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ABSTRACT
A sensitive, specific, reproducible and practical radioimmunoassay for the determination of 5-methoxytryptophol (ML) in pineal glands of different species has been developed. High-affinity specific antisera were produced by immunization of sheep with ML–bovine serum albumin. Iodinated ML, used as the radiolabel, was synthesized by direct iodination of ML using 1,3,4,6,-tetrachloro-3,6-diphenylglycouril as the oxidant. Sensitivity of the assay was 0·005 pmol/tube. The validity of the assay was checked using classical techniques. Cross-reactivity with other indoles was negligible. Parallel inhibition curves were obtained for rat, hamster, sheep and tortoise pineal homogenates. Using thin-layer chromatography, tortoise pineal immunoreactivity also co-chromatographed with standard ML. Samples (n = 5) with ML concentrations of 0·013, 0·052 and 0·209 pmol/tube had intra-assay coefficients of variation of 9·8, 5·7 and 7·6% respectively. Their respective interassay coefficients of variation were 17·7 16·5 and 11·4% (n = 8). The pineal concentration of ML was found to be species dependent. Afternoon ML levels were 0·052± 0·002 (s.e.m.) pmol/gland in the rat (n = 16), 0·539 ±0·089 pmol/gland in the hamster (n = 16), 1·73±0·225 nmol/g in the sheep (n = 10) and 7·15± 0·465 pmol/gland in the tortoise (n = 4). The ratio of ML:melatonin content in the pineal gland also showed a large interspecies variation with values of 0·02 in the rat, 0·22 in the sheep, 2·7 in the hamster and 17 in the tortoise.
J. Endocr. (1986) 110, 177–184
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The circadian rhythms of many night-shift workers are maladapted to their imposed behavioural schedule, and this factor may be implicated in the increased occurrence of cardiovascular disease (CVD) reported in shift workers. One way in which CVD risk could be mediated is through inappropriate hormonal and metabolic responses to meals. This study investigated the responses to standard meals at different circadian times in a group of night-shift workers on a British Antarctic Survey station at Halley Bay (75 degrees S) in Antarctica. Twelve healthy subjects (ten men and two women) were recruited. Their postprandial hormone and metabolic responses to an identical mixed test meal of 3330 kJ were measured on three occasions: (i) during daytime on a normal working day, (ii) during night-time at the beginning of a period of night-shift work, and (iii) during the daytime on return from night working to daytime working. Venous blood was taken for 9 h after the meal for the measurement of glucose, insulin, triacylglycerol (TAG) and non-esterified fatty acids. Urine was collected 4-hourly (longer during sleep) on each test day for assessment of the circadian phase via 6-sulphatoxymelatonin (aMT6s) assay. During normal daytime working, aMT6s acrophase was delayed (7.7+/-1.0 h (s.e.m.)) compared with that previously found in temperate zones in a comparable age-group. During the night shift a further delay was evident (11.8+/-1.9 h) and subjects' acrophases remained delayed 2 days after return to daytime working (12.4+/-1.8 h). Integrated postprandial glucose, insulin and TAG responses were significantly elevated during the night shift compared with normal daytime working. Two days after their return to daytime working, subjects' postprandial glucose and insulin responses had returned to pre-shift levels; however, integrated TAG levels remained significantly elevated. These results are very similar to those previously found in simulated night-shift conditions; it is the first time such changes have been reported in real shift workers in field conditions. They provide evidence that the abnormal metabolic responses to meals taken at night during unadapted night shifts are due, at least in part, to a relative insulin resistance, which could contribute to the documented cardiovascular morbidity associated with shift work. When applied to the 20% of the UK workforce currently employed on shift work, these findings have major significance from an occupational health perspective.
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ABSTRACT
Breeding activity was similarly advanced in ewes given continuous (s.c. implant) or timed (oral dose at 15.30 h) melatonin treatments or subjected to a short (8 h light: 16 h darkness) artificial photoperiod. Treatments commenced in mid-June and were terminated in mid-November. Weekly and serial blood samples were collected before and after treatments commenced, to ascertain the effects on plasma prolactin, LH and FSH concentrations. In addition, serial blood samples were collected for 24 h plasma prolactin and melatonin estimations before and after cessation of the treatments.
