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ABSTRACT

Melatonin-binding sites have previously been identified in the suprachiasmatic nucleus (SCN) and median eminence (ME) of the rat. We have further investigated the localization of melatonin-binding sites in the rat hypothalamus and pituitary using the ligand [¹²⁵I] iodomelatonin and in-vitro autoradiography. The presence of specific melatonin-binding sites in the SCN is confirmed; however the second area of melatonin binding is identified as the pars tuberalis of the pituitary and not the ME as previously described. No other areas which bound melatonin were found in either the pituitary or the hypothalamus.

α-Melanotrophin was detected by radioimmunoassay in the pituitary glands of fetal rats from day 17 of gestation. The pituitary content of α-melanotrophin increased regularly, at a gradually decreasing rate, throughout gestation and in the postnatal period. Concentrations of α-melanotrophin in the plasma of fetal and newborn rats were below the detection limit of the radioimmunoassay (10 pmol/l). Detectable concentrations were first found in young rats on day 3 after birth and did not differ significantly from those in their mothers throughout the period of suckling. Plasma concentrations of α-melanotrophin were raised in pregnant rats during the last 4 days of gestation and after parturition. They returned to basal levels in the 2 weeks after delivery. After weaning at 3 weeks of age, a large increase in the plasma concentration of α-melanotrophin was detected in juvenile rats. Plasma levels had returned to the normal adult range by 6 weeks of age. The increases in α-melanotrophin in the blood were thought to be the result of non-specific stress effects. The data did not provide evidence for a role for α-melanotrophin in reproductive processes in the rat.

Radioimmunoassay measurements of α-melanotrophin in plasma have identified a diurnal rhythm in male rats. Animals maintained on a 12 h light: 12 h darkness photoperiod had raised levels of plasma α-melanotrophin during the dark phase. Time-series analysis gave a fitted mean level of α-melanotrophin of 52·4 pmol/l, an amplitude of 12·1 pmol/l and peak levels 2·2 h before dawn.

Measurements throughout the oestrous cycle in female rats showed that similar variations between the dark and light phases occurred on the 2 days of dioestrus. The raised levels during the dark period were, however, absent on the nights of pro-oestrus and oestrus. During this pro-oestrous/oestrous period, plasma α-melanotrophin levels were below average but higher than the normal minimum levels found during the light period.

ABSTRACT

The effect of gastric inhibitory polypeptide (GIP), glucagon-like peptide-1(7–36) amide, (GLP-1(7–36) amide), glucagon-like peptide-2 (GLP-2), glucagon and insulin on fatty acid synthesis in explants of rat adipose tissue from various sites was investigated. GIP, GLP-1(7–36) amide and insulin stimulated fatty acid synthesis, as determined by measuring the incorporation of [¹⁴C]acetate into saponifiable fat, in a dose-dependent manner, over the concentration range 5–15 ng/ml (0·87–2·61 nmol/l) for insulin and 0·5–7·5 ng/ml for GIP (0·10–1·50 nmol/l) and GLP-1(7–36) amide (0·15–2·27 nmol/l). Insulin and GIP caused a significantly greater stimulation of [¹⁴C]acetate incorporation into fatty acids in omental adipose tissue than in either epididymal or subcutaneous adipose tissue. Both GIP and GLP-1(7–36) amide had the ability to stimulate fatty acid synthesis within the physiological range of the circulating hormones. At lower concentrations of the hormones, GLP-1(7–36) amide was a more potent stimulator of fatty acid synthesis than GIP in omental adipose tissue culture; the basal rate of fatty acid synthesis was 0·41±0·03 pmol acetate incorporated/mg wet weight tissue per 2 h; at 0·10 nmol hormone/l 1·15±0·10 and 3·40±0·12 pmol acetate incorporated/mg wet weight tissue per 2 h for GIP and GLP-1(7–36) amide respectively (P < 0·01). GLP-2 and glucagon were without effect on fatty acid synthesis in omental adipose tissue. The study indicates that GIP and GLP-1(7–36) amide, in addition to stimulating insulin secretion, may play a direct physiological role in vivo, in common with insulin, in promoting fatty acid synthesis in adipose tissue.

