Analysis of coupling between optic lobe circadian pacemakers in the cricket Gryllus bimaculatus

The coupling mechanism between weakly coupled two optic lobe circadian pacemakers in the cricket Gryllus bimaculatus was investigated by recording the locomotor activity, under light-dark cycles with various lengths, after the optic nerve was unilaterally severed. The activity rhythm split into two components under the light cycles different from 24 h: one was readily entrained to the light cycle and the other only loosely entrained or freeran. Additional removal of the optic lobe on the intact side resulted in a loss of the entrained component and that on the blinded side caused the reverse effect, indicating that the entrained component was driven by the pacemaker on the intact side and the other by the one on the blinded side. The synchronization between the two components was achieved only in light cycles with a limited length between 23 and 25 h. Without this range, the desynchronization of the components occurred. In the split rhythm, the phase-dependent modulation of the period of freerunning component and the mutual suppression of locomotor activity during the subjective day phase were clearly observed. The suppression was also evident in the lights-on peak that was the masking effect of light. The light cycle with dim light significantly reduced the ratio of animals with the pacemaker coupling as well as the magnitude of the period modulation. These results suggest (1) that the mutual coupling is achieved only when the difference in the periods between the two pacemakers is within an allowable range, (2) that the photic information is also involved in the mechanism of mutual coupling, and (3) that the suppression of activity occurs at the regulatory center for locomotion.Abbreviations CT circadian time - DD constant darkness - LL constant light - LD light to dark cycle - T length of light to dark cycle - freerunning period     

Mutual interactions between optic lobe circadian pacemakers in the cricket Gryllus bimaculatus

The coupling mechanism between the bilaterally paired optic lobe circadian pacemakers in the cricket Gryllus bimaculatus was investigated by recording locomotor activity, under constant light or constant red light, after the optic nerve was unilaterally severed.

A specific area of the compound eye in the cricket Gryllus bimaculatus sends photic information to the circadian pacemaker in the contralateral optic lobe

The circadian locomotor rhythm of the cricket Gryllus bimaculatus is primarily regulated by a pair of interacting optic lobe circadian pacemaker systems. The interaction involves phase-dependent modulation of the free-running period and phase-dependent suppression of activity. Since photic information has been shown to be involved in the interaction, we examined the regional difference in photoreception for the interaction within cricket compound eyes. The activity rhythm of animals receiving partial reduction of one compound eye combined with severance of the contralateral optic nerve split into entrained and free-running components under a 13-h light to 13-h dark cycle. All the animals operated on showed a phase-dependent suppression of activity, and most animals showed a phase-dependent modulation of the period of the free-running component. However, removal of the dorsocaudal area of the compound eye resulted in a severe reduction of the amplitude of the phase-dependent-period modulation. These results suggest that the dorsocaudal portion of the compound eye is a specific region receiving photic signals that are transmitted to the circadian pacemaker in the contralateral optic lobe and that the phase-dependent suppression of activity is caused by a mechanism separate from that for the period modulation.     

Unequal coupling between locomotor pacemakers of the German cockroach,Blattella germanica (L.)

Male adult German cockroaches, Blattella germanica (L.), expressed robust locomotor circadian rhythmicity under 28 degrees C and constant darkness (DD) conditions. By surgically severing the connections between the optic lobes and midbrain and their subsequent regeneration, we demonstrated that the locomotor circadian pacemaker was located in the optic lobes and that it controlled the locomotor circadian rhythm through neural pathways. From the results that unilaterally optic tract severed males still showed locomotor circadian rhythmicity (93.1%, n=29) without significantly changing the circadian period (tau) or level of motor activity, we concluded that the right and left optic lobes each contain a circadian pacemaker competent to drive the locomotor circadian rhythm. These two pacemakers were strongly coupled since only one rhythm was expressed when the pacemakers were independently exposed to opposite lighting conditions (DD or LL) at the same time. However, an unequal contribution of each pacemaker in determining the overt circadian period was found under constant dim light (10 lux) conditions, revealing a major-minor coupling relationship between these two pacemakers, so that the unilaterally blinded male expressed either a LL-rhythm with a circadian period of 24.27+/-0.21 h (41.7%, n=24) or a DD-rhythm with a circadian period of 23.43+/-0.19 h (58.3%, n=24). However, higher intensity of photic information (200-300 lux) could overpower this relationship and cause the minor pacemaker to lead the rhythm.     

