSD) isoform expression (hence glucocorticoid metabolism) that occurs in ovarian follicles; and (4) the attribution of general anti-inflammatory properties of glucocorticoids to their interference with prostaglandin synthesis. We interpret the evidence cited to hypothesise that corticosteroids serve an anti-inflammatory role during ovulation, thereby promoting rapid healing of the wound left by follicular rupture. Other possible levels of glucocorticoid action in the ovaries are also considered.

Animals↗

PubMed1998

Corticosteroid metabolism in human granulosa-lutein cells.

OBJECTIVE: The aims of this study were to determine the type and level of 11 beta-hydroxysteroid dehydrogenase (11 beta HSD) in human granulosa-leutein cells (GLE) shortly before ovulation and to correlate activity with the outcome of treatment in patients undergoing in vitro fertilization and embryo transfer (IVF/ET). DESIGN: GLC from 32 patients undergoing IVF/ET were tested for type and level of 11 beta HSD activity in relation to treatment outcome. PATIENTS: Periovulatory follicles were aspirated by ultrasound guided transvaginal puncture following a standard controlled ovarian stimulation protocol, approximately 36 h after administering an ovulation-inducing dose of human chorionic gonadotrophin (HCG). GLC were separated from follicular fluid by density-gradient centrifugation and taken for measurement of 11 beta HSD activity in vitro; oocytes were used for IVF/ET. MEASUREMENTS: Interconversion of cortisol (F) and cortisone (E), and dexamethasone (D) and 11-dehydrodexamethasone (DHD) was measured in standardized assays comprising incubation of GLC with 3H-labelled substrate, with separation of substrate and product by thin-layer radiochromatography. RESULTS: Conversion of F to E varied from 10.5 to 30.9% while that of E to F was between 2.4 and 44.6%. In the GLC of 25 patients in whom both activities were measured, dehydrogenase (F to E) activity predominated in 13 and reductase (E to F) in 12. By contrast, D (substrate for 11 beta HSD2 but not 11 beta HSD1) showed less than 1% metabolism in this system while DHD (substrate for 11 beta HSD1 and 11 beta HSD2) was converted significantly (65.6-90.5%) to D in the four patients tested. There was no significant difference in the interconversion of F and E between patients who became pregnant and those who did not. CONCLUSIONS: The dehydrogenase and oxoreductase reactions catalysed by 11 beta HSD both occur in granulosa-lutein cells at the time of follicular rupture, probably due to 11 beta HSD1. A lack of measurable conversion of dexamethasone to 11-dehydrodexamethasone suggests that dehydrogenation due to 11 beta HSD2 is low or absent. Neither type nor level of 11 beta HSD activity measured under the present assay conditions correlates with IVF outcome.

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed1998

Expression of oestrogen receptor alpha and beta in cultured human ovarian surface epithelial cells.

Ovarian surface epithelial (OSE) cells participate in the formation of the ovarian cortex and are potential targets of oestrogen action. Oestrogens typically act through nuclear oestrogen receptors (ER) of which there are two known subtypes: ERalpha and ERbeta. In view of the potential importance of oestrogen as a local regulator of OSE cell function, we screened for ERalpha and ERbeta mRNA in primary OSE cell cultures by reverse transcriptase-polymerase chain reaction (RT-PCR) analysis, and used freshly isolated granulosa cells (GC) and granulosa-lutein cells (GLC) as positive controls. OSE cells, scraped from the ovarian surface of women undergoing laparotomy for benign gynaecological conditions, were cultured for up to 21 days to obtain enough cells for mRNA extraction. GC were obtained from spontaneously cyclic women undergoing total hysterectomy; while GLC were obtained from follicular aspirates of gonadotrophin-stimulated in-vitro fertilization patients. Total RNA (1 microg) was reverse transcribed into single-stranded cDNA for PCR (30 cycles) using primers selected to give specific ERalpha and ERbeta products. The ERalpha and ERbeta PCR products, authenticated by cloning and sequencing, were both weakly detectable by Southern analysis in cultured OSE cells and readily detectable in GC and GLC. These results show that cultured human OSE express both ERalpha and ERbeta mRNA, consistent with a role for oestrogen in the regulation of OSE cell function in vivo.

