This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Johnston, J. D. Articles by Hazlerigg, D. G. Search for Related Content PubMed PubMed Citation Articles by Johnston, J. D. Articles by Hazlerigg, D. G.

Multiple Effects of Melatonin on Rhythmic Clock Gene Expression in the Mammalian Pars Tuberalis

School of Biological Sciences (J.D.J, D.G.H.), University of Aberdeen, Zoology Building, Aberdeen AB24 2TZ, Scotland, United Kingdom; Neurobiologie des Rhythmes laboratory, Unité Mixte de Recherche 7518 Centre National de la Recherche Scientifique/Université Louis Pasteur (B.B.T., M.M.-P.),67084 Strasbourg, France; and Medical Research Council Human Reproductive Sciences Unit (H.A., G.A.L.), Centre for Reproductive Biology, The Queen’s Medical Research Institute, Edinburgh EH16 4TJ, Scotland, United Kingdom

Address all correspondence and requests for reprints to: David Hazlerigg, School of Biological Sciences, University of Aberdeen, Zoology Building, Tillydrone Avenue, Aberdeen AB24 2TZ, Scotland, United Kingdom. E-mail: d.hazlerigg{at}abdn.ac.uk.

	Abstract

Top Abstract Introduction Materials and Methods Results Discussion References

 
In mammals, changing day length modulates endocrine rhythms via nocturnal melatonin secretion. Studies of the pituitary pars tuberalis (PT) suggest that melatonin-regulated clock gene expression is critical to this process. Here, we considered whether clock gene rhythms continue in the PT in the absence of melatonin and whether the effects of melatonin on the expression of these genes are temporally gated. Soay sheep acclimated to long photoperiod (LP) were transferred to constant light for 24 h, suppressing endogenous melatonin secretion. Animals were infused with melatonin at 4-h intervals across the final 24 h, and killed 3 h after infusion. The expression of five clock genes (Per1, Per2, Cry1, Rev-erb, and Bmal1) was measured by in situ hybridization. In sham-treated animals, PT expression of Per1, Per2, and Rev-erb showed pronounced temporal variation despite the absence of melatonin, with peak times occurring earlier than predicted under LP. The time of peak Bmal1 expression remained LP-like, whereas Cry1 expression was continually low. Melatonin infusion induced Cry1 expression at all times and suppressed other genes, but only when they showed high expression in sham-treated animals. Hence, 3 h after melatonin treatment, clock gene profiles were driven to a similar state, irrespective of infusion time. In contrast to the PT, melatonin infusions had no clear effect on clock gene expression in the suprachiasmatic nuclei. Our results provide the first example of acute sensitivity of multiple clock genes to one endocrine stimulus and suggest that rising melatonin levels may reset circadian rhythms in the PT, independently of previous phase.

	Introduction

Top Abstract Introduction Materials and Methods Results Discussion References

 
MELATONIN, THE PRINCIPAL product of the vertebrate pineal gland, undergoes marked temporal variation in secretion and drives a wide range of rhythmic physiology. In mammals, the synthesis and secretion of pineal melatonin occurs at night, under the control of the circadian clock present in the suprachiasmatic nuclei (SCN) of the hypothalamus. The duration of this nocturnal signal varies in proportion to the length of the night, and thence melatonin secretion transduces both daily and seasonal time throughout the body (1).

The pituitary pars tuberalis (PT) is strongly implicated in the photoperiodic regulation of prolactin secretion and, because of its high density of melatonin receptors, has become a key model tissue for the understanding of melatonin signal readout (2, 3, 4). It is now clear that melatonin is a key regulator of clock gene expression in the mammalian PT, because modulation of the endogenous profile of melatonin secretion, by manipulation of ambient photoperiod or by administration of exogenous melatonin, changes PT clock gene profiles (5, 6, 7, 8, 9).

To date, the documented effects of melatonin in the PT can be divided into those occurring in the morning (Per1) and those occurring in the evening (Cry1, Rev-rb). The former depend on melatonin withdrawal at the end of the night, probably mediated by the resultant increase in cAMP production (2, 5). In rodents unable to synthesize melatonin, either because of genetic deficiency (10) or pinealectomy (11), rhythmic expression of Per1 expression is absent, suggesting that melatonin is necessary for maintained rhythmic gene expression in this tissue.

Collectively, these observations suggest that the PT contains a circadian oscillator that is either driven or modulated by melatonin and raise questions about 1) whether a rhythmic melatonin signal is necessary for the 24-h variation of clock gene expression in the PT and 2) whether melatonin actions on gene expression in the PT is restricted to time windows around the morning and evening changes in illumination. To test these ideas, we returned to the sheep as a model because this species has been extensively used for investigations into the molecular basis of melatonin-dependent photoperiodic responses (2, 6, 12). We placed animals in constant light for one 24-h cycle (thereby suppressing endogenous melatonin secretion) and treated them at 4-h intervals with either melatonin-containing sc implants or sham procedure.