Plasma prolactin levels were significantly reduced immediately following the start of the melatonin (implant and oral) and short-photoperiod treatments but 'rebounded' to levels greater than control values. The normal seasonal (spring) rise in plasma prolactin was noted in the following year. Before the onset of breeding activity, mean plasma LH and FSH concentrations and LH pulse frequency did not change following any of the treatments. The 24-h plasma melatonin profile accurately reflected the various applied treatments but had re-entrained to the prevailing (natural) photoperiod 1 week after termination of the treatments. There were no significant group differences in 24-h plasma prolactin levels 1 week before or 1 and 11 weeks after the treatments had ceased.
Such treatments, although successfully advancing the onset of breeding activity and modifying the seasonal plasma prolactin rhythm, were not manifested through any apparent change in peripheral LH or FSH.
J. Endocr. (1987) 112, 103–111
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ABSTRACT
The effect of melatonin on the stimulated release of LH from prepubertal female rat pituitary cells in vitro was investigated. Significant inhibition of LH-releasing hormone and calcium ionophore-induced LH release was seen but not of potassium-induced release. These results suggest a specific interaction between melatonin and the endogenous events leading to LH release, and may implicate melatonin as an important neuroendocrine component of pubertal development in this species.
J. Endocr. (1985) 107, 107–112
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ABSTRACT
Ewes were treated with s.c. implants of melatonin in mid-April, mid-May and mid-June. From mid-June, other animals were given oral doses of melatonin daily at 16.30 h and another group was maintained under a short (8 h light: 16 h darkness) artificial photoperiod (lights out 16.30 h). Serial blood samples were taken from all animals in June and July. Plasma prolactin concentrations were significantly reduced in ewes treated in May and June (implant, oral and photoperiod treatments) but not in those treated in April. After treatment in June, prolactin levels were significantly suppressed after 7 days of oral and implant melatonin therapy, and after 28 days of a short artificial photoperiod. Melatonin treatment appeared more efficient than an artificial photoperiod in reducing plasma prolactin concentrations.
J. Endocr. (1986) 108, 287–292
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The circadian rhythms of most night shift workers do not adapt fully to the imposed behavioural schedule, and this factor is considered to be responsible for many of the reported health problems. One way in which such disturbances might be mediated is through inappropriate hormonal and metabolic responses to meals, on the night shift. Twelve healthy subjects (four males and eight females) were studied on three occasions at the same clock time (1330 h), but at different body clock times, after consuming test meals, first in their normal environment, secondly after a forced 9 h phase advance (body clock time approximately 2230 h) and then again 2 days later in the normal environment. They were given a low-fat pre-meal at 0800 h, then a test meal at 1330 h with blood sampling for the following 9 h. Parameters measured included plasma glucose, non-esterified fatty acids (NEFAs), triacylglycerol (TAG), insulin, C-peptide, proinsulin and glucose-dependent insulinotropic polypeptide, and urinary 6-sulphatoxymelatonin. In contrast with a previous study with a high-fat pre-meal, postprandial glucose and insulin responses were not affected by the phase shift. However, basal plasma NEFAs were lower immediately after the phase shift (P < 0.05). Incremental (difference from basal) TAG responses were significantly higher (P < 0.05) immediately after the phase shift compared with before. Two-day post-phase shift responses showed partial reversion to baseline values. This study suggests that it takes at least 2 days to adapt to eating meals on a simulated night shift, and that the nutritional content of the pre-meals consumed can have a marked effect on postprandial responses during a simulated phase shift. Such findings may provide a partial explanation for the increased occurrence of cardiovascular disease reported in shift workers.
