Kenji Tomioka

Chronobiological analysis of a new clock mutant, Toki, in Drosophila melanogaster.

We have isolated a new semidominant clock mutant Toki on the second chromosome in Drosophila melanogaster. This mutant differs from the wild-type Canton-S in several properties as follows. Larger values are obtained in the phase angle difference (phi, the time from lights-off in a 24-hr light-dark cycle to an activity offset), the ratio of activity time to rest time (alpha/rho) and the activity level. The free-running period (tau) is 25.3 hr, one hour longer than in the wild-type. In the phase response curve (PRC), the ratio between the delay and the advance portion is larger and the cross-over point occurs later, although there is no difference in amplitude of the mutants PRC (Type I). The rhythm is more sensitive to the light intensity, becoming obscure in darker condition. Toki interacts with other clock mutations, pers, perL and And, in such a way that tau s associated with these three X-linked mutations are lengthened and phi values become smaller.

Involvement of serotonin in the circadian rhythm of an insect visual system

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

SummaryThe 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.1.The majority (about 70%) of the animals showed a locomotor rhythm with 2 rhythmic components; one freerunning with a period of 25.33 ± 0.41 (SD) h and the other with 24.36 ± 0.37 (SD) h under constant light (Fig. 3A).2.Removal of the intact side optic lobe abolished the longer period component (Fig. 4A), while the operation on the operated side caused a reverse effect (Fig. 4B), indicating that the longer and the shorter period components are driven by the pacemaker on the intact and the operated side, respectively.3.The activity driven by a pacemaker was inhibited during the subjective day of the contralateral pacemaker (circadian time 0–10, Fig. 5).4.The freerunning periods of the two components were not constant but varied as a function of the mutual phase angle relationship (Figs. 3A, 7, 8).nThese results suggest that the 2 optic lobe pacemakers weakly couple to one another and that the cricket maintains a stable temporal structure in its behavior through the phase-dependent mututal inhibition of activity and the phase-dependent freerunning period modulation.

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.

Persistence of circadian ERG rhythm in the cricket with optic tract severed

We report here that the cricket Gryllus bimaculatus showed a circadian ERG rhythm in constant darkness (DD) and temperature (26 ~ even after the optic tracts (OT) were severed. This is the first detection, in insects, of circadian rhythm from the optic lobe (OL)-compound eye (CE) system neurally isolated from the central nervous system (CNS). The significance of this finding is not confined to the physiology of ERG rhythm but extends potentially to the physiology of locomotor rhythm. This cricket is diurnaIly active in the nymphal stage but becomes nocturnal 4 to 5 days after the imaginal molt [11]. Adult males which had become fully nocturnal were obtained from a laboratory stock kept in LD 12:12 (L: 0600-1800) and at a constant temperature of 26 ~ the standard environmental condition. The crickets, whose legs were cut off, were fixed to a supporting rod and Ag-AgC1 wire electrodes were chronically implanted into the immediate vicinity of the receptor layer of the compound eyes. The apparatus was so arranged that ERGs elicited by a 50 ms flash of green light at intervals of 1 h were recorded automatically. The stimulus intensities were always held below saturation for the ERG. The OT is relatively long in the cricket, connecting medulla to lobula [4]. A small square piece of head capsule was removed to make a window, through which the OT was cut with microscissors. Strict attention had to be paid to minimize damage to tracheae running into the OL. Finally, the piece of head capsule was replaced to close the window, and the wound was sealed with vaseline. The ERG was diphasic, composed of onand off-components. Amplitudes of the two components showed circadian rhythm in phase with each other. Hereafter, we refer to the peak-to-trough amplitude of the diphasic wave, in other words, the summation of amplitudes of onand off-components. In intact animals, the ERG amplitude changed synchronously with LD 12:12, peaking in the dark fraction. This rhythm freeran in the ensuing DD with a period (tau) a little shorter than 24 h. When the OT was cut unilaterally, the 4 crickets all exhibited this rhythm in both intact and operated eyes (Fig. 1). Moreover, in 2 of 3 animals in which the OT were bilaterally severed, the rhythm was obvious in both eyes under DD (Fig. 2). The remaining animal showed the rhythm only in one eye; the other eye was not rhythmic probably due to damage to tracheae during the operation. In both cases of unilateral and bilateral cuts, there was a subtle difference between taus of the two eyes obtained by the periodogram. However, since data are limited, it is premature to discuss bilateral organization of the rhythm of the two eyes as was done elegantly in the beetle [5].

