Activin A is a member of the transforming growth factor-β family and has known roles in the adrenal cortex, from which activin A is secreted. We aimed to find whether activin A induces secretion of catecholamines from chromaffin cells of the adrenal medulla, which neighbours the adrenal cortex in vivo. Using carbon fibre amperometry, we were able to measure catecholamine secretion in real-time from single chromaffin cells dissociated from the rat adrenal medulla. Activin A stimulated catecholamine secretion in a rapid and dose-dependent manner from chromaffin cells. This effect was fully reversible upon washout of activin A. The minimum dose at which activin A had a maximal effect was 2 nM, with an EC₅₀ of 1.1 nM. The degree of secretion induced by activin A (2 nM) was smaller than that due to membrane depolarization caused by an increase in the external K⁺ concentration from 5 to 70 mM. No response to activin A was seen when Ca²⁺ channels were blocked by Cd²⁺ (200 μM). We conclude from these findings that activin A is capable of stimulating a robust level of catecholamine secretion from adrenal chromaffin cells in a concentration-dependent manner. This occurs via the opening of voltage-gated Ca²⁺ channels, causing Ca²⁺ entry, thereby triggering exocytosis. These findings illustrate a new physiological role of activin A and a new mechanism in the control of catecholamine secretion from the adrenal medulla.

Exposure of male golden hamsters to short photoperiods of 6 h light: 18 h darkness led to testicular and accessory sex organ atrophy in 5 weeks. Short photoperiods also significantly depressed serum levels of LH, FSH, prolactin and testosterone in samples obtained by decapitation, but not in samples collected on the preceding day under ether anaesthesia. Injections of luteinizing hormone releasing hormone (LH-RH) at 09.00 h (lights on) or at 15.00 h (lights off) prevented testicular regression when compared with hamsters receiving injection vehicle only. However, the hamsters receiving LH-RH injections at lights on had significantly greater testicular weight and accessory sex organ (seminal vesicles and coagulating glands) weight and testosterone concentration than those receiving LH-RH at lights off. No increase in testicular weight was observed in hypophysectomized male hamsters given the same LH-RH injections and the same lighting regimen.

These results indicate that LH-RH alone can prevent, at least partially, testicular and sex organ atrophy and increase serum testosterone concentration by stimulating release of LH and FSH in hamsters exposed to short photoperiods, involving temporal difference of LH-RH action. Further implications of the results are discussed.

Abstract

A newly synthesised GH-releasing peptide, KP 102 (also named GHRP-2), was studied in an in vitro perifusion system of primary cultured ovine anterior pituitary cells. Application of KP 102 to the perifusion medium caused a dose-dependent increase in GH secretion. Dose-response relationships indicated that KP 102 had similar potency to GRF and was 10-fold more potent than earlier generations of GH-releasing peptide (GHRP-6 and GHRP-1) tested in same system. The response to a second application of KP 102 given within 1 h of initial application was significantly lower than the response to the first application. When KP 102 (or GRF) was applied first and then GRF (or KP 102) given 1 h later, the second response was not attenuated. When GRF and KP 102 were coadministered, an additive effect on release of GH was obtained. The effect of maximal dose of KP 102 (100nM) on GH release was totally abolished by [Ac-Tyr¹, d-Arg²] GRF 1-29 (1μM) which is believed to be a specific antagonist for the GRF receptor. Blockade of Ca²⁺ channels by Cd²⁺ (2mM) diminished the basal GH secretion and abolished the increase in GH release in response to KP 102 (100nM). These data suggest that the action of KP 102 is blocked by a GRF receptor antagonist and therefore acts through a different receptor to that employed by earlier generations of GH-releasing peptides. GH release in response to KP 102 involves an increase in Ca²⁺ influx and there is no cross-desensitization between KP 102 and GRF responses.

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.

Abstract

The present study was undertaken to investigate the metabolic regulation of insulin-like growth factor binding proteins (IGFBPs) gene expression in muscles from diabetic or fasted rat. The messenger RNA (mRNA) levels for IGFBP-2 and -4 were analysed by solution hybridization in heart, skeletal and smooth muscle and liver from fasted (3 days) and refed (6, 12, 24, 72 h) rats and rats made diabetic with streptozotocin. In aortic intima-media, the mRNA levels for IGFBP-2 and -4 were decreased by diabetes or fasting and were restored gradually by refeeding. The response of IGFBP-4 mRNA to diabetes appeared two days after injection of streptozotocin, while a significant decrease of IGFBP-2 mRNA was found after a diabetes duration of two weeks. Both diabetes and fasting decreased IGFBP-4 mRNA levels in heart muscle and skeletal muscle and refeeding restored mRNA for IGFBP-4 to normal level. IGFBP-2 mRNA was undetectable in heart muscle and skeletal muscle. In liver IGFBP-4 mRNA was abundantly expressed. It was slightly but significantly decreased by fasting and approached normality with refeeding, while no change was found in diabetic liver. In contrast, liver IGFBP-2 mRNA was much lower in amount than IGF-I mRNA and IGFBP-4 mRNA and was sharply elevated by fasting, and decreased by refeeding. In conclusion, 1) both IGFBP-2 and -4 mRNA in various tissues are regulated by diabetes or fasting; 2) the mRNA for IGFBP-2 is metabolically regulated in a discordant, organ-specific manner.

