Identification of the suprachiasmatic nucleus in birds

Circadian rhythms are generated by an internal biological clock. The suprachiasmatic nucleus (SCN) in the hypothalamus is known to be the dominant biological clock regulating circadian rhythms in mammals. In birds, two nuclei, the so-called medial SCN (mSCN) and the visual SCN (vSCN), have both been proposed to be the avian SCN. However, it remains an unsettled question which nuclei are homologous to the mammalian SCN. We have identified circadian clock genes in Japanese quail and demonstrated that these genes are expressed in known circadian oscillators, the pineal and the retina. Here, we report that these clock genes are expressed in the mSCN but not in the vSCN in Japanese quail, Java sparrow, chicken, and pigeon. In addition, mSCN lesions eliminated or disorganized circadian rhythms of locomotor activity under constant dim light, but did not eliminate entrainment under light-dark (LD) cycles in pigeon. However, the lesioned birds became completely arrhythmic even under LD after the pineal and the eye were removed. These results indicate that the mSCN is a circadian oscillator in birds.     

Complex bird clocks.

The circadian pacemaking system of birds comprises three major components: (i) the pineal gland, which rhythmically synthesizes and secretes melatonin; (ii) a hypothalamic region, possibly equivalent to the mammalian suprachiasmatic nuclei; and (iii) the retinae of the eyes. These components jointly interact, stabilize and amplify each other to produce a highly self-sustained circadian output. Their relative contribution to overt rhythmicity appears to differ between species and the system may change its properties even within an individual depending, for example, on its state in the annual cycle or its photic environment. Changes in pacemaker properties are partly mediated by changes in certain features of the pineal melatonin rhythm. It is proposed that this variability is functionally important, for instance, for enabling high-Arctic birds to retain synchronized circadian rhythms during the low-amplitude zeitgeber conditions in midsummer or for allowing birds to adjust quickly their circadian system to changing environmental conditions during migratory seasons. The pineal melatonin rhythm, apart from being involved in generating the avian pacemaking oscillation, is also capable of retaining day length information after isolation from the animal. Hence, it appears to participate in photoperiodic after-effects. Our results suggest that complex circadian clocks have evolved to help birds cope with complex environments.     

Hypothalamic regulation of circadian noradrenergic input to the chick pineal gland

Summary While the avian pineal gland contains circadian oscillators and photoreceptors capable of producing circadian rhythms of the hormone melatonin, it is extensively innervated by post-ganglionic fibers of the superior cervical ganglia which release norepinephrine (NE) rhythmically. Norepinephrine turnover is higher during subjective day than during subjective night. In mammals, this rhythmic input, which is higher in subjective night than subjective day, derives from the hypothalamic suprachiasmatic nuclei (SCN) and is essential for rhythmic melatonin production. The present study was designed to determine whether one of two candidates for the avian homologue of the mammalian SCN is necessary for rhythmic NE turnover in the chick pineal gland. Either electrolytic lesions or sham lesions were delivered to the periventricular preoptic nuclei (PPN) or to the visual suprachiasmatic nucleus (vSCN). After recovery, the rates of decline in [NE] were determined following pretreatment with -methyl-p-tyrosine, a tyrosine hydroxylase inhibitor, at mid-subjective day or at mid-subjective night. Birds receiving sham surgeries in either PPN or vSCN and birds receiving lesions of the PPN exhibited rhythmicity in NE turnover. No rhythm of NE turnover could be determined in birds with ablated vSCN.Abbreviations AMPT -methyl-p-tyrosine - DS supraoptic decussation - EBZ ear bar zero (see Methods) - GLv ventral lateral geniculate body - NE norepinephrine - PPN periventricular preoptic nuclei - RH retinohypothalamic projection - SCN suprachiasmatic nuclei - vSCN visual suprachiasmatic nucleus     

Circadian organization in the pigeon,Columba livia: the role of the pineal organ and the eye

