Subunit Messenger Ribonucleic Acids Research Articles

Differential Expression of GABA Receptor Subunit Messenger Ribonucleic Acids and Immunoreactivity in the Rat Neostriatum during Postnatal Development

γ-Aminobutyric acid (GABA) is a major inhibitory neurotransmitter in the neostriatum. Functions of GABA are mediated by GABA_A and GABA_B receptors in the neostriatum. In order to investigate the developmental expression of GABA receptor subunits (GABA_Aα₁, GABA_Aα₃ and GABA_Aα₆; GABA_Aβ₂ and GABA_Aβ₃, and GABA_BR₁ and GABA_BR₂) in the rat neostriatum, reverse transcriptase-polymerase chain reaction (RT-PCR) and immunofluorescence were performed. Tissues were obtained from rats on postnatal day (PND) 1, PND 7 and PND 14, and from adult rats. RT-PCR indicated that GABA_Aα₁ and GABA_Aα₆ mRNA levels were low, but that GABA_Aβ₂ and GABA_Aβ₃ mRNAs levels were high during the early postnatal period. Immunofluorescence revealed that GABA_Aα₁ immunoreactivity was only observed in striatal interneurons in PND 14 and in adult rats. Immunoreactivity for GABA_Aα₆ was only observed in PND 14 and adult animals. GABA_BR₁ immunoreactivity was found to be expressed by perikarya of striatal neurons at all ages examined. In contrast, GABA_BR₂ immunoreactivity was mainly observed in choline acetyltransferase-positive striatal interneurons in PND 14 and adult rats. The present results indicate that there are differential patterns of developmental expression of GABA receptor subunits in the neostriatum, with important implications for the development of GABA systems in rat basal ganglia.

Differential expression of N-methyl- d-aspartate receptor subunit messenger ribonucleic acids and immunoreactivity in the rat neostriatum during postnatal development

The present study was performed to investigate the patterns of gene expression of N-methyl- d-aspartate (NMDA) receptors (NRs) in the rat neostriatum during postnatal development. Reverse transcriptase–polymerase chain reactions (RT–PCR) indicated that levels of NR1, NR2A and NR2D mRNAs reached peak levels between postnatal days 7 (PND 7) and PND 14. The levels of NR2B and NR2C mRNAs were low at PND 1 and their levels increased at PND 7 and remained high in adults. Immunofluorescence combined with image analysis revealed that the levels of NR1 immunoreactivity rose to its maximum at PND 14. In contrast, NR1 immunoreactivity rose progressively in perikarya of striatal neurons. Levels of NR2A immunoreactivity in the neostriatum were highest in adults. However, levels of NR2A immunoreactivity were higher in striatal neurons at PND 1 and PND 7. Levels of NR2B immunoreactivity were highest at PND 7. In the perikarya of striatal neurons however, the highest levels of NR2B immunoreactivity were detected at PND 14 and adult striatal neurons. In addition, double immunofluorescence revealed that the levels of NR1 immunoreactivity increased but the levels of NR2A immunoreactivity were the same in parvalbumin (PV)-positive striatal interneurons of PND 14 and adult rats. NR2B immunoreactivity was not detected in PV-positive neurons of PND 14 rats, but intense NR2B labeling was seen in PV-positive neurons of adult rats. Last but not least, in choline acetyltransferase (ChAT)-positive striatal interneurons of PND 14 and adult rats, levels of NR1 and NR2A immunoreactivity was seen to increase. Level of NR2B immunoreactivity remained the same in ChAT-positive neurons of PND 14 and adult rats. The present results indicate that there are differential patterns of expression of NR mRNAs and immunoreactivity in the neostriatum during different stages of postnatal development. Different combinations of NR have been found in different subpopulations of striatal neurons at different developmental stages. NR1, NR2A and NR2B are the major NMDA receptor subunits expressed during development. The change in patterns of expression of NR may be related to the functional maturation of neurons in the neostriatum.