	Materials and Methods

Top Abstract Introduction Materials and Methods Results Discussion References

 
Animals and housing
All animals were treated in accordance with the U.K. Animals (Scientific Procedures) Act, 1986. Forty-eight pubertal female Soay sheep, obtained from specialist breeders in Scotland (12), were brought indoors in winter (November). They were housed in light-sealed rooms and fed a standardized diet of grass pellets (500 g per animal; Vitagrass, Cumbria, UK) given daily 1 h into the light phase. Hay and water were available ad libitum. White fluorescent strip lights provided approximately 160 lux at the animals’ eye level during the light phase, whereas dim red light (<5 lux) was provided during the dark phase.

Light and melatonin treatment
The animals were initially acclimatized under a winter-like short photoperiod (SP; 8 h light, 16 h darkness) for 4 wk and then exposed to long photoperiod (LP; 16 h light, 8 h dark) for 6 wk. Blood samples were collected twice weekly from the jugular vein from representative animals (n = 12), and the blood plasma separated by centrifugation within 30 min and stored at –20 C, to measure prolactin concentration as evidence of photoresponsiveness.

Over the final 24 h of the experiment, lights were left on to suppress endogenous pineal melatonin secretion. Animals were treated with a single sc implant of melatonin (Regulin; CEVA Animal Health, Chesham, Bucks, UK) placed on the inside of the hind leg using a standard injector, or a sham treatment with no implant as control at 4-h intervals across the final 24 h (n = 4 per time point), and killed 3 h after treatment by pentobarbital injection. Immediately before death, blood samples were collected from the jugular vein and processed as above to permit analysis of the effect of treatments on melatonin concentration. The hypothalamus and upper pituitary gland were removed within 10 min of death and snap frozen in isopentane kept at –30 C by dry ice. The expression of five clock genes (Per1, Per2, Cry1, Rev-erb, and Bmal1) was measured in the PT and SCN by in situ hybridization.

RIA
Prolactin concentrations were measured by a validated RIA (13). The lower limit of sensitivity was 0.5 ng/ml prolactin standard (NIH-PRL-S13), and the intraassay coefficient of variation was 8.3%. Melatonin concentrations were also measured by a standard RIA (14), using a commercial antibody (PF-1288; SPI-BIO, Paris, France). The sensitivity of the assay was 5 pg/ml, and the intraassay coefficient of variation was 12.0%.

In situ hybridization
Coronal cryosections (20 µm) cut through the SCN and PT were thaw-mounted onto poly-L-lysine-coated glass slides and stored at –80 C. Expression of the clock genes Per1, Per2, Cry1, Bmal1, and Rev-erb was then detected using radioactive riboprobes and procedures described previously (15).

Statistics
Weekly changes in plasma prolactin concentration during acclimation to LP were analyzed by one-way ANOVA with repeated measures. Clock gene expression profiles in sham-treated animals were analyzed by one-way ANOVA. The comparison between melatonin and sham treatment was analyzed by two-way ANOVA and Bonferroni’s post hoc testing when appropriate.

	Results

Top Abstract Introduction Materials and Methods Results Discussion References

 
Photosensitivity of experimental animals
The pretreatment exposure to LP induced a significant (P < 0.01, one-way ANOVA with repeated measures) increase in plasma prolactin concentrations from 19.2 ± 4.6, at the change from SP to LP, to 52.4 ± 11.1 ng/ml at wk 6, illustrating the physiological and photosensitive state of the experimental animals before the final day on constant light (LL) (Fig. 1).

View larger version (13K): [in this window] [in a new window]   FIG. 1. Photoresponsiveness of experimental animals. Female Soay sheep were brought indoors into controlled lighting conditions in November. They were initially exposed to a SP (8 h light, 16 h dark) for 4 wk and then transferred to LP (16 h light, 8 h dark) for 6 wk. Over this final 6-wk period, blood samples were taken from a subset (n = 12) of animals to monitor plasma prolactin concentration. Exposure to LP induced an increase in plasma prolactin (P < 0.01, one-way ANOVA with repeated measures), indicating that the animals used in this study were photoresponsive.

