SUMMARY

The effects of ovariectomy and ovariectomy and treatment with oestradiol benzoate (OB) on the basal concentration of thyrotrophin (TSH), the total concentrations and concentrations of free tri-iodothyronine (T₃) and thyroxine (T₄), and the concentrations of TSH, T₃ and T₄ observed after treatment with thyrotrophin releasing hormone (TRH) were studied in old (16–17 months of age) constant oestrous and young (3–4 months of age) oestrous rats. The untreated old control rats had significantly (P< 0·001) lower basal total T₄ concentrations and percentage and absolute concentrations of free T₄ and lower percentage and absolute concentrations of free T₃ than untreated young rats. The basal levels of TSH in these two groups were similar and the increases in TSH after injection of TRH were identical. Two weeks after ovariectomy, no significant additional differences in hormone concentrations between old and young rats were observed. However, release of TSH induced by TRH was increased by three- to fourfold in old rats after ovariectomy compared with nine- to tenfold in young ovariectomized rats (P<0·01). Basal T₄ concentrations remained unchanged in old ovariectomized rats treated for 7 days with 2 μg OB/day compared with both intact and ovariectomized rats. However, T₄ concentrations in OB-treated young rats were significantly (P<0·001) reduced. Treatment with OB significantly increased both basal and TRH-induced T₃ and TSH levels in old and young rats although the young rats showed a greater response (P<0·001). Two hours after injection of TRH, serum T₃ concentrations in old rats increased only after OB treatment and not after ovariectomy alone or in intact rats, whereas T₃ concentrations rose in all three groups of young animals.

These results indicate that (1) older female rats have lower total T₄, free T₄ and free T₃ concentrations and a lower TSH response to TRH, (2) OB treatment in young rats suppresses serum T₄ but increases serum T₃ and results in a greater TSH response to TRH and (3) at least one of the mechanisms accounting for the alterations in thyroid function observed in the older female rat, in addition to possible concomitant primary thyroid gland hypofunction, is a hyporesponsiveness of pituitary thyrotrophs to both endogenous negative feedback signals from low serum thyroid hormone concentrations and exogenous TRH stimulation.

SUMMARY

Old male rats of 22–24 months and young ones of 3–5 months were studied to find the effects of ageing, of orchidectomy and of orchidectomy and treatment with testosterone propionate (TP) on the basal serum concentrations of thyrotrophin (TSH) and on the total and free concentrations of tri-iodothyronine (T₃) and thyroxine (T₄) in the serum. The changes in TSH after treatment with thyrotrophin releasing hormone (TRH) were also observed. Intact old rats had significantly (P < 0·001) lower basal T₄ and T₃ as well as lower (P < 0·05) testosterone concentrations than were present in young rats. They also had higher basal TSH and per cent free T₄ but lower absolute free T₃ concentrations than had young rats. Two weeks after orchidectomy, basal TSH concentrations were slightly but significantly (P < 0·05) decreased in both young and old rats while T₄ decreased significantly (P < 0·05) only in the young. The responses of TSH to TRH were also reduced by orchidectomy in both age groups with the old rats being less responsive than the young. Orchidectomy and treatment with pharmacological doses of TP produced similar effects on the pituitary-thyrotrophic response for both old and young rats but a greater effect occurred in the basal T₄ response in young rats. In all groups basal TSH was influenced by orchidectomy or by treatment with TP but was always higher in the aged rat. Tri-iodothyronine concentration was always lower in the older rat and was not altered by orchidectomy or by treatment with TP in either young or old rats.

These results indicate that (1) in the male rat these age-specific effects on the thyroid–pituitary system are probably due, not only to a reduction in thyroid gland function and plasma T₄ protein-binding, but also to a concomitant hyporesponsiveness of the aged male rat pituitary thyrotroph to TRH stimulation and (2) there is probably a significant influence of testicular function on the pituitary–thyroid system of the male rat.

The present study was undertaken to determine the tissue-specific expression of estrogen receptor beta (ERbeta), and the effects of a daidzein supplement to the diet of pregnant sows on the expression of ERbeta, and type 1 insulin-like growth factor receptor (IGF-1R) genes in newborn piglets by using semi-quantitative RT-PCR. Eight sows received a dietary supplement of daidzein at a dosage of 8 mg per kg feed from day 85 of gestation, and six sows were used as controls. After parturition, 2 male neonatal piglets were selected from each litter for sampling. ERbeta mRNA was detected in intestine, lung, thymus, kidney, pituitary and hypothalamus tissues, but not in heart, adrenal, skeletal muscle, liver or placental tissues. Daidzein treatment significantly increased the birth weight of male piglets and markedly reduced the level of ERbeta mRNA in the hypothalamus, but not in the pituitary. An up-regulation of IGF-1R gene transcription was observed in skeletal muscles of newborn piglets. In addition, the IGF-1R mRNA was found to be most abundant in pituitary and hypothalamus, followed by skeletal muscle, thymus, and liver tissues in decreasing order. Our results demonstrate that (1) ERbeta is expressed in a tissue-specific manner in newborn piglets, (2) daidzein down-regulates ERbeta gene expression in the hypothalamus, possibly indicating central effects of daidzein, and (3) daidzein influences fetal growth associated with higher IGF-IR gene expression in skeletal muscle.