The effects of pinealectomy and blinding on the circadian locomotor activity rhythm in the Japanese newt,Cynops pyrrhogaster

We examined the effects of pinealectomy and blinding (bilateral ocular enucleation) on the circadian locomotor activity rhythm in the Japanese newt, Cynops pyrrhogaster. The pinealectomized newts were entrained to a light-dark cycle of 12 h light and 12 h darkness. After transfer to constant darkness they showed residual rhythmicity for at least several days which was gradually disrupted in prolonged constant darkness. Blinded newts were also entrained to a 12 h light/12 h dark cycle. In subsequent constant darkness they showed free-running rhythms of locomotor activity. However, the freerunning periods noticeably increased compared with those observed in the previous period of constant darkness before blinding. In blinded newts entrained to the light/dark cycle the activity rhythms were gradually disrupted after pinealectomy even in the presence of the light/dark cycle. These results suggest that both the pineal and the eyes are involved in the newt's circadian system, and also suggest that the pineal of the newt acts as an extraretinal photoreceptor which mediates the entrainment of the locomotor activity rhythm.Abbreviations circadian period - DD constant darkness - LD cycle, light-dark cycle - LD 12:12 light-dark cycle of 12 h light and 12 h darkness     

Organization of the Circadian System in Insects

The circadian systems of different insect groups are summarized and compared. Emphasis is placed on the anatomical identification and characterization of circadian pacemakers, as well as on their entrainment, coupling, and output pathways. Cockroaches, crickets, beetles, and flies possess bilaterally organized pacemakers in the optic lobes that appear to be located in the accessory medulla, a small neuropil between the medulla and the lobula. Neurons that are immunoreactive for the peptide pigment-dispersing hormone (PDH) arborize in the accessory medulla and appear to be important components of the optic lobe pacemakers. The neuronal architecture of the accessory medulla with associated PDH-immunoreactive neurons is best characterized in cockroaches, while the molecular machinery of rhythm generation is best understood in fruit flies. One essential component of the circadian clock is the period protein (PER), which colocalizes with PDH in about half of the fruit fly's presumptive pacemaker neurons. PER is also found in the presumptive pacemaker neurons of beetles and moths, but appears to have different functions in these insects. In moths, the pacemakers are situated in the central brain and are closely associated with neuroendocrine functions. In the other insects, neurons associated with neuroendocrine functions also appear to be closely coupled to the optic lobe pacemakers. Some crickets and flies seem to possess central brain pacemakers in addition to their optic lobe pacemakers. With respect to neuronal organization, the circadian systems of insects show striking similarities to the vertebrate circadian system. (Chronobiology International, 15(6), 567-594, 1998)     

Circadian organization in the lizardAnolis carolinensis: a multioscillator system

Summary The locomotor activity rhythm of normal anoles freerunning in conditions of constant darkness or constant dim light typically shows a bimodal pattern. Pinealectomy of freerunning anoles renders the lizards arrhythmic (continuously active). Pinealectomy of anoles entrained to light-dark (LD) cycles often causes a large phase advance of the lizards' daily activity onsets relative to light onsets. Also, the activity onsets of individual pinealectomized anoles entrained to LD cycles often alternate between two preferred phase relationships relative to the LD cycle. These data support the hypothesis that at least two circadian oscillators (or sets of oscillators) drive the overt rhythm of locomotor activity and the pineal acts either as a circadian pacemaker or as a coupling device for these oscillators.Abbreviation SCN suprachiasmatic nuclei     

Circadian Period Modulation and Masking Effects Induced by Repetitive Light Pulses in Locomotor Rhythms of the Cricket, Gryllus bimaculatus

Effects of 15 min light pulses given at various intervals (every 1, 2, 4, 6, 8 and 12 hr) under constant darkness on the locomotor rhythm were investigated in the adult male cricket, Gryllus bimaculatus. A single pulse per 24 hr induced period modulation in a circadian phase dependent manner, yielding a period modulation curve (PMC): the 15 min light pulse lengthened the period in the early subjective night (CT11-16) and shortened it during the late subjective night to the early subjective day (CT20-5). Frequent light pulses modulated the freerunning period of the rhythm dependent on the interval of the pulses: when compared with the freerunning period in DD (23.74 +/- 0.03 hr) the period was significantly shorter in intervals of 2 and 4 hr, but lengthened when the interval was 1 and 12 hr. Frequent light pulses also resulted in entrainment of the rhythm to run with the period of 24 hr and the ratio of the entrained animals varied from 12% to 72% depending on the interval of the light pulses. The period modulation and the entrainment by the repetitive light pulses could be interpreted according to the PMC. In about 15% of animals, the light pulses induced a rhythm dissociation, suggesting that the bilaterally paired circadian pacemakers have their own sensitivity to the entraining photic information. The light pulse caused a masking effect, i.e., an intense burst of activity. The magnitude of the light induced responses was dependent on the circadian phase. The strongest masking effect was observed in the subjective night. The phase of the prominent period modulation and of the marked masking effects well coincides with the previously reported sensitive phase of the photoreceptive system.     