View larger version (22K): [in this window] [in a new window]   FIG. 2. Melatonin implants induce physiological plasma melatonin concentrations and acutely inhibit plasma prolactin. Soay sheep acclimated to LP conditions were kept in LL for the final 24 h of the experiment. During this period, animals were given either melatonin infusion or sham treatment every 4 h and killed 3 h after treatment. A, Across the 24-h cycle, plasma melatonin concentration was suppressed in sham-treated animals but raised into the nocturnal physiological range by melatonin implants (P < 0.001 for effect of treatment, two-way ANOVA). B, Treatment with melatonin acutely suppressed plasma prolactin concentration (P < 0.01 for effect of treatment, two-way ANOVA).

Clock gene expression in the PT
Despite the suppressed plasma melatonin concentrations in sham-treated animals, the expression of four of the five clock genes analyzed in the PT exhibited significant (P < 0.05, one-way ANOVA) variation over the 24-h cycle (Fig. 3). For Rev-erb and Per1, which show peak PT expression in the early light phase in LP acclimated animals (6, 8), a melatonin-independent expression peak was seen after approximately 20 h of constant light treatment. For Per2, which peaks slightly later than Per1 under LP (6), a melatonin-independent increase in expression was seen after 24 h of exposure to LL. Similar to the case in LP acclimated animals (6), Bmal1 expression peaked between ZT12 and ZT16 in LL.

View larger version (32K): [in this window] [in a new window]   FIG. 3. Clock gene rhythms persist in the absence of rhythmic melatonin secretion. Soay sheep acclimated to LP conditions were kept in LL for the final 24 h of the experiment. Despite suppressed plasma melatonin concentration over the whole 24-h period, expression of Per1, Per2, Rev-erb, and Bmal1 mRNAs in the PT of sham-treated animals exhibited significant (P < 0.001, one-way ANOVA) variation in expression. Moreover, the relative timing of peak mRNA expression was similar to that reported in other clock tissues. Autoradiographs show representative examples of maximal and minimal expression of each gene. Scale bar, 2 mm.

View larger version (27K): [in this window] [in a new window]   FIG. 4. Melatonin acutely induces Cry1 mRNA expression throughout the 24-h cycle. Soay sheep acclimated to LP conditions were kept in constant light for the final 24 h of the experiment. During this period, animals were given either melatonin infusion or sham treatment every 4 h and killed 3 h after treatment. A, Representative autoradiographs of Cry1 expression in the PT of melatonin- vs. sham-treated animals. Scale bar, 5 mm. B, Quantified expression of Cry1 in the PT of animals treated with sham vs. melatonin across the 24-h cycle. Melatonin significantly increased Cry1 mRNA expression independently of infusion time (P < 0.001 for effect of treatment, two-way ANOVA; *, P < 0.05, sham vs. melatonin, Bonferroni’s post hoc test). Values are mean ± SEM of n = 4 animals per group.

View larger version (28K): [in this window] [in a new window]   FIG. 5. Melatonin acutely inhibits the endogenous elevation of expression of multiple clock genes. Soay sheep acclimated to LP conditions were kept in constant light for the final 24 h of the experiment. During this period, animals were given either melatonin infusion or sham treatment every 4 h and killed 3 h after treatment. Data show quantified expression of Per1, Per2, Rev-erb, and Bmal1 in the PT of animals treated with sham vs. melatonin across the 24-h cycle. Melatonin significantly inhibited gene expression (P < 0.05 for effect of treatment for Bmal1; P < 0.001 for effect of treatment for other clock genes, two-way ANOVA; *, P < 0.05, sham vs. melatonin, Bonferroni’s post hoc test). Values are mean ± SEM of n = 4 animals per group.

View larger version (18K): [in this window] [in a new window]   FIG. 6. Melatonin does not acutely regulate expression of clock genes in the ovine SCN. Soay sheep acclimated to LP conditions were kept in constant light for the final 24 h of the experiment. During this period, animals were given either melatonin infusion or sham treatment every 4 h and killed 3 h after treatment. Data show quantified expression of Per1, Per2, Cry1, Rev-erb, and Bmal1 in the SCN of animals treated with sham vs. melatonin across the 24-h cycle. For Rev-erb (P = 0.05), but not other clock genes (P > 0.05 for effect of treatment, two-way ANOVA), there was a significant effect of melatonin infusion on mRNA expression. Values are mean ± SEM of n = 4 animals per group.

	Discussion

Top Abstract Introduction Materials and Methods Results Discussion References

Since the classical lesioning experiments performed over 30 yr ago (17, 18), the SCN has been viewed as a master circadian pacemaker that drives and synchronizes overt rhythms of physiology and behavior. The recent identification of circadian oscillators in peripheral tissues and in immortalized cell lines suggests that the role of the SCN is primarily to coordinate circadian oscillations throughout the organism (19) and focuses attention on the mechanisms through which such circadian synchronization takes place. The precise manner by which a given SCN-dependent circadian signal influences peripheral oscillator function may be expected to reflect the physiological functions of the tissues upon which it acts. In mammals, melatonin is one such SCN-dependent circadian output, whose primary function is to modulate neuroendocrine physiology, by relaying information about changes in day length (20). The pineal melatonin signal communicates the phase of night onset and of the subsequent dawn; melatonin is secreted continuously between these transitions, giving a signal whose duration is proportional to the length of the night. This ability of the melatonin signal to represent night duration is crucial for photoperiodic responses in seasonal mammals (21).

