Light-Induced Variations in AP-1 Binding Activity and Composition in the Rat Suprachiasmatic Nucleus

Abstract : Expression of immediate early genes, including fos -like and jun -like genes, in the suprachiasmatic nucleus is believed to be part of the mechanism for photic entrainment of circadian rhythms to the environmental light/dark cycle. However, the effects of a light stimulus on activating protein-1 (AP-1) complexes in the suprachiasmatic nucleus remain unclear. The photic regulation of AP-1 DNA-binding activity and composition in the rat suprachiasmatic nucleus was evaluated by using an electrophoretic mobility shift assay. A light pulse given during subjective night induced an increase in AP-1 binding activity when either nuclear or whole-cell extracts from suprachiasmatic nuclei were used. Under constant dark conditions, proteins that are predominant components of AP-1 complexes are Fra-2 and Jun-D. Under light stimulation, c-Fos and Jun-B consistently increased, as expected, but this was also the case for Fra-2, Jun-D, and c-Jun, although to a lesser extent. An immunocytochemical study of the Fra-2 expression pattern demonstrated the presence of the protein in the ventrolateral as well as in the dorsomedial subdivisions of the suprachiasmatic nucleus. Light regulation of Fra-2 immunoreactivity, however, appeared to be restricted to the ventrolateral subdivision. It is concluded that light may be acting both by increasing constitutive AP-1 complexes and by inducing the expression of specific complexes.     

Photoentrainment, pharmacology, and phase shifts of the circadian rhythm in the rat pineal.

Photoentrainment of circadian rhythms in mammals is mediated by the retinohypothalamic projection to the suprachiasmatic nucleus of the hypothalamus. It should therefore be possible to mimic or block the effects of light on the circadian pacemaker with appropriate pharmacological agents. Such agents and their effects should be useful in identifying the neurotransmitters involved in photoentrainment and their mechanisms of action on the circadian pacemaker. The effects of light on the circadian rhythm in rat pineal serotonin N-acetyltransferase activity are described. Carbachol, a cholinergic agonist, was found to mimic the effects of light on this rhythm, including the acute reduction of nocturnal activity and phase-shifting of the free-running rhythm. These results raise the possibility that acetylcholine is involved in the photoentrainment of mammalian circadian rhythms.     

Circadian expression of clock genes in the rat eye and brain

The light sensing system in the eye directly affects the circadian oscillator in the mammalian suprachiasmatic nucleus (SCN). To investigate this relationship in the rat, we examined the circadian expression of clock genes in the SCN and eye tissue during a 24 h day/night cycle. In the SCN, rPer1 and rPer2 mRNAs were expressed in a clear circadian rhythm like rCry1 and rCry2 mRNAs, whereas the level of BMAL1 and CLOCK mRNAs decreased during the day and increased during the night with a relatively low amplitude. It seems that the clock genes of the SCN may function in response to a master clock oscillation in the rat. In the eye, the rCry1 and rCry2 were expressed in a circadian rhythm with an increase during subjective day and a decrease during subjective night. However, the expression of Opn4 mRNA did not exhibit a clear circadian pattern, although its expression was higher in daytime than at night. This suggests that cryptochromes located in the eye, rather than melanopsin, are the major photoreceptive system for synchronizing the circadian rhythm of the SCN in the rat.     

Avian Melatonin Synthesis: Photic and Circadian Regulation of Serotonin N-Acetyltransferase mRNA in the Chicken Pineal Gland and Retina