Change in concentrations of luteinizing hormone subunit messenger ribonucleic acids in the estrous cycle of beef cattle

Changes in the concentrations of LH subunit messenger ribonucleic acids (mRNAs) and in the LH content of the anterior pituitary of beef cattle were studied during the estrous cycle. Japanese beef cows were classified according to the expected day of the estrous cycle: stage I (early-luteal phase, days 1–4; day 1=day of ovulation), stage II (early-mid-luteal phase, days 5–10), stage III (late-mid-luteal phase, days 11–17) and stage IV (follicular phase, days 18–20), according to the morphology of the ovaries. The anterior pituitaries of the cows were collected and the levels of α and LHβ subunit mRNAs were determined by slot–blot analyses. The LH content of the anterior pituitary was measured by radioimmunoassay. The level of α subunit mRNA in the pituitary of cows was highest in stage I and decreased significantly by stage II ( P<0.05); thereafter it tended to increase. The level of LHβ subunit mRNA did not change significantly during the estrous cycle. The LH content of the pituitary of cows was low in stage I and tended to increase by stage II, then to decrease from stage II to III, and to increase significantly from stage III to IV ( P<0.05). These results suggest that the highest levels of gene expressions of α subunit in the anterior pituitary occur in the early-luteal phase of beef cows, while the LH content is increased most in the follicular phase. The enhanced gene expressions of common α subunit in the early-luteal phase could be important in replenishing the bovine anterior pituitary with LH, which is depleted of hormone by the LH surge or the enhanced pulsatile release.

Differential regulation of pituitary gonadotropin subunit messenger ribonucleic acid levels in photostimulated Siberian hamsters.

FSH levels begin to rise 3-5 days after male Siberian hamsters are transferred from inhibitory short photoperiods to stimulatory long photoperiods. In contrast, LH levels do not increase for several weeks. This differential pattern of FSH and LH secretion represents one of the most profound in vivo examples of differential regulation of the gonadotropins. The present study was undertaken to characterize the molecular mechanisms controlling differential FSH and LH synthesis and secretion in photostimulated Siberian hamsters. First, we cloned species-specific cDNAs for the three gonadotropin subunits: the common alpha subunit and the unique FSHbeta and LHbeta subunits. All three subunits share high nucleotide and predicted amino acid sequence identity with the orthologous cDNAs from rats. We then used these new molecular probes to examine the gonadotropin subunit mRNA levels from pituitaries of short-day male hamsters transferred to long days for 2, 5, 7, 10, 15, or 20 days. Short-day (SD) and long-day (LD) controls remained in short and long days, respectively, from the time of weaning. We measured serum FSH and LH levels by RIA. FSHbeta, LHbeta, and alpha subunit mRNA levels were measured from individual pituitaries using a microlysate ribonuclease protection assay. Serum FSH and pituitary FSHbeta mRNA levels changed similarly following long-day transfer. Both were significantly elevated after five long days (2.3- and 3.6-fold, respectively; P < 0.02) and declined thereafter, but they remained above SD control values through 20 long days. Alpha subunit mRNA levels also increased significantly relative to SD control values (maximum 2-fold increase after seven long days; P < 0.03), although to a lesser extent than FSHbeta. Neither serum LH nor pituitary LHbeta mRNA levels changed significantly following long-day transfer. The results indicate that long-day-associated increases in serum FSH levels in Siberian hamsters reflect an underlying increase in pituitary FSHbeta and alpha subunit mRNA accumulation.

Regulation of lutenizing hormone secretion and subunit messenger ribonucleic acid expression by gonadal steroids in perifused pituitary cells from male monkeys and rats.

The mechanisms by which gonadal steroids regulate gonadotropin secretion remain incompletely understood. As previous studies suggest that the pituitary actions of testosterone (T) and estradiol (E) differ in male primates and rodents, we compared the effects of 10 nM T, 0.1 nM E, and 10 nM dihydrotestosterone (DHT) on the LH response to hourly pulses of GnRH as well as the GnRH receptor (GnRH-R) and LH subunit messenger RNA (mRNA) levels in dispersed pituitary cells from intact male monkeys and rats. T suppressed (P < 0.01) and E increased (P < 0.05) GnRH-stimulated LH secretion by rat pituitary cells. With monkey pituitary cells, on the other hand, there was no significant effect of either T or DHT on GnRH-stimulated LH secretion. In E-treated monkey cells, a period of initial enhancement (P < 0.05) was followed by significant suppression (P < 0.05) of LH secretion. GnRH-R mRNA was unchanged by T or E in either rat or monkey cells. T suppressed LHbeta (P < 0.01) and alpha-subunit (P < 0.01) mRNAs, whereas E increased alpha-subunit (P < 0.01), but did not alter LHbeta mRNA levels in rat cells. In monkey cells, however, neither T nor E affected LHbeta or alpha-subunit mRNA levels significantly. Our results identify different regulatory mechanisms by which testicular steroid hormones control LH secretion by the pituitary in male primates and rodents. We propose that the primary site of androgen negative feedback in the male primate is to restrain GnRH pulsatile secretion, whereas in the male rat T also decreases gonadotropin synthesis and secretion by directly affecting the pituitary. E suppresses GnRH-stimulated LH secretion in the primate pituitary, but amplifies the action of GnRH in the rat. Our data also reveal that the action of T to suppress LH secretion and subunit mRNA in male rats is not through decreased GnRH-R gene expression.