The PT is a largely homogeneous population of type 1 melatonin (MT₁) receptor-expressing thyrotrophic cells (22, 23), making it an ideal tissue for the study of melatonin action. Long-term loss of melatonin synthesis or loss of functional MT₁ receptors abolishes rhythmic clock gene expression in the mammalian PT (10, 11, 24, 25), indicating that the molecular clockwork of this tissue is ultimately melatonin dependent. The data presented here, however, demonstrate clear temporal variation in PT expression of four core clock genes over a 24-h period in the absence of a rhythmic melatonin signal and thus strongly support the existence of an endogenous PT oscillator. We therefore suspect that arrhythmic Per1 gene expression in the PT of hamsters that have been pinealectomized for 7 d (11) derives from progressive damping of the PT circadian oscillator. Based on recent studies of damping in cell culture systems (26, 27, 28), we speculate that this is a result of desynchronization of circadian oscillators in individual PT cells. Accordingly, in genetically melatonin-synthesis- or melatonin-receptor-deficient mice (10, 24, 25), PT gene expression is likely arrhythmic because PT cells never become synchronized during fetal or postnatal development.

The present study has examined only one aspect of the information inherent in the endogenous melatonin signal: rising melatonin levels associated with night onset. Our data suggest that a dusk-like response (high Cry1, low Per, Rev-erb, and Bmal1) can be elicited by increasing melatonin levels occurring at any phase throughout the 24-h cycle. This suggests that PT cells exhibit a type 0 resetting response (29) to rising melatonin levels, that is to say that the PT molecular clockwork resets to a dusk state in response to rising melatonin, independent of its state immediately beforehand. This dusk state is transient, because our previous data show that a state of low Cry1 and high Bmal1 expression develops during continued melatonin presence through the middle to the end of the night (6). Because the effects of melatonin on cAMP-dependent signal transduction and gene expression persist during prolonged melatonin exposure (2), it is likely that cAMP-independent mechanisms account for disappearance of the dusk state under physiological conditions.

Interestingly, the PT of the Japanese quail also rhythmically expresses clock genes, of which Cry1 may be acutely induced by light. However, photic induction of Cry1 is subject to circadian gating in this tissue (30), suggesting that the avian PT may contain a more robust oscillator than is the case in mammals.

For sheep in a natural environment, the times of dawn and dusk move in opposite directions during the annual cycle, raising the possibility of a similar flexibility in the resetting response of the PT oscillator to declining melatonin levels associated with dawn. Although we have yet to perform an experiment explicitly to address this, our published data agree with this prediction; the timing of the dawn state, characterized by high Per1, can be delayed by an acute light manipulation in the preceding evening that delays the dawn-associated decline in melatonin secretion (8). Hence the circadian clockwork of the sheep PT shows a remarkable plasticity in its response to changing patterns of melatonin secretion.

Early work in the mammalian PT focused on the ability of melatonin to modulate adenylate cyclase activity (2). Melatonin-regulated expression of Per1, which is considered a cAMP-regulated immediate-early response gene in the PT (31), is likely to occur through this pathway (24). However, other clock genes are not directly sensitive to cAMP signaling, suggesting that melatonin-dependent signaling may occur by additional and as yet undefined signaling pathways. Consistent with this possibility, previous studies of many tissues, including the ovine PT, have revealed a wide range of intracellular pathways that are sensitive to melatonin (32). At present, we cannot entirely rule out the possibility that induction of Cry1 drives changes in the expression of other clock genes. However, we feel that this putative mechanism is unlikely, because it would require not only rapid transcription and translation of Cry1 but also a high level of mRNA instability for other clock genes to be affected within the short interval (3 h) between melatonin infusion and tissue collection used in this study.