Conversion of cholesterol to biologically active steroids is a multi-step enzymatic process. Along with some important enzymes, like cholesterol side-chain cleavage enzyme (P450scc) and 3β-hydroxysteroid dehydrogenase/isomerase (3β-HSD), several proteins play key role in steroidogenesis. The role of steroidogenic acute regulatory (StAR) protein is well established. A novel protein, BRE, found mainly in brain, adrenals and gonads, was highly expressed in hyperplastic rat adrenals with impaired steroidogenesis, suggesting its regulation by pituitary hormones. To further elucidate its role in steroidogenic tissues, mouse Leydig tumor cells (mLTC-1) were transfected with BRE antisense probes. Morphologically the BRE antisense cells exhibited large cytoplasmic lipid droplets and failed to shrink in response to human chorionic gonadotropin. Although cAMP production, along with StAR and P450scc mRNA expression, was unaffected in BRE antisense clones, progesterone and testosterone yields were significantly decreased, while pregnenolone was increased in response to human chorionic gonadotropin stimulation or in the presence of 22(R)OH-cholesterol. Furthermore, whereas exogenous progesterone was readily converted to testosterone, pregnenolone was not, suggesting impairment of pregnenolone-to-progesterone conversion, a step metabolized by 3β-HSD. That steroidogenesis was compromised at the 3β-HSD step was further confirmed by the reduced expression of 3β-HSD type I (3ß-HSDI) mRNA in BRE antisense cells compared with controls. Our results suggest that BRE influences steroidogenesis through its effects on 3β-HSD action, probably affecting its transcription.

Abstract

We have reported that MK-0677 is a novel, orally active GH secretagogue that stimulates an immediate and longlasting increase in serum GH levels in dogs. Significant elevations in IGF-I levels were associated with the increased GH secretion. Cortisol secretion was also increased following MK-0677 administration. In the current study, we determined the effect of repeat oral administration of MK-0677 on GH, IGF-I and cortisol levels; we also investigated if the GH and cortisol responses to MK-0677 are influenced by circulating IGF-I concentrations.

Following the initial oral administration of MK-0677, GH secretion (area under the time–response curve (AUC) ng/ml per h) was increased 7·9- to 9·8-fold (1·0 mg/kg), 5·6-fold (0·5 mg/kg) or 3·9-fold (0·25 mg/kg). With repeat MK-0677 administration, the GH response was decreased by 41–77%; GH concentrations remained significantly above control in the 0·5 mg/kg and 1·0 mg/kg groups. Individual beagle GH profiles indicated that the increased GH concentration was associated with an amplified GH pulsatile profile.

Serum IGF-I levels were significantly increased over control levels at all dosage levels by 480 min on the first day of MK-0677 administration. With repeated administration, IGF-I levels were increased up to 126% and remained elevated through 14 days, the longest treatment period evaluated. While daily MK-0677 administration appeared to increase IGF-I levels over 24 h, as evidenced by significant increases in the pretreatment IGF-I levels on days 4–14, no such increase was noted with alternate day MK-0677 administration; thus the dosage regimen modulated circulating IGF-I levels.

MK-0677 stimulated increases in cortisol secretion (AUC μg/dl per h) on the first day of treatment. A decreased cortisol response was observed following repeated daily treatment with MK-0677; in contrast, with alternate day treatment, no decrease in cortisol response to MK-0677 occurred.

A marked increase in circulating IGF-I concentrations following administration of exogenous GH resulted in a significant decrease in both the GH and cortisol response to MK-0677 compared with control animals. Our findings suggested, therefore, that circulating IGF-I concentrations regulate GH and cortisol response to MK-0677.

In summary, chronic oral administration of MK-0677 was associated with significant increases in GH and IGF-I levels that were maintained for the duration of the treatment. The GH profile following MK-0677 administration consisted of episodic increases above control. Compared with day 1, repeated daily treatment with MK-0677 resulted in an attenuated GH response that was associated with an increase in circulating IGF-I levels. The cortisol response was similarly reduced during chronic MK-0677 treatment, suggesting that IGF-I mediated negative feedback on both the GH and cortisol axes. The fact that similar attenuation of the GH and cortisol responses to MK-0677 on day 1 was observed if IGF-I levels were increased by treating animals with exogenous GH suggested that the attenuated response to MK-0677 that occurred during chronic treatment was mediated by increases in IGF-I rather than desensitization to MK-0677. Thus, a regulatory feedback loop apparently prevents hyperstimulation of the GH axis by MK-0677. We conclude that MK-0677 offers the potential of an orally active GH secretagogue that can maintain elevated IGF-I levels when administered chronically.