Regulation of Lutenizing Hormone Secretion and Subunit Messenger Ribonucleic Acid Expression by Gonadal Steroids in Perifused Pituitary Cells from Male Monkeys and Rats

Regulation of Lutenizing Hormone Secretion and Subunit Messenger Ribonucleic Acid Expression by Gonadal Steroids in Perifused Pituitary Cells from Male Monkeys and Rats

IL-12 receptor beta2 and CD30 expression in paranasal sinus mucosa of patients with chronic sinusitis.

The aetiology of chronic sinusitis is still poorly understood. The expression of T-helper 1 (Th1) and T-helper 2 (Th2) cell markers, interleukin (IL)-12 receptor beta2 subunit (IL-12Rbeta2) messenger ribonucleic acid (mRNA) and CD30, respectively, were investigated in the paranasal sinus mucosa of patients with chronic sinusitis in an attempt to elucidate the involvement of Th1 and Th2 cells in this disease. Anterior ethmoidal mucosae were surgically obtained from two groups of patients with chronic sinusitis: those who had allergic rhinitis (allergic group, n=11) and those without allergy (nonallergic group, n=11). IL-12Rbeta2 mRNA was quantified by means of the reverse transcription polymerase chain reaction, and CD30-positive cells were examined immunohistochemically. Both IL-12Rbeta2 mRNA and CD30 were expressed in the sinus mucosa of the allergic and nonallergic groups. The proportion of mononuclear cells which were CD30-positive in the sinus mucosa was significantly greater in the allergic than in the nonallergic group. The expression levels of IL-12Rbeta2 mRNA were virtually equivalent in both groups. These results suggest a T-helper 2-dominated mucosal reaction in the allergic compared to the nonallergic group, and indicate T-helper 1 activity in the sinus mucosa of both groups. The ubiquity of T-helper 1 cells suggests that they play a role in maintaining local mucosal defences against foreign antigens, which continually enter the upper respiratory tract.

Regulation of Pituitary Follistatin and Inhibin/Activin Subunit Messenger Ribonucleic Acids (mRNAs) in Male and Female Rats: Evidence for Inhibin Regulation of Follistatin mRNA in Females

Regulation of Pituitary Follistatin and Inhibin/Activin Subunit Messenger Ribonucleic Acids (mRNAs) in Male and Female Rats: Evidence for Inhibin Regulation of Follistatin mRNA in Females

Regulation of pituitary follistatin and inhibin/activin subunit messenger ribonucleic acids (mRNAs) in male and female rats: evidence for inhibin regulation of follistatin mRNA in females.

The regulation of FSHbeta messenger RNA (mRNA) expression is complex and involves signals from the hypothalamus and gonads. Additionally, the local (pituitary) production of activin and follistatin appears to serve as an important modulator of endocrine signals for FSHbeta regulation. The purpose of these studies was to identify factors controlling pituitary activin/inhibin subunit and follistatin mRNA production in male and female rats. Both males and females expressed the follistatin, inhibin alpha, and betaB mRNAs, whereas the betaA mRNA was not detected. In males, levels of FSHbeta and follistatin were higher than those in females. After gonadectomy, levels of FSHbeta and follistatin increased in both sexes, whereas betaB rose only in females. In males, blockade of GnRH action from the time of castration prevented the increase in FSHbeta and follistatin, suggesting that GnRH is the primary stimulus for these gene products. In females, treatment with a GnRH antagonist only partially prevented the rise in FSHbeta, follistatin, and betaB expression, suggesting that other factors were also important. Passive immunoneutralization of circulating inhibin increased FSHbeta and follistatin (but not betaB), providing evidence that inhibin is a physiological regulator of follistatin. Replacement of estradiol at the time of ovariectomy prevented the increase in betaB mRNA, suggesting that gonadal steroids may also act via local factors to regulate FSHbeta. In summary, these studies provide evidence that GnRH, gonadal steroids, and gonadal peptides probably regulate FSHbeta expression at least in part via the intrapituitary activin/follistatin system.