Additional to its role in seasonal neuroendocrine control, melatonin may influence the function of the SCN itself. Exogenous melatonin administration has been shown to synchronize circadian behavioral rhythmicity in rodents (33, 34) and in humans (35), and melatonin can reportedly phase-shift circadian rhythms of electrical activity in mouse SCN slice preparations (36). Knockout of the MT₁ and MT₂ melatonin receptor subtypes abolishes phase-shifting responses to melatonin in mice and acute effects of melatonin on SCN electrical activity (37, 38). Hence it is possible that circadian effects of melatonin are mediated through melatonin receptors located in the SCN. Nevertheless, a recent study reported no effect of melatonin injection on clock gene expression in the rat SCN (39). In sheep, melatonin receptors are at best weakly expressed in the SCN (40), and in the present study a mild effect of melatonin injection was seen for only one gene, Rev-erb, at one time point only. Possibly our data and that of Poirel et al. (39) reflect different coupling between melatonin receptors and circadian molecular clockwork in SCN compared with PT cells. Alternatively the heterogeneous cellular composition of the SCN and its sensitivity to multiple inputs in addition to melatonin (41) may account for these results.

A final point of interest is the regulation of plasma prolactin concentration in this study. We have previously described a LP-induced increase in prolactin concentration within 24 h (8). In this study, the animals required approximately 4–6 wk of LP exposure before showing such a rise. This difference can be explained by two reasons. In the current study, animals were likely to exhibit early refractoriness to shortening winter photoperiod when they were transferred to controlled lighting regimes; furthermore, animals were not habituated to blood sampling and it is therefore likely that initial prolactin concentrations were affected by stress of experimental manipulation at this stage. During the final 24 h of this experiment, melatonin caused an acute inhibition of prolactin release in the sheep under constant light. This is unlikely to reflect a direct effect on lactotroph cells, which do not appear to express melatonin receptors (22, 42) and are unresponsive to melatonin in primary cell culture (43). It is unclear whether melatonin acts at the level of the PT or hypothalamus to acutely suppress prolactin concentration. Although hypothalamic monoamines are not believed to drive photoperiodic changes of prolactin secretion (44, 45), there is some evidence that melatonin may stimulate secretion of the prolactin-inhibiting factor dopamine from hypothalamic neurons (46, 47). Alternatively, melatonin infusions may have acutely inhibited the secretion of prolactin-releasing factor(s), termed tuberalin, from the PT (43, 48, 49, 50, 51). However, the acute effects of melatonin on tuberalin secretion are as yet unknown.

In summary, we propose that the PT may become an important model tissue to address the function of peripheral circadian oscillators as well as the mechanisms of photoperiodic time measurement. A strength of the PT as a circadian model is that it has a well defined circadian input, melatonin. As well as testing the generality of the results described here in other species, we are currently developing in vitro techniques to further evaluate the effect of melatonin on the PT circadian clockwork. These studies will extend beyond the scope of the current in vivo work and will hopefully provide novel insights into the chronotherapeutic effects of melatonin.

	Acknowledgments

 
We are grateful to Marshall Building staff (Edinburgh) and Nigel Graham and Jean-Michel Fustin (Aberdeen) for technical assistance and to Prof. Ueli Schibler (University of Geneva, Switzerland) for the generous gift of the Rev-erb probe template. Purified preparations of ovine prolactin were provided by the National Hormone and Peptide Program.

	Footnotes

 
Present address for J.D.J.: School of Biomedical and Molecular Sciences, University of Surrey, Guildford GU2 7XH, United Kingdom.

This work was supported by the Medical Research Council (to H.A. and G.A.L.) and a grant from the Biotechnology and Biological Sciences Research Council (to D.G.H.).

First Published Online November 3, 2005

¹ J.D.J. and B.B.T. are joint first authors.

Abbreviations: LL, Constant light; LP, long photoperiod; MT₁, melatonin type 1; PT, pars tuberalis; SCN, suprachiasmatic nuclei; SP, short photoperiod; ZT, zeitgeber time.

Received August 29, 2005.

Accepted for publication October 26, 2005.

	References

Top Abstract Introduction Materials and Methods Results Discussion References

 