Summary The roles of the pineal organ and the eye in the control of circadian locomotor rhythmicity were studied in the pigeon (Columba livia). Neither pinealectomy nor blinding abolished the circadian rhythms in constant dim light conditions (LLdim). All the pinealectomized birds and the blinded birds entrained to light-dark (LD) cycles with no discernible anticipatory activity. However, the birds which had been both pinealectomized and blinded showed no circadian rhythms in prolonged LLdim. These birds entrained to LD cycles with anticipatory activity and showed residual rhythmicity for a while after transfer from LD cycles to LLdim. Continuous administration of melatonin induced suppression of the circadian rhythms and reduced total amount of locomotor activity in LLdim. These results suggest that not only the pineal organ but also the eye (perhaps the retina) is involved in the pigeon's circadian system.Abbreviations NAT N-acetyltransferase - LLdim constant dim light - cadian period - SCN suprachiasmatic nucleus - circadian activity time - LD light-dark     

The pineal gland influences rat circadian activity rhythms in constant light.

Mammalian circadian organization is believed to derive primarily from circadian oscillators within the hypothalamic suprachiasmatic nuclei (SCN). The SCN drives circadian rhythms of a wide array of functions (e.g., locomotion, body temperature, and several endocrine processes, including the circadian secretion of the pineal hormone melatonin). In contrast to the situation in several species of reptiles and birds, there is an extensive literature reporting little or no effect of pinealectomy on mammalian circadian rhythms. However, recent research has indicated that the SCN and circadian systems of several mammalian species are highly sensitive to exogenous melatonin, raising the possibility that endogenous pineal hormone may provide feedback in the control of overt circadian rhythms. To determine the role of the pineal gland in rat circadian rhythms, the effects of pinealectomy on locomotor rhythms in constant light (LL) and constant darkness (DD) were studied. The results indicated that the circadian rhythms of pinealectomized rats but not sham-operated controls dissociated into multiple ultradian components in LL and recoupled into circadian patterns only after 12-21 days in DD. The data suggest that pineal feedback may modulate sensitivity to light and/or provide coupling among multiple circadian oscillators within the SCN.     

Complete suprachiasmatic lesions eliminate circadian rhythmicity of body temperature and locomotor activity in golden hamsters

The effects of suprachiasmatic and control lesions on the circadian rhythms of locomotor activity and body temperature were studied in golden hamsters (Mesocricetus auratus) maintained in constant light as well as constant darkness. Large suprachiasmatic lesions, but not control lesions, eliminated circadian rhythmicity in locomotor activity as well as in body temperature. Analysis of the robustness of the rhythms of locomotor activity and body temperature in unlesioned and lesioned animals suggests that, because body temperature rhythmicity is more robust than locomotor rhythmicity, lesions that spare a small number of suprachiasmatic cells might abolish the latter but not the former. Our results do not support the hypothesis that the body temperature rhythm is controlled by a circadian pacemaker distinct from the main pacemaker located in the suprachiasmatic nuclei.     

Daily melatonin administration synchronizes circadian patterns of brain metabolism and behavior in pinealectomized house sparrows,Passer domesticus