Journal of Endocrinology (1997) 152, 183–192

Abstract

L-692,585 is a 2-hydroxypropyl derivative of L-692,429, both novel non-peptidyl growth hormone (GH) secretagogues. The effects of single and repeated intravenous administration of L-692,585 on serum or plasma GH and other hormones in beagles were evaluated. In a balanced 8-dog dose-ranging study, compared to the saline control with a mean (± s.e.m.) after-dose serum GH peak of 6·1 ± 1·3 ng/ml, L-692,585 significantly increased (P<0·05) peak GH concentrations 4·3-fold (32·5 ± 7·0 ng/ml) at a dose of 0·005 mg/kg, 7-fold (49·4±10·6 ng/ml) at a dose of 0·02 mg/kg, and 21-fold (134·3±29·0 ng/ml) at a dose of 0·10 mg/kg. Total GH release, expressed as area under the curve, showed a similar dose-dependent increase. Peak GH levels were recorded at 5 or 15 min after dosing with the levels returning to near baseline by 90 min. Serum cortisol levels were increased above saline control levels in a dose-dependent manner; however, the increases were modest compared to the GH increases. Based on peak responses and total GH release, L-692,585 was 10- to 20-fold and 2- to 2·5-fold more potent than L-692,429 and the growth hormone releasing peptide, GHRP-6, respectively. When L-692,585 was administered once daily for 14 consecutive days at 0, 0·01 or 0·10 mg/kg to each of 6 dogs, peak plasma GH levels and total GH release on days 1, 8 and 15 significantly increased in a dose-dependent manner, and no desensitization was evident. Mean peak levels ranged from 42 to 50 ng/ml and from 64 to 100 ng/ml for the 0·01 and 0·10 mg/kg doses, respectively. Six hours after dosing, IGF-1 levels were increased on each of the sampling days. Mean adrenocorticotrophic hormone and cortisol levels were modestly elevated transiently on each of the sampling days, while prolactin, insulin and thyroxine levels were unaltered over the course of the study.

These studies demonstrated that L-692,585 is a potent GH secretagogue that induces an acute, transient increase in GH levels. The magnitude of the response is not down-regulated following repeated daily administration for 14 days. IGF-1 levels are increased for an extended period following each treatment.

Journal of Endocrinology (1994) 143, 399–406

The possible direct effect of corticosterone on release of pituitary prolactin was examined in a system using incubation for 8 h. Corticosterone at either 0·1 or 1 μg/ml medium had no significant effect on in-vitro prolactin release but 10 or 100 μg/ml medium produced a significant inhibition of release of prolactin. Release of LH, FSH and thyroid-stimulating hormone were not altered by 0·1, 1 or 10 μg corticosterone/ml, indicating that its action at the concentration of 10 μg/ml was specific on release of prolactin. Corticosterone injected at doses of 1 or 5 mg/kg into hypophysectomized rats with two pituitary grafts underneath the kidney capsule produced a significant fall in serum levels of prolactin when compared with control hypophysectomized rats with two pituitary grafts. Examination with the electron microscope showed that about one third of the lactotrophes from adrenalectomized rats after corticosterone injection exhibited patterns which suggested a decrease in protein synthesis when compared with lactotrophes from adrenalectomized rats given only the vehicle injection. These observations indicated that inhibition of release of prolactin by corticosterone could be exerted directly on the pituitary gland, and that the rise of serum levels of prolactin after adrenalectomy might have been due to the removal of direct inhibition by corticosterone.

Male rats were adrenalectomized and 2–3 weeks later, concentrations of dopamine and noradrenaline in the medial basal hypothalamus were measured and found not to be different from values in intact rats. Dopamine metabolism also was not altered in the median eminence. The dopaminergic agonist, l-DOPA, inhibited, and the antagonists, pimozide and haloperidol, stimulated release of prolactin in both adrenalectomized and intact rats. Serotonin (5-HT) metabolism in the medial basal hypothalamus and anterior hypothalamus of adrenalectomized rats was not significantly different from values in intact rats, but a higher concentration of 5-HT was observed in the medial basal hypothalamus of adrenalectomized rats when compared with the values in intact rats. A serotonergic agonist, fluoxetine, and an antagonist, cyproheptadine, had no apparent effect on release of prolactin in intact rats, but fluoxetine produced a significant rise, and cyproheptadine, a significant lowering of serum levels of prolactin in adrenalectomized rats. These results suggest that 5-HT, but not dopamine, may be involved in the rise of prolactin after adrenalectomy.