1. Simonneaux V, Ribelayga C 2003 Generation of the melatonin endocrine message in mammals: a review of the complex regulation of melatonin synthesis by norepinephrine, peptides, and other pineal transmitters. Pharmacol Rev 55:325–395[Abstract/Free Full Text]
2. Hazlerigg DG, Morgan PJ, Messager S 2001 Decoding photoperiodic time and melatonin in mammals: what can we learn from the pars tuberalis? J Biol Rhythms 16:326–335[Abstract/Free Full Text]
3. Johnston JD 2004 Photoperiodic regulation of prolactin secretion: changes in intra-pituitary signalling and lactotroph heterogeneity. J Endocrinol 180:351–356[Abstract]
4. Kell CA, Stehle JH 2005 Just the two of us: melatonin and adenosine in rodent pituitary function. Ann Med 37:105–120[CrossRef][Medline]
5. Messager S, Ross AW, Barrett P, Morgan PJ 1999 Decoding photoperiodic time through Per1 and ICER gene amplitude. Proc Natl Acad Sci USA 96:9938–9943[Abstract/Free Full Text]
6. Lincoln G, Messager S, Andersson H, Hazlerigg D 2002 Temporal expression of seven clock genes in the suprachiasmatic nucleus and the pars tuberalis of the sheep: evidence for an internal coincidence timer. Proc Natl Acad Sci USA 99:13890–13895[Abstract/Free Full Text]
7. Dardente H, Menet JS, Poirel VJ, Streicher D, Gauer F, Vivien-Roels B, Klosen P, Pevet P, Masson-Pevet M 2003 Melatonin induces Cry1 expression in the pars tuberalis of the rat. Mol Brain Res 114:101–106[Medline]
8. Hazlerigg DG, Andersson H, Johnston JD, Lincoln GA 2004 Molecular characterization of the long-day response in the Soay sheep, a seasonal mammal. Curr Biol 14:334–339[CrossRef][Medline]
9. Johnston JD, Ebling FJP, Hazlerigg DG 2005 Photoperiod regulates multiple gene expression in the suprachiasmatic nuclei and pars tuberalis of the Siberian hamster (Phodopus sungorus). Eur J Neurosci 21:2967–2974[CrossRef][Medline]
10. Sun ZS, Albrecht U, Zhuchenko O, Bailey J, Eichele G, Lee CC 1997 RIGUI, a putative mammalian ortholog of the Drosophila Period gene. Cell 90:1003–1011[CrossRef][Medline]
11. Messager S, Garabette ML, Hastings MH, Hazlerigg DG 2001 Tissue-specific abolition of Per1 expression in the pars tuberalis by pinealectomy in the Syrian hamster. NeuroReport 12:579–582[CrossRef][Medline]
12. Lincoln GA 2002 Neuroendocrine regulation of seasonal gonadotrophin and prolactin rhythms: lessons from the Soay ram model. Reprod Suppl 19:131–147
13. McNeilly AS, Andrews P 1974 Purification and characterization of caprine prolactin. J Endocrinol 60:359–367[Abstract/Free Full Text]
14. Fraser S, Cowan P, Franclin M, Franey C, Arendt J 1983 Direct radioimmunoassay for melatonin in plasma. Clin Chem 29:396–397[Free Full Text]
15. Lincoln GA, Johnston JD, Andersson H, Wagner G, Hazlerigg DG 2005 Photorefractoriness in mammals: dissociating a seasonal timer from the circadian-based photoperiod response. Endocrinology 146:3782–3790[Abstract/Free Full Text]
16. Lincoln GA, Andersson H, Loudon A 2003 Clock genes in calendar cells as the basis of annual timekeeping in mammals: a unifying hypothesis. J Endocrinol 179:1–13[Abstract]
17. Moore RY, Eichler VB 1972 Loss of a circadian adrenal corticosterone rhythm following suprachiasmatic lesions in the rat. Brain Res 42:201–206[CrossRef][Medline]
18. Stephan FK, Zucker I 1972 Circadian rhythms in drinking behavior and locomotor activity of rats are eliminated by hypothalamic lesions. Proc Natl Acad Sci USA 69:1583–1586[Abstract/Free Full Text]
19. Reppert SM, Weaver DR 2002 Coordination of circadian timing in mammals. Nature 418:935–941[CrossRef][Medline]
20. Goldman BD 2001 Mammalian photoperiodic system: formal properties and neuroendocrine mechanisms of photoperiodic time measurement. J Biol Rhythms 16:283–301[Abstract/Free Full Text]
21. Bartness TJ, Powers JB, Hastings MH, Bittman EL, Goldman BD 1993 The timed infusion paradigm for melatonin delivery: what has it taught us about the melatonin signal, its reception, and the photoperiodic control of seasonal responses? J Pineal Res 15:161–190[Medline]
22. Klosen P, Bienvenu C, Demarteau O, Dardente H, Guerrero H, Pevet P, Masson-Pevet M 2002 The mt1 melatonin receptor and RORß receptor are co-localized in specific TSH-immunoreactive cells in the pars tuberalis of the rat pituitary. J Histochem Cytochem 50:1647–1657[Abstract/Free Full Text]
23. Dardente H, Klosen P, Pevet P, Masson-Pevet M 2003 MT1 melatonin receptor mRNA expressing cells in the pars tuberalis of the European hamster: effect of photoperiod. J Neuroendocrinol 15:778–786[Medline]
24. von Gall C, Garabette ML, Kell CA, Frenzel S, Dehghani F, Schumm-Draeger PM, Weaver DR, Korf HW, Hastings MH, Stehle JH 2002 Rhythmic gene expression in pituitary depends on heterologous sensitization by the neurohormone melatonin. Nat Neurosci 5:234–238.[CrossRef][Medline]
25. von Gall C, Weaver DR, Moek J, Jilg A, Stehle JH, Korf HW 2005 Melatonin plays a crucial role in the regulation of rhythmic clock gene expression in the mouse pars tuberalis. Ann NY Acad Sci 1040:508–511[CrossRef][Medline]