Pineal Gland and Circadian Locomotor Activity Rhythm in the Lacertid Gallotia galloti eisentrauti: Pinealectomy Induces Arrhythmicity

In order to contribute to a comparative view on lacertids, the effect of pinealectomy on the freerunning activity displayed under constant darkness and temperature (27.5°C ± 0.5) has been studied in the lizard Gallotia galloti eisentrauti . Animals showed an entrained motor activity rhythm under an initial light-dark (12:12 hours) routine and freerunning circadian periods ranging between 24.1 and 25.5 h during constant darkness (periodograms obtained by Sokolove & Bushell's method). After pinealectomy, most animals showed no significant circadian rhythm, their locomotor activity becoming diffuse throughout the whole 24 h period. Thus, the pineal gland seems to play an important role as a main pacemaker regulating the endogenous activity rhythm under constant conditions. This result contrasts with that found in Podarcis sicula where after pinealectomy only changes in length of the freerunning period were found.     

Kinetics in metabolic control of time measurement in photoperiodism

Abstract

The period length of the locomotor activity rhythm of Drosophila melanogaster wild form is under conditions of continuous weak red light 23.38 h, whereas die eye mutants Ly3 with a 23.71 h mean period and JK 84 with 23.14 h differ significantly. This might be due to a changed perception of light and not the result of a change in the circadian pacemaker by the mutation.

The mutant sine oculis exhibits a normal activity rhythm if the complex eyes are not completely reduced. If this is the case, the activity pattern is either less rhythmic, composed of several rhythms with different periods or truely arrhythmic depending on the individual fly.

Since the mutation in sine oculis affects in addition to the complex eye the distal part of die medulla and the lamina of the optic lobe, it is suggested that the circadian pacemakers for the locomotor activity rhythm is localized in these parts.     

Localization of the circadian pacemakers of Hemideina thoracica (Orthoptera; Stenopelmatidae)

The location of the circadian pacemakers of the orthopteran Hemideina thoracica (White) has been investigated through observation of the effects of surgical removal of brain tissues (principally optic lobes and tracts) on free-running and entrained locomotor rhythms. Bilobectomy and severance of optic tracts invariably resulted in arrhythmicity, whereas rhythmicity was sustained following unilateral lobectomy, generally with increases in the free-running period (FRP) and decreases in both the active-phase lengths and activity-to-rest ratios of the rhythm. Bilobectomized subjects could be entrained by temperature cycles, but exhibited no transients or residual rhythmicity, indicating that temperature brought about a direct response or masking effect. These results support the hypothesis that the circadian locomotor pacemakers of Hemideina are located within each optic lobe, and that there are no extraoptic centers for the control of the timing of locomotor activity. Although confirmation of the pacemaker role of the optic lobes requires transplantation of the tissues, the conclusion may be drawn by inference from other studies (e.g., Leucophaea maderae--Page, 1983; Gryllus bimaculatus--Tomioka and Chiba, 1986). Light entrainment continued after surgical binding and blackening of the compound eyes and ocelli, supporting the view that direct illumination of neural tissue through the cuticle may be one possible pathway for light entrainment.     

Isolation of an insect circadian clock

Summary Under constant conditions the compound eyes of the ground beetleAnthia sexguttata exhibit sensitivity changes in a very clear circadian rhythm. Usually the rhythms in both eyes in constant darkness are mutually coupled. After transection of the optic tract between the lobula and the supraesophageal ganglion the circadian rhythms of the two eyes continue without interruption, but coupling between them is abolished. Even if the entire supraesophageal ganglion is removed, leaving the optic ganglia intact, the circadian rhythms in the eyes continue without interruption independently. But the rhythm is abolished if the region of the lobula is damaged.The experiments show thatAnthia has circadian pacemakers in the left and right optic ganglia in or close to the lobula. These pacemakers can function independently from the rest of the brain and control circadian rhythms of physiological events.Supported by the Deutsche Forschungsgemeinschaft, Sonderforschungsbereich 45 Vergleichende Neurobiologie des Verhaltens E1     