Abstract: The circadian rhythms in melatonin production in the chicken pineal gland and retina reflect changes in the activity of serotonin N -acetyltransferase (arylalkylamine N -acetyltransferase; AA-NAT; EC 2.3.1.87). Here we determined that the chicken AA-NAT mRNA is detectable in follicular pineal cells and retinal photoreceptors and that it exhibits a circadian rhythm, with peak levels at night. AA-NAT mRNA was not detected in other tissues. The AA-NAT mRNA rhythm in the pineal gland and retina persists in constant darkness (DD) and constant lighting (LL). The amplitude of the pineal mRNA rhythm is not decreased in LL. Light appears to influence the phase of the clock driving the rhythm in pineal AA-NAT mRNA in two ways: The peak is delayed by ∼6 h in LL, and it is advanced by >4 h by a 6-h light pulse late in subjective night in DD. Nocturnal AA-NAT mRNA levels do not change during a 20-min exposure to light, whereas this treatment dramatically decreases AA-NAT activity. These observations suggest that the rhythmic changes in chicken pineal AA-NAT activity reflect, at least in part, clock-generated changes in mRNA levels. In contrast, changes in mRNA content are not involved in the rapid light-induced decrease in AA-NAT activity.     

褪黑素对大鼠下丘脑薄片视交叉上核神经元放电昼夜节律性的调制

先用持续光照和松果腺切除预处理大鼠,然后制成下丘脑薄片,记录其视交叉上核（ＳＣＮ）神经元自发放电,观察其昼夜变化和褪黑素（ＭＥＬ）对它的影响。实验结果表明：⑴在正常光照（光照：黑暗＝１２：１２）条件下,ＳＣＮ神经元自发放电频率呈现昼夜低的节律性。在昼夜时间（ＣＴ）６－８出现放电高峰,频率约为８．３Ｈｚ;在ＣＴ１８－２０出现低谷,频率约为３．８Ｈｚ。松果腺切除后,ＳＣＮ神经元自发放电的昼夜节律性基本     

Species Differences Between Hamsters and Rats with Regard to the Putative Role of Serotonin in the Circadian System

In mammals, circadian rhythms of locomotor activity and many other behavioral and physiological functions are controlled by an endogenous pacemaker located in the hypothalamic suprachiasmatic nucleus (SCN). Among various other afferents, the SCN receives a dense serotonergic input from the mesencephalic raphe complex. Experimental evidence obtained so far in Syrian hamsters suggests that serotonin (5-HT) mimics the effect of nonphotic stimuli during subjective day and modulates photic input to the SCN during subjective night. These findings are consistent with a putative role of serotonergic pathways in the transmission of the state of arousal to the SCN. In this paper, we review recent evidence for different modes of 5-HT action and/or the involvement of different 5-HT receptor subtypes in hamsters and rats. In intact rats, 5-HT agonists induce photic-like phase shifts of locomotor activity and melatonin rhythms as well as c-Fos expression in the ventral SCN. These results suggest a role for 5-HT in the transmission of photic rather than nonphotic information to the rat SCN. Such a function of 5-HT would also explain why the circadian system of rats is less sensitive or even insensitive to nonphotic stimuli.     

Species Differences Between Hamsters and Rats with Regard to the Putative Role of Serotonin in the Circadian System

In mammals, circadian rhythms of locomotor activity and many other behavioral and physiological functions are controlled by an endogenous pacemaker located in the hypothalamic suprachiasmatic nucleus (SCN). Among various other afferents, the SCN receives a dense serotonergic input from the mesencephalic raphe complex. Experimental evidence obtained so far in Syrian hamsters suggests that serotonin (5-HT) mimics the effect of nonphotic stimuli during subjective day and modulates photic input to the SCN during subjective night. These findings are consistent with a putative role of serotonergic pathways in the transmission of the state of arousal to the SCN. In this paper, we review recent evidence for different modes of 5-HT action and/or the involvement of different 5-HT receptor subtypes in hamsters and rats. In intact rats, 5-HT agonists induce photic-like phase shifts of locomotor activity and melatonin rhythms as well as c-Fos expression in the ventral SCN. These results suggest a role for 5-HT in the transmission of photic rather than nonphotic information to the rat SCN. Such a function of 5-HT would also explain why the circadian system of rats is less sensitive or even insensitive to nonphotic stimuli.     

Circadian and seasonal rhythms of 5-HT receptor subtypes, membrane anisotropy and 5-HT release in hippocampus and cortex of the rat.