Second trimester levels of maternal serum total activin A and placental inhibin/activin alpha and betaA subunit messenger ribonucleic acids in Down syndrome pregnancy.

Previous data have shown that inhibin A (alpha/betaA) is increased about twofold in maternal serum samples from Down syndrome pregnancy. Our objectives were to determine whether activin A (betaA/betaA) was similarly increased in maternal serum from pregnancies affected with fetal Down syndrome, and to investigate whether increased expression of each inhibin/activin subunit occurred in placental tissue from cases of fetal Down syndrome. Maternal serum total activin A levels were measured in 20 cases of fetal Down syndrome and 100 unaffected pregnancy samples. In addition, analysis of inhibin/activin alpha and betaA subunit mRNA levels was performed in placental tissue extracts from six cases of fetal Down syndrome and six tissues with a normal karyotype. The median total activin A level in the Down syndrome cases was 0.82 MoM (multiples of the median); values did not differ significantly (P = 0.36, Mann-Whitney U analysis) from those in unaffected pregnancies. The inhibin alpha subunit/GAPDH mRNA ratio, but not that of betaA subunit/GAPDH mRNA, was significantly greater (P < 0.01, ANOVA) in placental tissue from Down syndrome than in control placental tissue. Unlike inhibin A, activin A is not significantly increased in Down syndrome relative to unaffected pregnancy. Furthermore, increased amounts of maternal serum inhibin A in Down syndrome pregnancy probably result from increased placental expression of inhibin alpha, but not betaA, subunit.

Posttranscriptional Regulation of Thyrotropin -Subunit Messenger Ribonucleic Acid by Thyroid Hormone in Murine Thyrotrope Tumor Cells: A Conserved Mechanism across Species

Posttranscriptional Regulation of Thyrotropin -Subunit Messenger Ribonucleic Acid by Thyroid Hormone in Murine Thyrotrope Tumor Cells: A Conserved Mechanism across Species

Inhibin‐B and pro‐αC‐containing inhibins in amniotic fluid from chromosomally normal and Down syndrome pregnancies

In second-trimester Down syndrome pregnancies, levels of inhibin-A (the alpha-betaA dimer) in maternal serum and amniotic fluid (AF) are significantly higher and lower than in normal pregnancy, respectively. Since AF also contains inhibin-B (the alpha-betaB dimer) and precursor inhibins, we have examined whether the secretion of these inhibin isoforms may also be altered in association with Down syndrome. AF from 45 Down syndrome and 150 chromosomally normal pregnancies between 16 and 19 weeks' gestation were analysed, blinded to whether the sample was from a Down syndrome or a normal pregnancy. The median (10th-90th percentiles) inhibin-B level in the control pregnancies increased from 310.0 (80.8-1112.5) pg/ml at 16 weeks to 459.5 (193.7-1386.8) pg/ml at 19 weeks' gestation. The corresponding figures for precursor inhibins (pro-alphaC inhibins) were 541.8 (206.9-1322.8) pg/ml at 16 weeks and 1391.8 (433.3-2652.6) pg/ml at 19 weeks. Expressed as multiples of the median (MOM), the levels of inhibin-B and pro-alphaC inhibins in the Down syndrome samples were 0.85 and 0.79, respectively. Neither was significantly different from the controls. These data suggest that, of the three inhibin subunits, abnormal production or secretion of the inhibin betaA-subunit may underlie the decreased inhibin-A levels previously observed in Down syndrome. Confirmation of this by quantitative assessment of the inhibin subunit messenger ribonucleic acids would now be useful.