Journal of Endocrinology (1991) 130, 267–272

Abstract

To define the hormonal influences that are directly involved in the hair follicle cycles of animals with differing patterns of fibre growth and moulting, we have investigated the possible presence of IGF-I and melatonin receptors on the dermis and hair follicles of cashmere and Angora goats, sampled in February, March and June, using quantitative in vitro autoradiography. The presence of IGF-I receptors in the dermis of both breeds of goat was determined using cryostat sections incubated with 50 pm ¹²⁵I-labelled IGF-I in the presence or absence of 50 nm IGF-I. Sections of the growing tip of deer antlers containing the cartilaginous zone, a tissue known to contain high concentrations of specific IGF-I receptors, were used as a positive control. As the production of antler velvet uniquely involves the generation of hair follicles de novo, the presence of IGF-I receptors in the velvet-producing region was also investigated. In both breeds of goat, specific ¹²⁵I-IGF-I binding was localised over the inner and outer root sheath, the matrix, the germinal matrix, the dermal papilla and the sebaceous glands and satisfied the basic kinetic criteria considered to be representative of a specific IGF-I receptor. Analysis of saturation isotherms using a one-site binding model revealed dissociation constants (K _d) in the range 0·1–0·9 nm and theoretical maximal numbers of binding sites (B _max) between 21·4 and 45·6 fmol/mg tissue. K _d and B _max values derived from cashmere and Angora goats sampled at different times of the year did not differ significantly between breeds or sampling times. Specific ¹²⁵I-IGF-I binding was also localised to the developing follicles on the deer antler dermis. The presence of melatonin receptors within the goat dermis was also investigated. Sections were incubated with 100 pm 2-[¹²⁵I]iodomelatonin with or without 0·1 μm melatonin, along with sections of sheep pars tuberalis which are known to contain high levels of high-affinity melatonin receptors. No displaceable 2-[¹²⁵I]iodomelatonin binding was found on any sections of the cashmere or Angora skin analysed. It is therefore concluded that melatonin receptors are not present on the hair follicles or associated structures. IGF-I receptors are present on the hair follicle and sebaceous gland and may be involved in the growth of both seasonally and non-seasonally produced fibre and in the development of antler velvet.

Journal of Endocrinology (1996) 151, 55–63

ABSTRACT

The receptors for TSH have been studied in human thyroid tissue to assess their density and binding characteristics in various disease states. A single set of similar independent receptors appeared to be present in both healthy and pathological thyroid tissue. Their binding affinity for bovine TSH averaged 1·1 × 10¹⁰l/mol in healthy tissue and, with the exception of papillary carcinoma which showed some reduction, was not significantly altered in the various disease states studied. No receptors with low binding affinity were found. The number of receptors was significantly greater in toxic diffuse goitre and in hyperfunctioning follicular adenoma (but these tissues came from patients given antithyroid drugs and often iodine preoperatively), and was reduced in Hashimoto's thyroiditis. In well-differentiated thyroid carcinoma, the number of receptors was similar to or greater than in normal tissue, but in undifferentiated and medullary carcinoma, and in lymphoma of the thyroid, receptors were completely absent. Tracer-binding data obtained with human TSH were uniformly lower than the corresponding data obtained with bovine TSH, but showed an analogous pattern of differences amongst the various normal and pathological tissues.

J. Endocr. (1984) 102, 369–374

Abstract

The pars tuberalis (PT) of the anterior pituitary is characterized by the presence of a high concentration of melatonin receptors, and acute exposure of cells from this tissue to melatonin inhibits the accumulation of cyclic AMP (cAMP) stimulated by forskolin. Conversely, exposure of ovine PT (oPT) cells to melatonin for periods of up to 16 h causes a progressive increase in subsequent basal and forskolin-stimulated production of cAMP. These observations are consistent with the possibility that the PT is involved in the mediation of melatonin-dependent phenomena in mammals. If the chronic effects of exposure to melatonin are indeed functionally significant, then one would anticipate that those responses of oPT cells known to be dependent upon levels of cAMP would also show an enhanced response to stimulation following prolonged exposure to the hormone. In the present study, the activation of cAMP-dependent protein kinase and the synthesis of secretory protein by oPT cells were found to be sensitized by prolonged exposure to physiological concentrations of melatonin. In the case of the synthesis of secretory protein this effect of melatonin was confined to those proteins whose synthesis has been shown to be sensitive to melatonin in acute experiments. These observations support the hypothesis that melatonin-induced sensitization modulates the putative biosynthetic and secretory function of the PT.

The present study also examined the mechanism of sensitization of oPT cells by melatonin. The development of sensitization was not affected by simultaneous exposure of oPT cells to forskolin (1 μm) during pretreatment with melatonin. This observation suggests that melatonin-induced sensitization occurs independently of the established acute effects of the hormone on cAMP levels in oPT cells. Since no effects of melatonin upon any other signalling cascade have been observed in these cells, the most plausible explanation for this finding is that sensitization is a direct consequence of prolonged activation of melatonin receptors. Such a mechanism might be linked to the partial down-regulation of melatonin receptors known to occur in oPT cells in response to prolonged exposure to the hormone. In order to test this hypothesis further, the process of recovery from the sensitizing effects of melatonin was examined. The recovery of oPT cells from the sensitizing effects of exposure to melatonin (100 pm, 16 h) took place gradually and, even after an interval of 16 h, cells that had previously been exposed to melatonin for 16 h remained sensitized to approximately 20% of the extent seen immediately following pretreatment with melatonin for 16 h. In contrast to the previously reported insensitivity of the development of sensitization to the protein synthesis inhibitor, cycloheximide, the recovery of oPT cells from melatonin-induced sensitization was completely blocked by cycloheximide (10 μg/ml). Taken together, these observations are consistent with the hypothesis that melatonin-induced sensitization of oPT cells is the result of a reduction in levels of certain as yet unidentified protein(s), involved in the tonic inhibition of adenylate cyclase activity, occurring in parallel with the down-regulation of melatonin receptors, and that, conversely, the resynthesis of these factor(s) is a prerequisite for the return of oPT cells to the desensitized condition.

Journal of Endocrinology (1994) 142, 127–138