Paul Pévet

Kisspeptin Mediates the Photoperiodic Control of Reproduction in Hamsters

The KiSS-1 gene encodes kisspeptin, the endogenous ligand of the G-protein-coupled receptor GPR54. Recent data indicate that the KiSS-1/GPR54 system is critical for the regulation of reproduction and is required for puberty onset. In seasonal breeders, reproduction is tightly controlled by photoperiod (i.e., day length). The Syrian hamster is a seasonal model in which reproductive activity is promoted by long summer days (LD) and inhibited by short winter days (SD). Using in situ hybridization and immunohistochemistry, we show that KiSS-1 is expressed in the arcuate nucleus of LD hamsters. Importantly, the KiSS-1 mRNA level was lower in SD animals but not in SD-refractory animals, which spontaneously reactivated their sexual activity after several months in SD. These changes of expression are not secondary to the photoperiodic variations of gonadal steroids. In contrast, melatonin appears to be necessary for these seasonal changes because pineal-gland ablation prevented the SD-induced downregulation of KiSS-1 expression. Remarkably, a chronic administration of kisspeptin-10 restored the testicular activity of SD hamsters despite persisting photoinhibitory conditions. Overall, these findings are consistent with a role of KiSS-1/GPR54 in the seasonal control of reproduction. We propose that photoperiod, via melatonin, modulates KiSS-1 signaling to drive the reproductive axis.

Daily variation in the concentration of melatonin and 5-methoxytryptophol in the human pineal gland: effect of age and Alzheimer's disease

Melatonin and 5-methoxytryptophol (ML) were measured in human pineals (38 controls, 16 subjects with Alzheimers disease). Time of death had a major influence on the indole concentrations with significantly higher melatonin levels occurring at night (22.00-10.00 h) and significantly higher ML levels occurring during the day (10.00-22.00 h). This daily variation disappeared in both the older subjects (55-92 years) and in the Alzheimer patients (55-89 years).

Melatonin: Both master clock output and internal time-giver in the circadian clocks network

Daily rhythms in physiological and behavioral processes are controlled by a network of circadian clocks, reset by inputs and delivering circadian signals to the brain and peripheral organs. In mammals, at the top of the network is a master clock located in the suprachiasmatic nuclei (SCN) of the hypothalamus, mainly reset by ambient light. The nocturnal synthesis and release of melatonin by the pineal gland are tightly controlled by the SCN clock and inhibited by light exposure. Several roles of melatonin in the circadian system have been identified. As a major hormonal output, melatonin distributes temporal cues generated by the SCN to the multitude of tissue targets expressing melatonin receptors. In some target structures, like the Pars tuberalis of the adenohypophysis, these melatonin signals can drive daily rhythmicity that would otherwise be lacking. In other target structures, melatonin signals are used for the synchronization (i.e., adjustment of the timing of existing oscillations) of peripheral oscillators, such as the fetal adrenal gland. Due to the expression of melatonin receptors in the SCN, endogenous melatonin is also able to feedback onto the master clock, although its physiological significance needs further characterization. Of note, pharmacological treatment with exogenous melatonin can synchronize the SCN clock. From a clinical point of view, provided that the subject is not exposed to light at night, the daily profile of circulating melatonin provides a reliable estimate of the timing of the human SCN. During the past decade, a number of melatonin agonists have been developed for treating circadian, psychiatric and sleep disorders. These drugs may target the SCN for improving circadian timing or act indirectly at some downstream level of the circadian network to restore proper internal synchronization.

The suprachiasmatic nucleus controls the daily variation of plasma glucose via the autonomic output to the liver: are the clock genes involved?

In order to drive tissue‐specific rhythmic outputs, the master clock, located in the suprachiasmatic nucleus (SCN), is thought to reset peripheral oscillators via either chemical and hormonal cues or neural connections. Recently, the daily rhythm of plasma glucose (characterized by a peak before the onset of the activity period) has been shown to be directly driven by the SCN, independently of the SCN control of rhythmic feeding behaviour. Indeed, the daily variation in glucose was not impaired unless the scheduled feeding regimen (six‐meal schedule) was associated with an SCN lesion. Here we show that the rhythmicity of both clock‐gene mRNA expression in the liver and plasma glucose is not abolished under such a regular feeding schedule. Because the onset of the activity period and hyperglycemia are correlated with an increased sympathetic tonus, we investigated whether this autonomic branch is involved in the SCN control of plasma glucose rhythm and liver rhythmicity. Interestingly, hepatic sympathectomy combined with a six‐meal feeding schedule resulted in a disruption of the plasma glucose rhythmicity without affecting the daily variation in clock‐gene mRNA expression in the liver. Taking all these data together, we conclude that (i) the SCN needs the sympathetic pathway to the liver to generate the 24‐h rhythm in plasma glucose concentrations, (ii) rhythmic clock‐gene expression in the liver is not dependent on the sympathetic liver innervation and (iii) clock‐gene rhythmicity in liver cells is not sufficient for sustaining a circadian rhythm in plasma glucose concentrations.