Adult↗

PubMed1997

Decrease in cerebral free magnesium concentration following closed head injury and effects of VA-045 in rats.

1. We examined the alterations in cerebral free Mg2+ concentration in closed head injury (CHI) in rats and the effects of VA-045, a novel apovincaminic acid derivative, on them with in vivo 31P-NMR. 2. Free Mg2+ decreased by about 30% within 20 min after head impact and, afterward, it gradually decreased further to reach about 60% of the control level after 3 hr. VA-045 inhibited the decrease. 3. In nonimpacted rats, VA-045 did not alter the free Mg2+ level. 4. The decrease in cerebral free Mg2+ following CHI may be a critical factor in the development of irreversible tissue injury, and VA-045 may prevent it.

Animals↗

PubMed1997

Role of androgens in follicle maturation and atresia.

Androgens are products of progestogen metabolism, intermediates in oestrogen biosynthesis and local regulators of ovarian function. Current understanding of intraovarian androgen formation, metabolism and action is reviewed, highlighting the contribution of androgens to the paracrine regulation of follicular maturation and atresia. Any factor that alters intracellular cAMP levels is a potential modulator of granulosa cell differentiation, and hence follicular development. Androgen appears to modulate gonadotrophin action on granulosa cells through amplification of cAMP-mediated post-receptor signalling. Here it is argued that during intermediate stages of follicular development, locally produced androgen acts via granulosa cell androgen receptors (AR) to promote follicle-stimulating hormone (FSH)-induced granulosa cell differentiation through amplifying cAMP-mediated post-receptor signalling. During late pre-ovulatory follicular development, higher concentrations of cAMP caused by stimulation with luteinizing hormone (LH) suppress granulosa cell proliferation and down-regulate some of the genes induced by FSH at earlier stages of pre-ovulatory development, including aromatase activity. Other granulosa cell functions, including progesterone synthesis, are enhanced by the high concentrations of cAMP induced by LH. There is experimental evidence from studies of rat and non-human primate (common marmoset) ovaries that AR levels in granulosa cells decline during pre-ovulatory follicular maturation. Since androgens augment FSH-induced cAMP formation and action, loss of AR could be a means of avoiding inappropriately high cAMP levels and hence avoiding premature activation of 'high-tone' cAMP-response genes that lead to atresia. Negative regulation of the granulosa cell AR could be part of the intra-ovarian mechanism that determines which follicle(s) becomes dominant and secretes oestrogen in the normal menstrual cycle.

Androgens↗

PubMed1997

Location and developmental regulation of androgen receptor in primate ovary.

Locally produced androgens act via granulosa cell androgen receptors to modulate follicular responsiveness to gonadotrophins and thereby contribute to the paracrine regulation of ovarian function. We used quantitative androgen receptor immunocytochemistry to assess androgen receptor distribution in relation to pre-ovulatory follicular development in the common marmoset (Callithrix jacchus), a New World primate that ovulates two to four follicles in each approximately 28 day ovarian cycle. Ovaries from four adult females in the late follicular phase and from four in the luteal phase were fixed in 4% paraformaldehyde and subjected to an immunocytochemical analysis using a polyclonal androgen receptor antibody with detection by a standard avidin-biotin-peroxidase technique for alkaline phosphatase. Specific androgen receptor immunostaining occurred mainly in granulosa cell nuclei, with little or no specific staining in theca, stroma or oocytes. Granulosa cell androgen receptor immunostaining was most abundant in healthy preantral/early antral follicles, being low or absent from pre-ovulatory follicles and corpora lutea. Differences in granulosa cell androgen receptor immunostaining between immature (0.1-1.0 mm diameter) and pre-ovulatory (> or = 2.0 mm diameter) follicles were quantified using a videodensitometric analysis of grey-scale values. Readings were taken from the granulosa cell layers of 53 immature follicles and 10 pre-ovulatory follicles in late follicular phase ovaries. The average androgen receptor level in granulosa cells of immature follicles proved to be 4.2-fold higher (P < 0.01) than that in granulosa cells of pre-ovulatory follicles. Because other evidence suggests that paracrine androgen action in granulosa cells converts from stimulation to inhibition as follicles mature, we speculate that a development-related reduction in androgen receptor numbers serves to "protect' granulosa cells against the inhibitory action of androgen, thereby promoting pre-ovulatory follicular dominance in primate ovarian cycles.