Functional diversities of two activity components of circadian rhythm in genetical splitting mice (CS strain)

CS mice, an inbred strain, showed two distinctive characteristics in the circadian rhythm of locomotor activity: (1) large variation in the freerunning period, and (2) spontaneous rhythm splitting under continuous darkness. In the splitting rhythm there was a positive correlation between the freerunning period of the evening component and the activity time of the morning component. The phase-shifting effect of a 15-min light pulse was examined on the two activity components of the splitting rhythm. There were significant differences in the amount of light-induced phase response between the two components. A light pulse during the late subjective night induced a phase advance shift only in the morning component, while a light pulse during the early subjective night induced a phase delay shift only in the evening component. These results indicate functional diversities of the two activity components in the circadian locomotor rhythm of CS mice, and suggest that the circadian system in CS mice consists of two mutually coupled oscillators which have different circadian periods and different responsiveness to light. The CS mouse is a useful model to explore a genetic background of oscillator coupling in the circadian system of nocturnal rodents. Accepted: 19 November 1998     

Pineal-dependent locomotor activity of lamprey,Lampetra japonica,measured in relation to LD cycle and circadian rhythmicity

Summary Locomotor activity of the river lamprey, Lampetra japonica, was investigated under a light-dark (LD 1212) cycle and under continuous dark conditions. Intact lampreys were entrained to the light:dark cycle. They were active mainly in the early half of the dark period and inactive in light period. The light:dark entrainment continued in 72.7% of lampreys after the removal of bilateral eyes, but additional pinealectomy made the entrainment disappear in all lampreys. When lampreys were pinealectomized with their eyes intact, light: dark entrainment was abolished in most cases. The results indicate that the pineal organ of the lamprey is a photoreceptive organ responsible for synchronizing locomotor activity to LD cycle. Under continuous dark conditions, the locomotor activity began to free-run with a period of 21.3 ± 0.9 h (mean ± SD, n = 53). This circadian rhythmicity was not affected by the removal of lateral eyes but was abolished by pinealectomy. The pineal organ appears to function as an oscillator, or as one of the oscillators, for the circadian locomotor rhythm of lampreys.Abbreviations DD continuous dark - LD light:dark     

Circadian organization in Aplysia californica.

Substantial progress has been made in unraveling the organization of the circadian system of Aplysia californica. There are at least three circadian pacemakers in Aplysia. One has been localized in each eye and a third lies outside the eyes. Removal of the eyes disrupts the free-running locomotor activity rhythm; however, an extraocular oscillator can mediate a free-running rhythm in some eyeless animals. Although photoreceptors sufficient for entrainment of the ocular oscillator have been localized in the retina, photoreceptors outside the eyes are capable of "driving" a diurnal rhythm of locomotor activity and may also influence entrainment of ocular pacemakers. Finally, attention has been focused on the optic nerve as a coupling pathway between various parts of the system. The evidence suggests that information transmitted in the optic nerves is involved in entrainment of the ocular pacemaker by light, and in ocular control of the locomotor activity rhythm.     

Photoperiodic modulation of circadian rhythms in the cricket Gryllus bimaculatus

The waveform and the free-running period of circadian rhythms in constant conditions are often modulated by preceding lighting conditions. We have examined the modulatory effect of variable length of light phase of a 24h light cycle on the ratio of activity (alpha) and rest phase (rho) as well as on the free-running period of the locomotor rhythm in the cricket Gryllus bimaculatus. When experienced the longer light phases, the alpha/rho-ratio was smaller and the free-running period was shorter. The magnitude of changes in alpha/rho-ratio was dependent on the number of cycles exposed, while the free-running period was changed by a single exposure, suggesting that there are separate regulatory mechanisms for the waveform and the free-running period. The neuronal activity of the optic lobe showed the alpha/rho-ratio changing with the preceding photoperiod. When different photoperiodic conditions were given to each of the two optic lobe pacemakers, the alpha/rho-ratio of a single pacemaker was rather intermediate between those of animals treated with either of the two conditions. These results suggest that the storage of the photoperiodic information occurs at least in part in the optic lobe pacemaker, and that the mutual interaction between the bilateral optic lobe pacemakers is involved in the photoperiodic modulation.     