Specific serotonin binding (5-HT1, 5-HT1A, and 5-HT2 subtypes) and membrane anisotropy were measured at 2 h intervals over a 24 h period in the hippocampus and cortex of Wistar WU rats, housed under a 12 h light-dark cycle, with lights on at 07.00. All experiments were performed both in March and December. In the hippocampus significant circadian rhythms could be ascertained for 5-HT1 binding sites in March and December while for 5-HT1A (subtype of 5-HT1) binding sites the circadian rhythm was only significant in March. The membrane anisotropy also showed significant variations only in March. Circadian rhythms were also found in the cortex for 5-HT1 (December) and 5-HT2 (March and December) binding sites as well as for the membrane anisotropy (December). A correlation was found between membrane anisotropy and 5-HT1 and 5-HT2 binding sites in hippocampus and cortex, respectively. A circadian rhythmicity was also observed for serotonin release as measured by in vivo voltammetry in both brain areas. The results obtained on the diurnal variations of serotonin receptor subtypes and serotonin release and the probable inverse relationship of these two parameters may be relevant in understanding the coupling of pre- and postsynaptic activity.     

The circadian timing system and reproduction in mammals.

Circadian systems in a wide variety of organisms all appear to include three basic components: 1) biological oscillators that maintain a self-sustained circadian periodicity in the absence of environmental time cues; 2) input pathways that convey environmental information, especially light cues, that can entrain the circadian oscillations to local time; and 3) output pathways that drive overt circadian rhythms, such as the rhythms of locomotor activity and a variety of endocrine rhythms. In mammals, the circadian system is employed in the regulation of reproductive physiology and behavior in two very important ways. 1) In some species, there is a strong circadian component in the timing of ovulation and reproductive behavior, ensuring that these events will occur at a time when the animal is most likely to encounter a potential mate. 2) Many mammals exhibit seasonal reproductive rhythms that are largely under photoperiod regulation; in these species, the circadian system and the pineal gland are crucial components of the mechanism that is used to measure day length. The rhythm of pineal melatonin secretion is driven by a neural pathway that includes the circadian oscillator(s) in the suprachiasmatic nuclei. Melatonin is secreted at night in all mammals, and the duration of each nocturnal episode of melatonin secretion is inversely related to day length. The pineal melatonin rhythm appears to serve as an internal signal that represents day length and that is capable of regulating a variety of seasonal variations in physiology and behavior.     

Serotonin agonist quipazine induces photic-like phase shifts of the circadian activity rhythm and c-Fos expression in the rat suprachiasmatic nucleus

Nonphotic stimuli can reset and entrain circadian activity rhythms in hamsters and mice, and serotonin is thought to be involved in the phase-resetting effects of these stimuli. In the present study, the authors examined the effect of the serotonin agonist quipazine on circadian activity rhythms in three inbred strains of rats (ACI, BH, and LEW). Furthermore, they investigated the effect of quipazine on the expression of c-Fos in the mammalian circadian pacemaker, the suprachiasmatic nucleus (SCN). Quipazine reduced the amount of running wheel activity for 3 h after treatment, however, no long-term changes in tau and in the activity level were observed. More important, quipazine induced significant phase advances of the activity rhythm and c-Fos production in the SCN at the end of the subjective night (Circadian Time [CT] 22), whereas neither phase shifts nor c-Fos induction were observed during the subjective day. Quipazine injections also resulted in moderate phase delays at the beginning of the subjective night (CT 14). A similar phase-response characteristic typically can be observed for photic stimuli. By contrast, nonphotic stimuli normally produce phase advances during the subjective day. The present results suggest species differences between the hamster and the rat with respect to the serotonergic action on circadian timekeeping and indicate that serotonergic pathways play a role in the transmission of photic information to the SCN of rats.     