Animals↗

PubMed1997

Differential expression of messenger ribonucleic acids encoding 11beta-hydroxysteroid dehydrogenase types 1 and 2 in human granulosa cells.

In glucocorticoid target organs 11beta-hydroxysteroid dehydrogenase (11betaHSD) regulates the levels of active glucocorticoids available to glucocorticoid receptors. To date two isoforms of 11betaHSD, NADP-dependent type 1 11betaHSD (11betaHSD1) with predominant reductase activity and NAD-dependent type 2 11betaHSD (11betaHSD2) with dehydrogenase activity have been identified. Human ovarian granulosa cells have been shown to possess both dehydrogenase and reductase 11betaHSD activities and express 11betaHSD1 mRNA. However, whether 11betaHSD2 mRNA is also present or if the expression of either mRNA is developmentally regulated in the human ovary is unknown. We therefore used northern analysis to examine 11betaHSD1 and 11betaHSD2 mRNA levels in non-luteinized and luteinizing granulosa cells, corpora lutea (CL) and ovarian stroma obtained from human ovaries. Here we show that non-luteinized granulosa cells express relatively high levels of 11betaHSD2 mRNA but not 11betaHSD1. Conversely, luteinizing granulosa cells abundantly express 11betaHSD1 mRNA but not 11betaHSD2. CL also expresses 11betaHSD2 to lesser extent. Neither 11betaHSD mRNA is detectable in ovarian stroma. These results indicate that mRNAs encoding both 1lbetaHSD isozymes are present in human granulosa cells and they are developmentally--but differentially--regulated during preovulatory follicular development. The existence of developmentally regulated 11betaHSD in human granulosa cells is important new evidence that glucocorticoids, acting directly on the ovary, serve physiologically significant roles in the regulation of folliculogenesis.

11-beta-Hydroxysteroid Dehydrogenases↗

PubMed1997

Differential regulation of aromatase and androgen receptor in granulosa cells.

During follicular development, androgen acts in three distinct ways. During the early stage of follicular differentiation, androgen acts as an enhancer of FSH-stimulated follicular differentiation. As follicular differentiation progresses, this effect is decreased and androgen is mainly utilized as a substrate for estrogen synthesis under increasing stimulation of FSH and LH. These two events are mediated by androgen receptor (AR) and aromatase (P450arom), respectively. In the rat and marmoset monkey, AR and P450arom are predominantly expressed in granulosa cells, and both are developmentally regulated. The expression of AR is highest in preantral/early antral follicles and gradually decreases as follicles mature, whereas expression of P450arom is increased as follicular differentiation progresses. We propose that differential regulation of these two androgen-utilizing factors contributes to the smooth transition of developing follicles from the early stage of differentiation to the fully mature ovulatory status. A failure of this transition due to improper androgen stimulation might result in follicular atresia.

Androgens↗

PubMed1996

Androgen receptor gene expression in rat granulosa cells: the role of follicle-stimulating hormone and steroid hormones.