Pigment-dispersing hormone-immunoreactive neurons in the cockroach Leucophaea maderae share properties with circadian pacemaker neurons

Neurons immunoreactive with antisera against the crustacean peptide -pigment dispersing hormone fullfill several anatomical criteria proposed for circadian pacemakers in the brain of the cockroach Leucophaea maderae. These include position of somata, projections to the lamina and midbrain and possible coupling pathways between the two pacemakers through commissural fibers. In behavioral experiments combined with lesion studies and immunocytochemical investigations we examined whether the presence of pigment-dispersing hormone-immunoreactive arborizations in the midbrain of the cockroach correlates with the presence of circadian locomotor activity. No rhythm was detected after severing both optic stalks in any animal for at least 12 days. Within the same time pigment-dispersing hormone-immunoreactive fibers in the midbrain disappeared. Two to seven weeks after the operation some of the cockroaches regained circadian locomotor activity, while others remained arrhythmic. In all cockroaches which regained rhythmic behavior pigment-dispersing hormone-immunoreactive fibers had regenerated and had largely found their original targets within the brain. In all arrhythmic cockroaches either none or very little regeneration had occurred. The period of the regained circadian activity inversely correlated with the number of regenerated immunoreactive commissural fibers. These data provide further evidence for the involvement of pigment-dispersing hormone-immunoreactive neurons in circadian clocks of orthopteroid insects.Abbreviations DD constant darkness - PDH pigment-dispersing hormone - PDHLI pigmentdispersing hormone-like immunoreactivity - PDFL a pigment-dispersing factor containing cells in the lamina - PDFMe pigment-dispersing factor containing cells in the medulla - QV quantification value     

Circadian organization in the lizardSceloporus occidentalis: The effects of pinealectomy, blinding, and melatonin

Summary Pinealectomy of the iguanid lizardSceloporus occidentalis freerunning in either continuous illumination or continuous darkness typically causes changes in the period of the activity rhythm as well as changes in the amount of daily activity (). Blinding also alters the period of the freerunning activity rhythm. Continuous long term administration of melatonin via subcutaneous capsules causes a significant lengthening of the period of the activity rhythm (as well as a decrease in ) of pinealectomized and/or blinded lizards showing that melatonin exerts its action at extrapineal and extraocular sites. However, the amount of lengthening induced by melatonin is significantly greater in pinealectomized lizards than in intact lizards. The results indicate that the pineal (and possibly the eyes) act as coupling devices or as the loci of circadian pacemakers within a multioscillator system. Melatonin may function as a chemical messenger between the pineal (or eyes) and the rest of the circadian system.     

Circadian locomotor rhythms in the desert iguana I. The role of the eyes and the pineal

Summary The pineal and the eyes are known to be important components in the circadian system of some species of lizards; their effects may be mediated by the hormone melatonin. We examined the role played by these structures in the desert iguana (Dipsosaurus dorsalis). Surgical removal of the pineal had no effect on circadian locomotor rhythms, even though this procedure abolished the circadian rhythm of melatonin in the blood. Furthermore, when the isolated pineal of Dipsosaurus was studied in organ culture, it showed no circadian rhythm of melatonin secretion, as do pineals of some other lizard species, although it did produce large quantities of this hormone. Bilateral ocular enucleation had only small effects on the freerunning period of locomotor rhythms, without affecting melatonin levels in the blood. Behavioral circadian rhythms persisted in desert iguanas subjected to both enucleation and pinealectomy. These data suggest that neither the pineal nor the eyes are central components of the circadian pacemaking system in Dipsosaurus, nor is melatonin critically involved in maintaining its organization.Abbreviations CT circadian time - ZT zeitgeber time - LL constant light - LD light-dark cycle - DD constant darkness - freerunning circadian period     

Entrainment of the circadian locomotor activity rhythm in the Japanese newt by melatonin injections

We examined whether melatonin can act as a synchronizing agent within the circadian system of amphibians by testing the ability of melatonin injections to entrain the circadian locomotor activity rhythm of a newt (Cynops pyrrhogaster). Under constant darkness, all newts (13 cases) showing the free-running rhythms were subcutaneously injected with 10 g melatonin at the same time every other day for at least 30 days. Subsequently, they were injected with vehicle (1% ethanolic saline) instead of melatonin for at least another 30 days. In 10 of the 13 newts, the locomotor activity rhythms could be entrained to a period of 24 h by melatonin injections but not by vehicle injections. During the entrained steady-state, the active phase of an activity-rest cycle preceded the time of melatonin injections as previously reported in other diurnal species. These results suggest that the endogenous circadian rhythm of melatonin concentration may be involved in synchronizing circadian oscillator(s) within the newt's circadian system.