Recent research in our laboratory has indicated that in sparrows the visual suprachiasmatic nucleus (vSCN) is metabolically rhythmic such that 2-deoxy[¹⁴C]glucose (2DG) uptake and specific binding of 2[¹²⁵I]iodomelatonin (IMEL) are high during subjective day for up to 10 circadian cycles in constant darkness (DD). These rhythms damp to arrhythmicity in pinealectomized birds (PINX). The present study was designed to test the hypothesis that exogenous melatonin rhythmically applied can restore disrupted behavioral and cerebral rhythmicity. Pinealectomized house sparrows were placed in constant dim light and allowed to become arrhythmic. Experimental birds received 0.86 mM melatonin in 0.01% ethanol (ETOH) to drink for 12 of every 24 h for 14 days. Control birds received 0.01% ETOH only. Behavioral rhythmicity was restored by melatonin but not by ETOH. Birds were injected with 2DG 6 or 18 h following the beginning of melatonin (for experimental birds: MT06 and MT18 respectively) or ETOH (for control birds: ET06 and ET18 respectively) administration, allowed to survive 1 h and killed for 2DG and IMEL autoradiography. The data indicated 2DG rhythmicity such that uptake was high at MT18 in vSCN and several visual, auditory and limbic system structures in birds receiving melatonin but not in birds receiving ETOH. Similarly, IMEL binding rhythms were restored in vSCN and other visual, auditory and limbic system structures in birds receiving melatonin but not in those receiving ETOH. These data indicate that melatonin cycles are responsible for generating and/or driving a wide array of cerebral metabolic rhythms and that this influence is inhibitory.     

Circadian rhythms of indoleamines and serotonin N-acetyltransferase activity in the pineal gland

Conclusion The circadian rhythm of melatonin synthesis in the pineal glands of various species has been summarized. The night-time elevation of melatonin content is in most if not all cases regulated by the change of N-acetyltransferase activity. In mammals, the N-acetyltransferase rhythm is controlled by the central nervous system, presumably by suprachiasmatic nuclei in hypothalamus through the superior cervical ganglion. In birds, the circadian oscillator that regulates the N-acetyltransferase rhythm is located in the pineal glands. The avian pineal gland may play a biological clock function to control the circadian rhythms in physiological, endocrinological and biochemical processes via pineal hormone melatonin.     

Early development of circadian rhythmicity in the suprachiamatic nuclei and pineal gland of teleost,flounder (Paralichthys olivaeus), embryos

Circadian rhythms enable organisms to coordinate multiple physiological processes and behaviors with the earth's rotation. In mammals, the suprachiasmatic nuclei (SCN), the sole master circadian pacemaker, has entrainment mechanisms that set the circadian rhythm to a 24‐h cycle with photic signals from retina. In contrast, the zebrafish SCN is not a circadian pacemaker, instead the pineal gland (PG) houses the major circadian oscillator. The SCN of flounder larvae, unlike that of zebrafish, however, expresses per2 with a rhythmicity of daytime/ON and nighttime/OFF. Here, we examined whether the rhythm of per2 expression in the flounder SCN represents the molecular clock. We also examined early development of the circadian rhythmicity in the SCN and PG. Our three major findings were as follows. First, rhythmic per2 expression in the SCN was maintained under 24 h dark (DD) conditions, indicating that a molecular clock exists in the flounder SCN. Second, onset of circadian rhythmicity in the SCN preceded that in the PG. Third, both 24 h light (LL) and DD conditions deeply affected the development of circadian rhythmicity in the SCN and PG. This is the first report dealing with the early development of circadian rhythmicity in the SCN in fish.     

Circadian regulation of bird song, call, and locomotor behavior by pineal melatonin in the zebra finch

As both a photoreceptor and pacemaker in the avian circadian clock system, the pineal gland is crucial for maintaining and synchronizing overt circadian rhythms in processes such as locomotor activity and body temperature through its circadian secretion of the pineal hormone melatonin. In addition to receptor presence in circadian and visual system structures, high-affinity melatonin binding and receptor mRNA are present in the song control system of male oscine passeriform birds. The present study explores the role of pineal melatonin in circadian organization of singing and calling behavior in comparison to locomotor activity under different lighting conditions. Similar to locomotor activity, both singing and calling behavior were regulated on a circadian basis by the central clock system through pineal melatonin, since these behaviors free-ran with a circadian period and since pinealectomy abolished them in constant environmental conditions. Further, rhythmic melatonin administration restored their rhythmicity. However, the rates by which these behaviors became arrhythmic and the rates of their entrainment to rhythmic melatonin administration differed among locomotor activity, singing and calling under constant dim light and constant bright light. Overall, the study demonstrates a role for pineal melatonin in regulating circadian oscillations of avian vocalizations in addition to locomotor activity. It is suggested that these behaviors might be controlled by separable circadian clockworks and that pineal melatonin entrains them all through a circadian clock.     