Melatonin Effects on Behavior: Possible Mediation by the Central GABAergic System

The best described function of the pineal hormone melatonin is to regulate seasonal reproduction, with its daily production and secretion varying throughout the seasons or the photoperiod. Additionally, a number of behavioral effects of the hormone have been found. This review describes the effects of melatonin in rodent behavior. We focus on: (a) inhibitory effects (sedation, hypnotic activity, pain perception threshold elevation, anti-convulsive activity, anti-anxiety effects); and (b) direct effects on circadian rhythmicity (entrainment, resynchronization, alleviation of jet-lag symptoms, phase-shifting). Most of these effects are clearly time-dependent, with a peak of melatonin activity during the night. One of the possible mechanisms of action for melatonin in the brain is the interaction with the GABAergic system, as suggested by neurochemical and behavioral data. Finally, some pineal hormone effects might be candidates as putative therapies for several human disorders.

Feeding Cues Alter Clock Gene Oscillations and Photic Responses in the Suprachiasmatic Nuclei of Mice Exposed to a Light/Dark Cycle

The suprachiasmatic nuclei (SCN) of the hypothalamus contain the master mammalian circadian clock, which is mainly reset by light. Temporal restricted feeding, a potent synchronizer of peripheral oscillators, has only weak influence on light-entrained rhythms via the SCN, unless restricted feeding is coupled with calorie restriction, thereby altering phase angle of photic synchronization. Effects of daytime restricted feeding were investigated on the mouse circadian system. Normocaloric feeding at midday led to a predominantly diurnal (60%) food intake and decreased blood glucose in the afternoon, but it did not affect the phase of locomotor activity rhythm or vasopressin expression in the SCN. In contrast, hypocaloric feeding at midday led to 2-4 h phase advances of three circadian outputs, locomotor activity rhythm, pineal melatonin, and vasopressin mRNA cycle in the SCN, and it decreased daily levels of blood glucose. Furthermore, Per1 and Cry2 oscillations in the SCN were phase advanced by 1 and 3 h, respectively, in hypocalorie- but not in normocalorie-fed mice. The phase of Per2 and Bmal1 expression remained unchanged regardless of feeding condition. Moreover, the shape of behavioral phase-response curve to light and light-induced expression of Per1 in the SCN were markedly modified in hypocalorie-fed mice compared with animals fed ad libitum. The present study shows that diurnal hypocaloric feeding affects not only the temporal organization of the SCN clockwork and circadian outputs in mice under light/dark cycle but also photic responses of the circadian system, thus indicating that energy metabolism modulates circadian rhythmicity and gating of photic inputs in mammals.

Immunohistochemical evidence for the presence of melatonin in the pineal gland, the retina and the Harderian gland.

SummaryThe presence of melatonin is demonstrated in the pineal gland, the retina and the Harderian gland in some mammalian and non-mammalian vertebrates, using a specific fluorescence labelled antibody technique. Four different potent antibodies against melatonin have been used and compared. In the pineal gland of hamsters, mice, rats and snakes, specific fluorescence, mostly restricted to the cytoplasm of the cells, is detected in pinealocytes. Fluorescence is also detected in the pineal organ of fishes, tortoises and lizards, but it has not been possible, from cryostat sections of fresh tissue, to assert which kind of cell is reacting (photoreceptor cells or interstitial ependymal cells). In the retina, fluorescence is almost exclusively restricted to the outer nuclear layer. In the Harderian gland of mammals and reptiles, fluorescence is localized in the secretory cells of the alveoli and mostly restricted to the cytoplasm surrounding the nucleus. These results are discussed in relation to the concept of melatonin synthesis at extrapineal sites independent of pineal production.

Melatonin sees the light: blocking GABA-ergic transmission in the paraventricular nucleus induces daytime secretion of melatonin.