Effect of long-term exposure to constant dim light on the circadian system of rats

The newly discovered multi-oscillatory nature of the mammalian circadian clock system and the cloning of the genes involved in the molecular mechanism that generates circadian rhythmicity have opened new approaches for understanding how mammals are temporally organized and how the mammalian circadian system reacts to the lack of normal synchronization cues. In the present study we investigated the effects of long-term exposure to constant red dim light on the pattern of the expression of Period 1 in the suprachiasmatic nuclei of the hypothalamus and of Arylalkylamine N-acetyltransferase(Aa-nat) in the retina and pineal gland. Our data demonstrate that Period 1 mRNA expression in the suprachiasmatic nuclei of the hypothalamus was not affected by exposure to constant red dim light for 60 days, whereas Aa-nat mRNA expression in the retina and in the pineal gland was significantly affected, since in some animals (20-30%) Aa-nat mRNA levels were found to be higher during the subjective day. A circadian rhythm of serum melatonin and locomotor activity was present in all the animals tested. In 4 animals serum melatonin levels were high during the subjective day. Our data suggest that long-term exposure to constant red dim light may induce desynchronization between the circadian rhythm of locomotor activity and serum melatonin levels.     

Long-Lasting Effects of Sepsis on Circadian Rhythms in the Mouse

Daily patterns of activity and physiology are termed circadian rhythms and are driven primarily by an endogenous biological timekeeping system, with the master clock located in the suprachiasmatic nucleus. Previous studies have indicated reciprocal relationships between the circadian and the immune systems, although to date there have been only limited explorations of the long-term modulation of the circadian system by immune challenge, and it is to this question that we addressed ourselves in the current study. Sepsis was induced by peripheral treatment with lipopolysaccharide (5 mg/kg) and circadian rhythms were monitored following recovery. The basic parameters of circadian rhythmicity (free-running period and rhythm amplitude, entrainment to a light/dark cycle) were unaltered in post-septic animals compared to controls. Animals previously treated with LPS showed accelerated re-entrainment to a 6 hour advance of the light/dark cycle, and showed larger phase advances induced by photic stimulation in the late night phase. Photic induction of the immediate early genes c-FOS, EGR-1 and ARC was not altered, and neither was phase-shifting in response to treatment with the 5-HT-1a/7 agonist 8-OH-DPAT. Circadian expression of the clock gene product PER2 was altered in the suprachiasmatic nucleus of post-septic animals, and PER1 and PER2 expression patterns were altered also in the hippocampus. Examination of the suprachiasmatic nucleus 3 months after treatment with LPS showed persistent upregulation of the microglial markers CD-11b and F4/80, but no changes in the expression of various neuropeptides, cytokines, and intracellular signallers. The effects of sepsis on circadian rhythms does not seem to be driven by cell death, as 24 hours after LPS treatment there was no evidence for apoptosis in the suprachiasmatic nucleus as judged by TUNEL and cleaved-caspase 3 staining. Overall these data provide novel insight into how septic shock exerts chronic effects on the mammalian circadian system.     

Paraventricular Thalamus Lesions Alter Circadian Period and Activity Distribution in the Blinded Rat

The paraventricular thalamic nucleus (PVT) is reciprocally connected with the suprachiasmatic nucleus, the mammalian circadian pacemaker. In this study, we examine the effects of PVT lesions on the free-running circadian rhythm of locomotor activity in the blinded, albino rat. In the blinded rat, lesions of the PVT cause a period lengthening of the free-running circadian rhythm and a change in the pattern of locomotor activity. In animals with complete PVT lesions, locomotor activity is concentrated in late subjective night as compared to the pre-lesion control. Our results suggest that the PVT participates in the regulation of suprachiasmatic pacemaker function and that this regulation may be modified by retinal input.     

Paraventricular Thalamus Lesions Alter Circadian Period and Activity Distribution in the Blinded Rat

The paraventricular thalamic nucleus (PVT) is reciprocally connected with the suprachiasmatic nucleus, the mammalian circadian pacemaker. In this study, we examine the effects of PVT lesions on the free-running circadian rhythm of locomotor activity in the blinded, albino rat. In the blinded rat, lesions of the PVT cause a period lengthening of the free-running circadian rhythm and a change in the pattern of locomotor activity. In animals with complete PVT lesions, locomotor activity is concentrated in late subjective night as compared to the pre-lesion control. Our results suggest that the PVT participates in the regulation of suprachiasmatic pacemaker function and that this regulation may be modified by retinal input.