26. Nagoshi E, Saini C, Bauer C, Laroche T, Naef F, Schibler U 2004 Circadian gene expression in individual fibroblasts: cell-autonomous and self-sustained oscillators pass time to daughter cells. Cell 119:693–705[Medline]
27. Welsh DK, Yoo SH, Liu AC, Takahashi JS, Kay SA 2004 Bioluminescence imaging of individual fibroblasts reveals persistent, independently phased circadian rhythms of clock gene expression. Curr Biol 14:2289–2295[CrossRef][Medline]
28. Carr AJ, Whitmore D 2005 Imaging of single light-responsive clock cells reveals fluctuating free-running periods. Nat Cell Biol 7:319–321[CrossRef][Medline]
29. Roenneberg T, Daan S, Merrow M 2003 The art of entrainment. J Biol Rhythms 18:183–194[Abstract/Free Full Text]
30. Yasuo S, Watanabe M, Tsukada A, Takagi T, Iigo M, Shimada K, Ebihara S, Yoshimura T 2004 Photoinducible phase-specific light induction of Cry1 gene in the pars tuberalis of Japanese quail. Endocrinology 145:1612–1616[Abstract/Free Full Text]
31. Morgan PJ, Ross AW, Graham ES, Adam C, Messager S, Barrett P 1998 oPer1 is an early response gene under photoperiodic regulation in the ovine pars tuberalis. J Neuroendocrinol 10:319–323[CrossRef][Medline]
32. Vanecek J 1998 Cellular mechanisms of melatonin action. Physiol Rev 78:687–721[Abstract/Free Full Text]
33. Redman J, Armstrong S, Ng KT 1983 Free-running activity rhythms in the rat: entrainment by melatonin. Science 219:1089–1091[Abstract/Free Full Text]
34. Grosse J, Velickovic A, Davis FC 1996 Entrainment of Syrian hamster circadian activity rhythms by neonatal melatonin injections Am J Physiol 270:R533–R540
35. Arendt J, Skene DJ 2005 Melatonin as a chronobiotic. Sleep Med Rev 9:25–39[CrossRef][Medline]
36. Gillette MU, McArthur AJ 1996 Circadian actions of melatonin at the suprachiasmatic nucleus. Behav Brain Res 73:135–139[CrossRef][Medline]
37. Liu C, Weaver DR, Jin X, Shearman LP, Pieschl RL, Gribkoff VK, Reppert SM 1997 Molecular dissection of two distinct actions of melatonin on the suprachiasmatic circadian clock. Neuron 19:91–102[CrossRef][Medline]
38. Jin X, von Gall C, Pieschl RL, Gribkoff VK, Stehle JH, Reppert SM, Weaver DR 2003 Targeted disruption of the mouse Mel(1b) melatonin receptor. Mol Cell Biol 23:1054–1060[Abstract/Free Full Text]
39. Poirel VJ, Boggio V, Dardente H, Pevet P, Masson-Pevet M, Gauer F 2003 Contrary to other non-photic cues, acute melatonin injection does not induce immediate changes of clock gene mRNA expression in the rat suprachiasmatic nuclei. Neuroscience 120:745–755[CrossRef][Medline]
40. Bittman EL, Weaver DR 1990 The distribution of melatonin binding sites in neuroendocrine tissues of the ewe. Biol Reprod 43:986–993[Abstract]
41. Piggins HD, Cutler DJ 2003 The roles of vasoactive intestinal polypeptide in the mammalian circadian clock. J Endocrinol 177:7–15[Abstract]
42. Williams LM, Lincoln GA, Mercer JG, Barrett P, Morgan PJ, Clarke IJ 1997 Melatonin receptors in the brain and pituitary gland of hypothalamo-pituitary disconnected Soay rams. J Neuroendocrinol 9:639–643[CrossRef][Medline]
43. Stirland JA, Johnston JD, Cagampang FR, Morgan PJ, Castro MG, White MR, Davis JR, Loudon AS 2001 Photoperiodic regulation of prolactin gene expression in the Syrian hamster by a pars tuberalis-derived factor. J Neuroendocrinol 13:147–157[CrossRef][Medline]
44. Lincoln GA, Clarke IJ 1995 Evidence that melatonin acts in the pituitary gland through a dopamine-independent mechanism to mediate effects of daylength on the secretion of prolactin in the ram. J Neuroendocrinol 7:637–643[CrossRef][Medline]
45. Lincoln GA, Clarke IJ 2002 Noradrenaline and dopamine regulation of prolactin secretion in sheep: role in prolactin homeostasis but not photoperiodism. J Neuroendocrinol 14:36–44[CrossRef][Medline]
46. Shieh KR, Chu YS, Pan JT 1997 Circadian change of dopaminergic neuron activity: effects of constant light and melatonin. Neuroreport 8:2283–2287[Medline]
47. Chu YS, Shieh KR, Yuan ZF, Pan JT 2000 Stimulatory and entraining effect of melatonin on tuberoinfundibular dopaminergic neuron activity and inhibition on prolactin secretion. J Pineal Res 28:219–226[CrossRef][Medline]
48. Hazlerigg DG, Hastings MH, Morgan PJ 1996 Production of a prolactin releasing factor by the ovine pars tuberalis. J Neuroendocrinol 8:489–492[CrossRef][Medline]
49. Morgan PJ, Webster CA, Mercer JG, Ross AW, Hazlerigg DG, MacLean A, Barrett P 1996 The ovine pars tuberalis secretes a factor(s) that regulates gene expression in both lactotropic and nonlactotropic pituitary cells. Endocrinology 137:4018–4026[Abstract]
50. Lafarque M, Oliveros L, Aguado L 1998 Effect of adenohypophyseal pars tuberalis secretions on pars distalis prolactin liberation. Medicina 58:36–40
51. Johnston JD, Cagampang FR, Stirland JA, Carr AJ, White MR, Davis JR, Loudon AS 2003 Evidence for an endogenous per1- and ICER-independent seasonal timer in the hamster pituitary gland. FASEB J 17:810–815[Abstract/Free Full Text]