Despite a pronounced inhibitory effect of light on pineal melatonin synthesis, usually the daily melatonin rhythm is not a passive response to the surrounding world. In mammals, and almost every other vertebrate species studied so far, the melatonin rhythm is coupled to an endogenous pacemaker, i.e. a circadian clock. In mammals the principal circadian pacemaker is located in the suprachiasmatic nuclei (SCN), a bilateral cluster of neurons in the anterior hypothalamus. In the present paper we show in the rat that bilateral abolition of γ‐aminobutyric acid (GABA), but not vasopressin, neurotransmission in an SCN target area, i.e. the paraventricular nucleus of the hypothalamus, during (subjective) daytime results in increased pineal melatonin levels. The fact that complete removal of the SCN results in a pronounced increase of daytime pineal mRNA levels for arylalkylamine N‐acetyltransferase (AA‐NAT), i.e. the rate‐limiting enzyme of melatonin synthesis, further substantiates the existence of a major inhibitory SCN output controlling the circadian melatonin rhythm.

Daily Rhythms of Melatonin Binding Sites in the Rat Pars tuberalis and Suprachiasmatic Nuclei; Evidence for a Regulation of Melatonin Receptors by Melatonin Itself

Using quantitative autoradiography, the density of melatonin binding sites has been measured in the rat pars tuberalis (PT) and suprachiasmatic nuclei (SCN) every 4 h throughout a 24-hour period in animals kept in a light regime of 12L/12D (with lights on at 07.00 h). Slices of PT and SCN were incubated in the presence of 180 and 172 pM, respectively, of 2-125I-melatonin. In both structures investigated, specific 2-125I-melatonin binding sites showed similar rhythms throughout the 24-hour period with a maximum at 16.00 h (PT: 46.9 +/- 2.8 fmol/mg protein, n = 5 and SCN: 5.12 +/- 0.30 fmol/mg protein, n = 5) and a minimum at 4.00 h (PT: 28.5 +/- 4.5 fmol/mg protein, n = 5 and SCN: 3.07 +/- 0.39 fmol/mg protein, n = 5). Similar experiments performed on PT of animals kept in constant light (LL) for 3 days revealed a lack of variations of melatonin binding site density, all the values being significantly higher than those of the respective 12L/12D group (concentration of 2-125I-melatonin used: 180 pM). All these preliminary results were confirmed by saturation studies performed at 16.00 and 4.00 h using quantitative autoradiography and in 12L/12D animals, using radioreceptor binding assays on isolated PT membranes. In 12L/12D animals, the maximum number of melatonin binding sites (Bmax) of both SCN and PT was significantly higher at 16.00 h than at 4.00 h. In all these cases, however, the dissociation constant (Kd) failed to show any significant daily variation.(ABSTRACT TRUNCATED AT 250 WORDS)

Suprachiasmatic control of melatonin synthesis in rats: inhibitory and stimulatory mechanisms

The suprachiasmatic nucleus (SCN) controls the circadian rhythm of melatonin synthesis in the mammalian pineal gland by a multisynaptic pathway including, successively, preautonomic neurons of the paraventricular nucleus (PVN), sympathetic preganglionic neurons in the spinal cord and noradrenergic neurons of the superior cervical ganglion (SCG). In order to clarify the role of each of these structures in the generation of the melatonin synthesis rhythm, we first investigated the day‐ and night‐time capacity of the rat pineal gland to produce melatonin after bilateral SCN lesions, PVN lesions or SCG removal, by measurements of arylalkylamine N‐acetyltransferase (AA‐NAT) gene expression and pineal melatonin content. In addition, we followed the endogenous 48 h‐pattern of melatonin secretion in SCN‐lesioned vs. intact rats, by microdialysis in the pineal gland. Corticosterone content was measured in the same dialysates to assess the SCN lesions effectiveness. All treatments completely eliminated the day/night difference in melatonin synthesis. In PVN‐lesioned and ganglionectomised rats, AA‐NAT levels and pineal melatonin content were low (i.e. 12% of night‐time control levels) for both day‐ and night‐time periods. In SCN‐lesioned rats, AA‐NAT levels were intermediate (i.e. 30% of night‐time control levels) and the 48‐h secretion of melatonin presented constant levels not exceeding 20% of night‐time control levels. The present results show that ablation of the SCN not only removes an inhibitory input but also a stimulatory input to the melatonin rhythm generating system. Combination of inhibitory and stimulatory SCN outputs could be of a great interest for the mechanism of adaptation to day‐length (i.e. adaptation to seasons).