Expression of Chicken Thyroid-Stimulating Hormone -Subunit Messenger Ribonucleic Acid during Embryonic and Neonatal Development

The importance of thyroid hormone from embryonic through neonatal life has been documented in both avian and mammalian species. However, the regulation of thyroid hormone production during this period is not completely understood. The objective of this study was to characterize expression of chicken TSHβ messenger RNA (mRNA) compared with that of thyroid hormones and GH in embryonic and neonatal chickens. Total pituitary RNA was extracted on embryonic days (e-) 11, 13, 15, 17, and 19 and neonatal days (d-) 1, 3, 6, 9, and 12 and subjected to ribonuclease protection assays (RPA) for chicken TSHβ mRNA. TSHβ mRNA levels increased through e-19, with e-19 levels being greater than those at all other embryonic ages (P < 0.05). Levels decreased markedly on d-1, then slowly increased to d-6 and stayed elevated through d-12. RIAs were performed for T4, T3, and GH at the same ages. Serum T4 levels increased slowly from less than 1.0 ng/ml on e-11 to a peak of 6.6 ng/ml on d-1 (P < 0.05). After the peak on d-1, posthatch T4 levels stabilized between 3.5–4.5 ng/ml through d-12 (P < 0.05). T3 concentrations were less than 0.25 ng/ml on e-11, increased dramatically between e-19 and d-1 (P < 0.05), and remained high throughout the rest of the experiment, with a concentration of 3.25 ng/ml on d-6 (P < 0.05). GH levels for e-11 through e-17 were below the sensitivity of the GH RIA. On e-19, the GH level was 3 ng/ml and continued to increase through d-12 to a level of 130 ng/ml. As thyroid hormone levels were preceded by maximal TSHβ mRNA levels on e-19, we next determined whether TSHβ gene expression on e-19 was under TRH and T3 regulation. E-19 anterior pituitary cells were cultured in serum-free medium with either TRH (10−8) or T3 (10−8) for 20–24 h. Treatment with T3 significantly decreased levels of TSHβ mRNA (P < 0.05). However, TRH did not produce a significant increase in TSHβ mRNA, although TRH did increase TSHβ mRNA by 60%, on the average, in this study. Therefore, these results indicate that an increase in pituitary TSH production probably regulates thyroid hormone levels during late embryonic development and that negative feedback inhibition of TSH production by thyroid hormones also exists at this critical developmental stage.

Pituitary follistatin and inhibin subunit messenger ribonucleic acid levels are differentially regulated by local and hormonal factors.

Follistatins, activins, and inhibins are expressed in a wide range of tissues where they function as autocrine and/or paracrine factors. Activin B (beta B beta B) and inhibin B (alpha beta B) are the predominant forms expressed in the rat anterior pituitary. This study was designed to evaluate the regulation of the messenger RNAs (mRNAs) for inhibin alpha and beta B, and follistatin, relative to each other, using cultured rat anterior pituitary cells. Activin A stimulated follistatin (a maximal 4-fold stimulation by 6 h) and beta B (a maximal 1.7-fold stimulation after 2 h) mRNA levels. Although inhibin A dramatically decreased follistatin mRNA levels (34% of the control value after 24 h), it only marginally affected those of beta B (86% of the control value after 2 h). Follistatin inhibited the accumulation of its own mRNA (46% of the control value after 6 h), but had no statistically significant effect on beta B or alpha mRNA levels. Inhibin A was the only treatment that had an effect on alpha mRNA levels, causing a slight decrease (82% of the control value by 24 h). The effects of activin A and inhibin A on follistatin and beta B mRNA levels were dose dependent. Moreover, follistatin and inhibin A blocked the effects of activin A. Immunoneutralization experiments were performed to determine whether locally secreted activin B regulates the expression of these three mRNAs. A monoclonal antibody to activin B reduced follistatin and beta B mRNA levels (37% and 73% of the control value, respectively) and enhanced the stimulatory effect of exogenous activin A on these mRNAs (840% vs. 300% and 170% vs. 145% of the control value, respectively); there was no change in alpha mRNA accumulation. GnRH and activators of the protein kinase A (forskolin) and protein kinase C (12-O-tetradecanoylphorbol acetate) pathways also had differential effects on follistatin, beta B, and alpha mRNA levels. GnRH stimulated follistatin mRNA levels, but suppressed those of beta B. 12-O-Tetraphorbol acetate had no effect on beta B, but stimulated follistatin mRNA levels to the same extent as forskolin. Of these agents, only forskolin produced a marginal inhibitory effect on alpha mRNA accumulation. Testosterone decreased both follistatin and beta B mRNA levels without affecting those of alpha. The results of this study demonstrate that the local production of rat anterior pituitary follistatin, activin B, and inhibin B is regulated by hypothalamic, peripheral, and local factors in such a way that the ratios between activin B and its two inactivators, follistatin and inhibin B, are very tightly maintained.