In rat ovary, androgen receptor (AR) is predominantly expressed in granulosa cells and is developmentally regulated. However, the exact mechanism that is responsible for the regulation of AR in granulosa cells has not been elucidated. The aim of this study was to examine 1) the levels of AR messenger RNA (mRNA) expression in granulosa cells from follicles of different size and 2) the effects of FSH, 8-bromo-cAMP, androgen, and estrogen on AR mRNA levels in granulosa cells in vitro. The abundance of AR mRNA was examined by ribonuclease protection assay using 32P-labeled AR complementary RNA probe and related to that of P450aromatase (P450arom) mRNA, a well established maker of granulosa cell differentiation. In large follicles (> 400 microns in diameter), the abundance of AR mRNA was decreased to 51% of that in small follicles (< 200 microns; P < 0.01), whereas the abundance of P450arom mRNA increased to 277% (P < 0.01). In medium follicles (200-400 microns), the abundance of AR mRNA was maintained (101%), whereas the abundance of P450arom mRNA increased to 202% of that in small follicles (P < 0.05). Treatment with FSH (0-300 ng/ml) or 8-bromo-cAMP (0-4 mM) induced P450arom mRNA in the cultured granulosa cells in a dose-dependent manner; however, it did not affect the levels of AR mRNA expression. Treatment with 5 alpha-dihydrotestosterone (1 microM) resulted in a significant reduction in the abundance of AR mRNA to 67% of the control value (P < 0.05). This effect was reversed by the addition of FSH (100 ng/ml; P < 0.01). Treatment with diethylstilbestrol (1 microM), alone or in combination with FSH (100 ng/ml), did not have any significant effect, although these treatments tended to decrease the abundance of AR mRNA to 81% and 85%, respectively. Both 5 alpha-dihydrotestosterone and diethylstilbestrol dramatically enhanced the abundance of FSH-induced P450arom mRNA compared to the effect of FSH alone. These results indicate that 1) the down-regulation of AR mRNA expression takes place in granulosa cells of preovulatory follicles; 2) FSH is not directly responsible for this event; and 3) androgen down-regulates AR mRNA expression in immature granulosa cells, and this effect is reversed by FSH. We conclude that androgen and FSH jointly regulate AR mRNA expression in rat granulosa cells.

8-Bromo Cyclic Adenosine Monophosphate↗

PubMed1995

Developmental regulation of androgen receptor in rat ovary.

Androgen receptor (AR) distribution and developmental regulation in the rat ovary were examined by semiquantitative immunohistochemistry. Ovarian AR mRNA levels were also determined by Northern analysis of total RNA and compared with the levels of cytochrome P450aromatase (P450arom), an established marker of preovulatory follicular maturity. Hypophysectomized immature female rats were treated with recombinant human (rh)-FSH and/or rh-LH, or human menopausal gonadotrophin (HMG). AR was predominantly located in granulosa cells. There was no indication of specific AR immunoreactivity in thecal cells, but scattered stromal cells did stain positively. In control and LH-treated ovaries, only small preantral/early antral follicles were present. Granulosa cells in these follicles showed intense AR immunostaining. Treatment with FSH, FSH and LH or HMG stimulated varying degrees of preovulatory follicular development. In these follicles, the intensity of AR immunostaining progressively declined as follicular development progressed. In intact immature rats treated with FSH, the abundance of ovarian AR mRNA was significantly decreased to 35% of the control value while combined treatment of FSH and LH resulted in further down-regulation of AR mRNA expression to 17% of the control value. A decrease in the abundance of AR mRNA was accompanied by a simultaneous increase in the abundance of P450arom mRNA. Similar results were obtained in hypophysectomized immature rats treated with FSH and LH, suggesting an inverse relationship between AR mRNA expression and granulosa cell maturity. These results suggest that (1) the AR is most abundant in the granulosa cells of rat ovaries and (2) the expression of AR and its mRNA are developmentally regulated, being down-regulated during FSH-stimulated preovulatory follicular development.

Animals↗

PubMed1989

Seven-legged calf--dipygus with an extra foreleg at the pelvic region.

A male Holstein-Friesian calf with seven legs was examined macroscopically and radiographically. External features included two normal forelimbs, two normal hindlimbs (lateral hindlimbs), and two abnormal hindlimbs (medial hindlimbs) which were underdeveloped. Also, a rudimentary forelimb, which was attached to the pelvic region, was observed between both the medial hindlimbs. It consisted of an underdeveloped humerus, a duplicated ulna, several carpal bones, a partially duplicated metacarpal bone and three digits with three hoofs. This leg was connected with two sets of coxae by a irregular-shaped bone considered the vestigial vertebrae and ribs. Two penises and scrotums, three kidneys and testes were also observed. This calf is the first case of dipygus associated with pygopagus parasiticus in cattle. Based on these findings, the pathogenesis of this rare anomaly was briefly discussed from an embryological point of view.

Abnormalities, Multiple↗