Journal of Endocrinology (1994) 143, 235–242

Abstract

The involvement of protein kinase C (PKC) in the action of GH-releasing factor (GRF) and synthetic GH-releasing peptides (GHRP-2 and GHRP-6) was investigated in ovine somatotrophs in primary culture. In partially purified sheep somatotrophs, GRF and GHRP-2 caused translocation of PKC activity from the cytosol to the cell membranes and caused GH release in a dose- and time-dependent manner. GHRP-6 did not cause PKC translocation. The PKC inhibitors, calphostin C, staurosporine and chelerythrine, partially reduced GH release in response to GRF and GHRP-2 at doses which selectively inhibit PKC activity. These inhibitors totally abolished GH release caused by phorbol 12-myristate 13-acetate (PMA). Down-regulation of PKC by the treatment of cells with phorbol 12,13-dibutyrate for 16 h caused a significant (P<0·001) reduction in total PKC activity and totally abolished PKC translocation in response to a challenge with GRF, GHRP-2 or PMA. In addition, down-regulation abolished GH release in response to GRF, GHRP-2 or GHRP-6. Treatment of cells with H₈₉, a selective PKA inhibitor, totally blocked GH release caused by either GRF or GHRP-2 and partially reduced PMA-induced GH release. H₈₉ had no effect on PKC translocation caused by GRF, GHRP-2 or PMA and did not affect GH release caused by GHRP-6. These data suggest that GHRP-2 and GRF activate PKC in addition to stimulating adenylyl cyclase activity. Although the cAMP–protein kinase A (PKA) pathway is the major signalling pathway employed by GRF and GHRP-2, the activation of PKC may potentiate signalling via the cAMP–PKA pathway in ovine GH secretion. Importantly, the effect of PMA in increasing the secretion of GH from ovine somatotrophs is effected, in part, by up-regulation of the cAMP–PKA pathway. We conclude that there is cross-talk between the PKC pathway and the cAMP–PKA pathway in ovine somatotrophs during the action of GRF or GHRP.

Journal of Endocrinology (1997) 154, 219–230

Free fatty acids (FFAs) regulate insulin secretion in a complex pattern and induce pancreatic β-cell dysfunction in type 2 diabetes. Voltage-dependent Ca^{2 +} channels (VDCC) in β-cells play a major role in regulating insulin secretion. The aim of present study is to clarify the action of the FFA, linoleic acid, on VDCC in β-cells. The VDCC current in primary cultured rat β-cells were recorded under nystatin-perforated whole-cell recording configuration. The VDCC was identified as high-voltage-gated Ca^{2 +} channels due to there being no difference in current amplitude under holding potential between −70 and −40 mV. Linoleic acid (10 μM) significantly inhibited VDCC currents in β-cells, an effect which was fully reversible upon washout. Methyl-linoleic acid, which does not activate G protein coupled receptor (GPR)40, neither did alter VDCC current in rat β-cells nor did influence linoleic acid-induced inhibition of VDCC currents. Linoleic acid-induced inhibition of VDCC current was not blocked by preincubation of β-cells with either the specific protein kinase A (PKA) inhibitor, H89, or the PKC inhibitor, chelerythrine. However, pretreatment of β-cells with thapsigargin, which depletes intracellular Ca²⁺ stores, completely abolished linoleic acid-induced decrease in VDCC current. Measurement of intracellular Ca²⁺ concentration ([Ca²⁺]_i) illustrated that linoleic acid induced an increase in [Ca²⁺]_i and that thapsigargin pretreatment inhibited this increase. Methyl-linoleic acid neither did induce increase in [Ca²⁺]_i nor did it block linoleic acid-induced increase in [Ca²⁺]_i. These results suggest that linoleic acid stimulates Ca²⁺ release from intracellular Ca²⁺ stores and inhibits VDCC currents in rat pancreatic β-cells via Ca²⁺-induced inactivation of VDCC.

ABSTRACT

Effects of age and naloxone on testicular function were studied in golden hamsters exposed to short photoperiods. Subjection of hamsters to short photoperiods of 6 h light: 18 h darkness for 6 weeks led to testicular regression in young adult (5–6 months) or middle-aged (11–12 months) golden hamsters but not in prepubertal hamsters of 1–2 months of age. The middle-aged hamsters had decreased testis width by week 4 of treatment and the young hamsters by week 5. Daily injection of naloxone at the time of'lights on' partially prevented testicular regression in young and middle-aged hamsters but the extent of regression was significantly greater in the middle-aged animals.

Plasma LH and FSH concentrations were significantly reduced in hamsters placed in short photoperiods regardless of age or testicular weight, while naloxone treatment significantly increased the LH concentrations in all age groups. Plasma β-endorphin-like immunoreactivity was significantly increased by short photoperiod or older age.

These results indicated that (a) the sensitivity of the testicular suppression to short photoperiod increases as a function of age, (b) naloxone, a specific opiate receptor blocker, can partially prevent short photoperiod-induced testicular regression and (c) ageing and short photoperiods increase β-endorphin-like immunoreactivity. It is concluded that the opiate system may be involved in ageing and photoperiod regulation of reproductive function.

J. Endocr. (1984) 101, 1–6