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ABSTRACT
Specific, sensitive and direct radioimmunoassays have been used to determine the daily patterns of 5-methoxytryptophol (ML) and melatonin in the pineal glands of Syrian hamsters kept in different photo-periods: 8 h light: 16 h darkness (8L:16D), 14L: 10D and 16L: 8D. A rhythm in pineal ML was evident in animals in all the photoperiods, with high daytime levels (641±35 (s.e.m.) fmol/gland; n=162) which dropped to 119±16 fmol/gland (n = 44) 7·1– 7·5 h after lights out. The duration of low night-time ML levels was proportional to the length of the dark phase (1·2 h in 16L:8D, 5·4 h in 14L: 10D and 8·4 h in 8L: 16D). A marked daily rhythm in melatonin was also present in hamsters in the different photoperiods, with daytime levels of 323 ± 34 fmol/gland (n = 129) and night-time peak concentrations of 3676 ± 336 fmol/gland (n = 22). The duration of high nocturnal melatonin levels was dependent upon the length of the dark phase (4·1 h in 16L: 8D, 4·5 h in 14L: 10D and 12·5 h in 8L: 16D). Linear regression analysis revealed a statistically significant inverse relationship between pineal ML and melatonin levels in 8L: 16D (P< 0·001), 14L: 10D normal (P<0·05) and 14L: 10D shifted (P<0·001) photoperiods. After advancing the lighting schedule by 10 h (14L: 10D, lights off at 04.00 h), pineal ML and melatonin rhythms became entrained to the new lighting regimen.
The daily rhythms in pineal ML and melatonin in the Syrian hamster thus depend on the prevailing photoperiod, a reciprocal relationship existing between pineal ML and melatonin concentrations.
J. Endocr. (1987) 114, 301–309
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Although melatonin treatment has been shown to phase shift human circadian rhythms, it still remains ambiguous as to whether exogenous melatonin can entrain a free-running circadian system. We have studied seven blind male subjects with no light perception who exhibited free-running urinary 6-sulphatoxymelatonin (aMT6s) and cortisol rhythms. In a single-blind design, five subjects received placebo or 5 mg melatonin p.o. daily at 2100 h for a full circadian cycle (35-71 days). The remaining two subjects also received melatonin (35-62 days) but not placebo. Urinary aMT6s and cortisol (n=7) and core body temperature (n=1) were used as phase markers to assess the effects of melatonin on the During melatonin treatment, four of the seven free-running subjects exhibited a shortening of their cortisol circadian period (tau). Three of these had taus which were statistically indistinguishable from entrainment. In contrast, the remaining three subjects continued to free-run during the melatonin treatment at a similar tau as prior to and following treatment. The efficacy of melatonin to entrain the free-running cortisol rhythms appeared to be dependent on the circadian phase at which the melatonin treatment commenced. These results show for the first time that daily melatonin administration can entrain free-running circadian rhythms in some blind subjects assessed using reliable physiological markers of the circadian system.
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Patterns of plasma melatonin, similar in the duration of high levels to those found in winter, were induced in Suffolk-cross ewes kept in summer light (16 h light: 8 h darkness) by daily oral administration of melatonin (3 mg/13 μmol). The onset of oestrous cycles in these sheep occurred in August, 2–8 weeks before the onset of oestrous cycles in untreated ewes kept in natural light. The onset of oestrous cycles in a further group of ewes kept in winter light (8 h light: 16 h darkness) from mid-June was indistinguishable from that of the melatonin-treated ewes. Rams were excluded from the premises. These data indicate that melatonin alone in physiological quantities is sufficient to induce early onset of the breeding season in the ewe, and provide strong evidence for a hormonal role of melatonin in a short-day breeder.
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This study was undertaken to determine whether the internal clock contributes to the hormone and metabolic responses following food, in an experiment designed to dissociate internal clock effects from other factors. Nine female subjects participated. They lived indoors for 31 days with normal time cues, including the natural light: darkness cycle. For 7 days they retired to bed from 0000 h to 0800 h. They then underwent a 26-h 'constant routine' (CR) starting at 0800 h, being seated awake in dim light with hourly 88 Kcal drinks. They then lived on an imposed 27-h day (18 h of wakefulness, 9 h allowed for sleep), for a total of 27 days. A second 26-h CR, starting at 2200 h, was completed. During each CR salivary melatonin and plasma glucose, triacylglycerol (TAG), non-essential fatty acids (NEFA), insulin, gastric inhibitory peptide (GIP) and glucagon-like peptide-1 (GLP-1) were measured hourly. Melatonin and body temperature data indicated no shift in the endogenous clock during the 27-h imposed schedule. Postprandial NEFA, GIP and GLP-1 showed no consistent effects. Glucose, TAG and insulin increased during the night in the first CR. There was a significant effect of both the endogenous clock and sleep for glucose and TAG, but not for insulin. These findings may be relevant to the known increased risk of cardiovascular disease amongst shift workers.