Neural pathways involved in mutual interactions between optic lobe circadian pacemakers in the cricket Gryllus bimaculatus

The circadian locomotor rhythm of the cricket Gryllus bimaculatus is primarily generated by a pair of optic lobe circadian pacemakers. The two pacemakers mutually interact to keep a stable temporal structure in the locomotor activity. The interaction has two principal effects on the activity rhythm, i.e., phase-dependent modulation of the freerunning period and phase-dependent suppression of activity driven by the partner pacemaker. Both effects were mediated by neural pathways, since they were immediately abolished after the optic stalk connecting the optic medulla to the lobula was unilaterally severed. The neural pathways were examined by recording locomotor activity, under a 13 h light to 13 h dark cycle, after the optic nerves were unilaterally severed and the contralateral optic stalk was partially destroyed near the lobula. When the dorsal half of the optic stalk was severed, locomotor rhythm mostly split into two components: one was readily entrained to the given light-dark cycle and the other freeran with a marked fluctuation in freerunning period, where the period of the freerunning component was lengthened or shortened when the onset of the entrained component occurred during its subjective night or day, respectively. The phase-dependent modulation of activity was also observed in both components. However, severance of the ventral half of the optic stalk resulted in appearance only of the freerunning component; neither the phase-dependent modulation of its freerunning period nor the change in activity level was observed. These results suggest that neurons driving the mutual interaction and the overt activity rhythm run in the ventral half of the proximal optic stalk that includes axons of large medulla neurons projecting to the cerebral lobe and the contralateral medulla.

Light Cycles Given During Development Affect Freerunning Period of Circadian Locomotor Rhythm of period Mutants in Drosophila melanogaster.

We reared wild type (Canton-S) and period mutant flies, i.e., per(S) and per(L), of Drosophila melanogaster in constant darkness, constant light or 24h light dark cycles with various light to dark ratios throughout the development from embryo to early adult. The locomotor activity rhythms of newly eclosed individuals were subsequently monitored in the lighting conditions, in which they had been reared, for several days and then in constant darkness. Circadian rhythms were clearly exhibited in constant darkness even in flies reared in constant light and constant darkness as well as flies reared in light-dark cycles, but the freerunning period differed among groups. The results suggest that the circadian clock is assembled without any cyclical photic information, and that the light influences the developing circadian clock of Drosophila to alter the freerunning period. The effects of light on the rhythm differed in some aspects between per(L) flies and the other two strains. Possible mechanisms through which light affects the developing circadian clock are discussed. Copyright 1997 Elsevier Science Ltd. All rights reserved

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

Abstract 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.

Electrical properties of the cerebral prothoracicotropic hormone cells in diapausing and non-diapausing pupae of the tobacco hornworm, Manduca sexta

Abstract Prothoracicotropic hormone (PTTH) is an insect brain neuropeptide that is a primary factor regulating an insect development. Curtailment of its release is thought to be responsible for the pupal diapause of tobacco hornworm, Manduca sexta. The cell synthesizing and secreting the PTTH has been identified as a pair of neurosecretory cells in the pars lateralis on each brain hemisphere. Using intracellular recording techniques, we have demonstrated electrical properties of the PTTH cells in different physiological status, i.e., diapausing and developing pupae. In diapausing pupae, they showed threshold value increasing and input resistance decreasing with the progress of diapausing state, indicating that they were getting unexcitable. Spontaneous action potentials and excitatory postsynaptic potentials (EPSPs) were rarely observed in deeply diapausing state. Non-diapausing PTTH cells were almost silent except day-2, showing rather constant values of electrical properties. On day-2, a significant proportion of the cells had spontaneous action potentials, showing less negative membrane potential values than inactive cells. Exclusively inhibitory postsynaptic potentials (IPSPs) were observed in significant numbers of the cells during the period from day-2 to day-5. On the basis of the results obtained, we proposed a working hypothesis that electrical activities of the PTTH cell may be primarily regulated by its membrane properties which are further modulated by the synaptic mediation.

Analysis of postembryonic development of locomotor activity rhythm by corpora allata implantation in the cricket gryllus bimaculatus

Abstract 1. 1. Male crickets Gryllus bimaculatus show a drastic change in circadian rhythm from nymphal diurnality to adult nocturnality, in association with an increase in activity level several days after the imaginai moult. 2. 2. The corpora allata implantation into male 7th or 8th instar nymphs produced supernumerary instar nymphs in about 30% of the implanted animals, but did not affected the normal development in the remaining animals. 3. 3. The majority of the supernumerary instar nymphs were diurnal and sexually inactive, although their internal reproductive organs appeared to be fully mature. 4. 4. The supernumerary instar nymphs became nocturnal with an increase in activity level several days after the imaginai (9th) moult. 5. 5. The roles of the nervous system in the regulation of the rhythm reversal are discussed.