Pituitary follistatin and inhibin subunit messenger ribonucleic acid levels are differentially regulated by local and hormonal factors

Pituitary follistatin and inhibin subunit messenger ribonucleic acid levels are differentially regulated by local and hormonal factors

Corticosterone selectively increases follicle-stimulating hormone beta- subunit messenger ribonucleic acid in primary anterior pituitary cell culture without affecting its half-life

Corticosterone selectively increases follicle-stimulating hormone beta- subunit messenger ribonucleic acid in primary anterior pituitary cell culture without affecting its half-life

Direct actions of the luteinizing hormone-releasing hormone agonist, deslorelin, on anterior pituitary contents of luteinizing hormone (LH) and follicle-stimulating hormone (FSH), LH and FSH subunit messenger ribonucleic acid, and plasma concentrations of LH and FSH in castrated male cattle.

The objective in this study was to characterize direct effects of the LHRH agonist, deslorelin, on anterior pituitary gland function in male cattle in the absence of gonadal feedback. Castrated bulls (steers), 30 mo old, were allocated to four groups: group 1, control, no treatment (n = 8); group 2, five deslorelin implants (approximately 250 micrograms total deslorelin/day) for 42 days (n = 8); group 3, control+ LHRH (50 micrograms i.m.) at weekly intervals (n = 3); group 4, five deslorelin implants+LHRH as for group 3 (n = 3). Plasma LH was similar (p > 0.05) for steers in groups 1 and 2 on Day 0 and lower (p < 0.05) for steers in group 2 on Day 4, and continued to decrease to Day 41 (group 1, 1.71 +/- 0.20 ng/ml [mean +/- SEM]; group 2, 0.38 +/- 0.03 ng/ml [p < 0.001]). Mean plasma concentrations of FSH were similar (p > 0.05) for steers in groups 1 and 2 on Day 0 and lower (p < 0.05) for steers in group 2 on Day 7, and declined to Day 41 (group 1, 43.5 +/- 3.9 ng/ml; group 2, 17.5 +/- 1.5 ng/ml [p < 0.001]). Steers in group 3 showed increases in plasma LH after injection of LHRH on all occasions, while steers in group 4 did not show increases in plasma LH from Day 14 onward. Mean relative pituitary contents (arbitrary units) of LH beta- and FSH beta-subunit mRNAs were reduced on Day 42 in steers treated with deslorelin (LH beta: groups 1 and 3, 1.56 +/- 0.27; groups 2 and 4, 0.08 +/- 0.01 [p < 0.001]; FSH beta: groups 1 and 3, 1.01 +/- 0.08; groups 2 and 4, 0.34 +/- 0.07 [p < 0.001]). However, alpha-subunit mRNA was similar for control steers and steers treated with deslorelin (groups 1 and 3, 1.00 +/- 0.11; groups 2 and 4, 0.86 +/- 0.12 [p > 0.1]). Pituitary content of LH, but not FSH, was reduced in steers treated with deslorelin. In summary, steers treated with deslorelin showed desensitization to natural LHRH, and this was associated with reduced pituitary contents of LH and FSH beta-subunit mRNAs, a reduction in pituitary content of LH, and decreases in plasma concentrations of LH and FSH. This demonstrated, for the first time, a direct action of LHRH agonist on LH and FSH beta-subunit gene expression in cattle, independent of gonadal feedback. Also, there was a differential effect of treatment with deslorelin on gonadotropin alpha- and beta-subunit mRNA contents in the anterior pituitary.

Effect of testosterone on maturational gonadotropin subunit messenger ribonucleic acid levels in the goldfish pituitary.