Pineal regulation of circadian rhythms of 2-deoxy[14C]glucose uptake and 2[125I]iodomelatonin binding in the visual system of the house sparrow,Passer domesticus

The pineal gland and its hormone melatonin are crucial for the generation of circadian rhythms in several species of passerine birds. The sites and mechanisms by which they influence avian behavior are therefore of particular interest. Recent research employing several brain imaging techniques has indicated that the sites of melatonin action within the avian brain are wide-spread within the 4 major visual pathways. In this study, we have investigated whether the avian homologue of the mammalian suprachiasmatic nucleus, the visual suprachiasmatic nucleus (vSCN), and other visually sensitive structures express circadian rhythms of 2-deoxy[¹⁴C]glucose (2DG) uptake and 2[¹²⁵I]iodomelatonin (IMEL) binding in house sparrows,Passer domesticus, under constant environmental conditions in the presence or absence of the pineal gland. The results indicate that 2DG uptake in the vSCN is oscillatory in sham-operated sparrows but damps to arrhythmicity in pinealectomized birds, suggesting this structure contains a damped circadian oscillator independent of pineal input. We have also asked whether IMEL binding is rhythmic under these conditions in the same brains. These results indicate IMEL binding is rhythmic in several structures in the circadian, tectofugal, thalamofugal visual pathways and that pinealectomy increases the level of IMEL binding 2–4 fold suggesting that IMEL binding is down regulated by endogenous melatonin. However, the circadian rhythm of this binding is only gradually abolished, suggesting it too is regulated by a non-pineal circadian clock. These data are discussed in the context of the behavioral neurobiology of avian circadian systems and the neuroendocrine loop model.     

The contrary effect of destruction of the dorsal hippocampus and of the suprachiasmatic nucleus of the hypothalamus on the rhythmic organization of behavior and on anxiety in rats]

Dorsal hippocampal lesions increase the amplitude of the circadian rhythms of locomotion and the number of long-period cycles in the structure of forced swimming and simultaneously decrease anxiety in rats. Bilateral destruction of the circadian pacemaker (suprachiasmatic nuclei of hypothalamus) induces the opposite shifts in the rhythmic organization of behavior and anxiety of animals.     

Electrical and pharmacological properties of the suprachiasmatic nuclei

The suprachiasmatic nuclei (SCN) of the mammalian hypothalamus are in important circadian pacemaker. The electrical activity of these nuclei exhibits an intrinsic circadian rhythm. The rhythmicity of the SCN is also reflected in cyclic glucose consumption and serotonin metabolism. These rhythms are entrained to the light-dark cycle via the retinohypothalamic projection. This pathway, possibly together with a visual projection via the ventral lateral geniculate nuclei, innervates light-responsive SCN cells, which exhibit the functional properties of luminance detectors. The SCN contain various peptides, acetylcholine, and serotonin either intrinsically or in terminals of afferent projections. For acetylcholine it has been demonstrated that the SCN mediate the process of photic entrainment and light suppression of pineal synthetic activity. In the case of serotonin and vasopressin it seems certain that the SCN do not depend on their presence for generating circadian rhythms or for entrainment. Both substances may modulate the intrinsic pacemaker frequency through mechanisms that remain to be established.     