This article has been cited by other articles:

				J. M. Fustin, H. Dardente, G. C. Wagner, D. A. Carter, J. D. Johnston, G. A. Lincoln, and D. G. Hazlerigg Egr1 involvement in evening gene regulation by melatonin FASEB J, March 1, 2009; 23(3): 764 - 773. [Abstract] [Full Text] [PDF]

				J.M. Fustin, J.S. O'Neill, M.H. Hastings, D.G. Hazlerigg, and H. Dardente Cry1 Circadian Phase in vitro: Wrapped Up with an E-Box J Biol Rhythms, February 1, 2009; 24(1): 16 - 24. [Abstract] [PDF]

				S. M. Dupre, D. W. Burt, R. Talbot, A. Downing, D. Mouzaki, D. Waddington, B. Malpaux, J. R. E. Davis, G. A. Lincoln, and A. S. I. Loudon Identification of Melatonin-Regulated Genes in the Ovine Pituitary Pars Tuberalis, a Target Site for Seasonal Hormone Control Endocrinology, November 1, 2008; 149(11): 5527 - 5539. [Abstract] [Full Text] [PDF]

				D. J. MacGregor and G. A. Lincoln A Physiological Model of a Circannual Oscillator J Biol Rhythms, June 1, 2008; 23(3): 252 - 264. [Abstract] [PDF]

				F. J. Valenzuela, C. Torres-Farfan, H. G. Richter, N. Mendez, C. Campino, F. Torrealba, G. J. Valenzuela, and M. Seron-Ferre Clock Gene Expression in Adult Primate Suprachiasmatic Nuclei and Adrenal: Is the Adrenal a Peripheral Clock Responsive to Melatonin? Endocrinology, April 1, 2008; 149(4): 1454 - 1461. [Abstract] [Full Text] [PDF]

				M. J Paul, I. Zucker, and W. J Schwartz Tracking the seasons: the internal calendars of vertebrates Phil Trans R Soc B, January 27, 2008; 363(1490): 341 - 361. [Abstract] [Full Text] [PDF]

				G. C. Wagner, J. D. Johnston, I. J. Clarke, G. A. Lincoln, and D. G. Hazlerigg Redefining the Limits of Day Length Responsiveness in a Seasonal Mammal Endocrinology, January 1, 2008; 149(1): 32 - 39. [Abstract] [Full Text] [PDF]

This Article Abstract Full Text (PDF) Alert me when this article is cited Alert me if a correction is posted Citation Map Services Email this article to a friend Similar articles in this journal Similar articles in PubMed Alert me to new issues of the journal Download to citation manager Request Copyright Permission Citing Articles Citing Articles via HighWire Citing Articles via Google Scholar Google Scholar Articles by Johnston, J. D. Articles by Hazlerigg, D. G. Search for Related Content PubMed PubMed Citation Articles by Johnston, J. D. Articles by Hazlerigg, D. G.