In this study, we investigated the effects of testosterone and a nonaromatizable androgen, 11 beta-hydroxyandrosterone, on maturational gonadotropin (GtH-II) subunit gene expression in the goldfish pituitary. While testosterone treatment at physiological doses resulted in stimulation of GtH-II-alpha and -beta subunit mRNA production, time-course and dose-response studies performed on sexually immature goldfish of mixed sex, using a wider dose range exceeding physiological levels, demonstrated a biphasic response to in vivo androgen treatment. Time-related treatment with testosterone and 11 beta-hydroxyandrosterone (20 micrograms/fish) resulted in an initial inhibition of GtH-II subunit mRNA production (12-24 h) followed by stimulation at 72-96 h. In dose-response studies, treatment for 24 h with testosterone resulted in a significant stimulation at the low physiological doses of 0.2 and 2 micrograms/fish. At the supraphysiological level of 20 micrograms/fish, testosterone treatment resulted in no stimulation or in decreased GtH-II subunit mRNA levels compared to the control values. Similarly, treatment with 11 beta-hydroxyandrosterone resulted in a significant stimulation of GtH-II subunit mRNA levels at low physiological concentrations (0.2 microgram/ fish) and an inhibition, or no stimulation, at higher concentrations (2-20 micrograms/fish). In sexually mature goldfish of mixed sex, the biphasic effect of testosterone was not observed in vivo, and treatment with this steroid resulted in stimulation of GtH-II subunit mRNA production in a dose-related manner. To investigate the direct action of testosterone, studies were carried out using isolated goldfish pituitary fragments from goldfish of mixed sex in vitro. Treatment with testosterone at various concentrations was found to stimulate GtH-II subunit mRNA production in pituitary glands obtained from both sexually immature and sexually mature goldfish. Overall, the present study demonstrates a stimulatory effect of testosterone on GtH-II subunit mRNA levels in goldfish. The observed stimulation of basal GtH-II subunit mRNA production by testosterone occurs, in part, through a direct action at the level of the pituitary in both sexually immature and mature goldfish.

Transforming growth factor-β1 and -β2 induce inhibin and activin βB-subunit messenger ribonucleic acid levels in cultured human granulosa-luteal cells

To examine the effect of transforming growth factor-beta (TGF-beta) on inhibin and activin subunit messenger ribonucleic acids. Human granulosa-luteal cell culture model. Granulosa cells were obtained from women undergoing an IVF program in a private IVF clinic. Regularly menstruating women undergoing oocyte retrieval for IVF because of either tubal obstruction or infertility of the spouse. For each experiment, cells of two to four patients were pooled, enzymatically dispersed, separated from red blood cells by centrifugation through Ficoll-Paque and cultured in vitro in the presence of TGF-beta 1 or TGF-beta 2 and/or hCG whereafter cellular RNA was extracted for Northern or dot blot filter hybridization with inhibin alpha-, beta A, and beta B-subunit complementary DNA probes. Both TGF-beta 1 and TGF-beta 2 induced the expression of a 4.8-kb inhibin and activin beta B-subunit messenger (mRNA) transcript in a time- and dose-dependent manner but had no effect on alpha- or beta A-subunit mRNA levels. Human chorionic gonadotropin alone did not affect beta B-subunit mRNA levels, but when administered together with TGF-beta s, it prevented the induction of beta B-subunit mRNAs. Our results suggest that in human ovary, granulosa, or thecal cell-derived TGF-beta 1 or -beta 2 may eventually locally modulate in a paracrine or autocrine manner the relative expression levels of inhibin and activin subunits favoring the formation of the inhibin and activin dimers containing the beta B-subunit. The effect of TGF-beta is clearly different from that of gonadotropins, which potently induce the alpha- and beta A-subunit mRNAs, indicating that distinct components of the human ovarian inhibin and activin system are regulated differentially by endocrine and local factors.

Inhibin alpha and beta A subunit messenger ribonucleic acids expression in human pituitary adenomas: studies by quantitative reverse transcription-polymerase chain reaction.

Inhibin alpha and beta A subunit messenger ribonucleic acid (mRNA) levels were measured quantitatively by reverse transcription-polymerase chain reaction (RT-PCR) in human pituitary adenomas. The inhibin alpha subunit mRNA levels were undetectably low in cultured adenoma tissues, but beta A mRNA were 0.383 +/- 0.074 in 3 GH adenomas, 0.672 +/- 0.140 in 3 prolactinomas and 0.957 +/- 0.414 molecules/cell in 3 non-functioning adenomas. The addition of 10(-8) M activin A decreased the beta A mRNA levels within 4 h in 1 of 3 GH adenomas, 2 of 3 prolactinomas and 2 of 3 non-functioning adenomas, though the decreases were not statistically significant. The results showed an abundance of beta A subunit mRNA compared with alpha subunit mRNA in all human pituitary adenomas and a local role for activin in its own production through inhibin beta A mRNA subunit expression.