Differential elimination of circadian and ultradian rhythmicity by hypothalamic lesions in the common vole, Microtus arvalis

Effects of hypothalamic lesions on the ultradian and circadian organization of wheel running and feeding were studied in the common vole, Microtus arvalis. Circadian organization broke down within 30 days in continuous darkness in 24% of intact voles (n = 135). Ultradian rhythmicity of feeding (period 2-3 hr) persisted in constant conditions in all intact voles. Following lesions of the suprachiasmatic nuclei (SCN), circadian rhythmicity disappeared when lesions were complete (n = 8) or more extensive than 25% of the total SCN volume (n = 5). Absence of circadian rhythmicity was also found in animals with substantial lesions in the diencephalic paraventricular area (PVA) and in the retrochiasmatic area (RCA) and/or adjacent arcuate nucleus (Arc). Complete loss of ultradian and circadian organization occurred in eight voles with damage to the RCA and/or Arc. In three of these, the SCN was intact. The SCN is a likely candidate for a circadian pacemaker in voles (as in other rodents), while the loss of circadian rhythmicity following PVA and RCA/Arc lesions may be due to destruction of efferent pathways from the SCN. The RCA/Arc area is apparently necessary for the expression of ultradian rhythms. The intact SCN is neither necessary nor sufficient for the generation of ultradian rhythmicity.     

Neural control of circadian rhythms in plasma ACTH, plasma corticosterone and motor activity

The circadian rhythms for plasma ACTH and corticosterone (B), as well as motor activity, were explored in female rats after ocular enucleation (O-X), stereotaxic lesion of the suprachiasmatic nuclei (SCN-X) or of midbrain raphe nuclei (R-X), or both O-X and R-X, pharmacological blockade of the serotoninergic (5HT) system by pCPA, sometimes bypassed by 5-HTP, or 5-HT denervation of the SCN by local injection of 5,7-DHT. The three circadian rhythms explored responded quite differently to the treatments. In particular, the ACTH and B rhythms lost their usual close correlations. The amplitude and mean level of ACTH fluctuations were depressed after all treatments, but remained normal or were enhanced for B rhythm. ACTH rhythmicity actually was undetectable after SCN-X, pCPA and, in several rats, combined O-X and R-X, whereas persisting circadian and/or ultradian B and locomotor activity rhythms were always measured. The participation of the suprachiasmatic nuclei, the midbrain raphe nuclei and other possible 5-HT components in a complex circadian pacemaker system is discussed.     

Effects of preparation time on phase of cultured tissues reveal complexity of circadian organization

The phases of central (SCN) and peripheral circadian oscillators are held in specific relationships under LD cycles but, in the absence of external rhythmic input, may damp or drift out of phase with each other. Rats exposed to prolonged constant light become behaviorally arrhythmic, perhaps as a consequence of dissociation of phases among SCN cells. The authors asked whether individual central and peripheral circadian oscillators were rhythmic in LL-treated arrhythmic rats and, if rhythmic, what were the phase relationships between them. The authors prepared SCN, pineal gland, pituitary, and cornea cultures from transgenic Period1-luciferaserats whose body temperature and locomotor activity were arrhythmic and from several groups of rhythmic rats held in LD, DD, and short-term LL. The authors measured mPer1gene expression by recording light output with sensitive photomultipliers. Most of the cultures from all groups displayed circadian rhythms. This could reflect persistent rhythmicity in vivo prior to culture or, alternatively, rhythmicity that may have been initiated by the culture procedure. To test this, the authors cultured tissues at 2 different times 12 h apart and asked whether phase of the rhythm was related to culture time. The pineal, pituitary, and SCN cultures showed partial or complete dependence of phase on culture time, while peak phases of the cornea cultures were independent of culture time in rhythmic rats and were randomly distributed regardless of culture time in arrhythmic animals. These results suggest that in behaviorally arrhythmic rats, oscillators in the pineal, pituitary, and SCN had been arrhythmic or severely damped in vivo, while the cornea oscillator was free running. The peak phases of the SCN cultures were particularly sensitive to some aspect of the culture procedure since rhythmicity of SCN cultures from robustly rhythmic LD-entrained rats was strongly influenced when the procedure was carried out at any time except the 2nd half of the day.