Abstract

To investigate the effect of hypophyseal transection (HST) on GH secretagogue activity of the non-peptidyl GH secretagogue L-692,585 in the conscious pig, male castrated swine were randomly assigned to either a hypophyseal stalk transection group (HST; n=3) or to a sham-operated control group (SOC; n=3). Treatments administered were L-692,585 (100 γg/kg), human GH-releasing factor(1–29)NH₂ (GRF; 20 γg/kg) or L-692,585 (100 γg/kg) + GRF (20 γg/kg) on days −7 to −3 before surgery and days +3 to +8 after surgery. To evaluate the integrity of the pituitary gland, the animals were challenged with corticotropin-releasing hormone (CRH; 150 γg) or GnRH (150 ng/kg) both before and after surgery. Blood was collected from −60 to +180 min post treatment and assayed for GH, cortisol and LH.

Before surgery, no significant difference (P>0·05) in peak GH response (ng/ml) was present between the two groups (SOC vs HST) in response to L-692,585 (101 ± 12 vs 71 ± 9) or L-692,585 + GRF (171 ± 21 vs 174 ± 21). Only two out of three SOC vs three out of three HST pigs responded to GRF (13 ± 2 vs 25 ± 3) resulting in a significant difference between groups. Following surgery, significant differences were present in peak GH response (ng/ml) between SOC and HST groups following L-692,585 (79 ± 6 vs 13·8 ± 1·0); however, the response to L-692,585 + GRF was similar (115 ± 8 vs 94 ± 7). All animals responded to GRF; however, a significant difference was present between groups due to the magnitude of the responses. Whereas the cortisol responses (ng/ml) to L-692,585 in the SOC and HST groups were similar before surgery, a significant difference was present after surgery (44·4 ± 6·4 vs 14·6 ± 2·1). No significant difference was noted between the HST and SOC groups in response to CRH or GnRH either before or after surgery.

These results indicated that L-692,585 induced an immediate GH response in the intact animal in contrast to GRF where the GH release was variable. L-692,585 also stimulated an immediate increase in cortisol levels. Transection of the hypophyseal stalk dramatically decreased but did not ablate the GH or cortisol response to L-692,585. Co-administration of L-692,585 + GRF induced an immediate GH response of similar magnitude in the intact and HST animal. We conclude that L-692,585 has a direct but limited action at the level of the pituitary and that an intact hypophyseal stalk is required for a maximal GH and cortisol response. L-692,585 acts with GRF at the level of the pituitary to induce a maximal GH response. These findings suggest that L-692,585 stimulates GH secretion by acting in combination with GRF and interrupting the inhibitory tone of somatostatin on the somatotroph.

Journal of Endocrinology (1996) 148, 371–380

ABSTRACT

A sensitive and specific double-antibody enzyme-linked immunoassay, using a synthetic analogue of human relaxin for standard and immunogen, was developed for the measurement of human relaxin (hRLX) in serum and plasma. No cross-reactivity was observed for human insulin, human insulin-like growth factor-I, hGH, human chorionic gonadotropin, hFSH, hLH or human prolactin. The assay was used to monitor RLX concentrations in samples from men, non-pregnant and pregnant women, and in pregnant rhesus monkeys infused with hRLX. RLX was not detected in serum from men nor from non-pregnant women, while a concentration of 600 ng/l was measured in pooled sera from two pregnant women (pregnancies achieved by in-vitro fertilization). Immunoreactive RLX (1·1 μg/g) was found in human corpora lutea taken from ectopic pregnancies at 7 weeks. In an experiment with a pregnant rhesus monkey infused with human RLX analogue, less than 1·5% of the maternal concentration was measured in the fetal circulation. Even though preliminary, these data suggest a low level of transfer of human analogue relaxin across the placenta in a rhesus monkey. Further studies of the physiology of RLX in human pregnancy will be facilitated by the availability of this immunoassay.

Journal of Endocrinology (1989) 120, 449–457

The solute carrier (SLC) family is a large group of membrane transport proteins. Their dysfunction plays an important role in the pathogenesis of thyroid cancer. The most well-known SLC is the sodium-iodide symporter (NIS), also known as sodium/iodide co-transporter or solute carrier family 5 member 5 (SLC5A5) in thyroid cancer. The dysregulation of NIS in thyroid cancer is well documented. The role of NIS in the uptake of iodide is critical in the treatment of thyroid cancer, radioactive iodide (RAI) therapy in particular. In addition to NIS, other SLC members may affect the autophagy, proliferation, and apoptosis of thyroid cancer cells, indicating that an alteration in SLC members may affect different cellular events in the evolution of thyroid cancer. The expression of the SLC members may impact the uptake of chemicals by the thyroid, suggesting that targeting SLC members may be a promising therapeutic strategy in thyroid cancer.