The two-oscillator circadian system of tree shrews (Tupaia belangeri) and its response to light and dark pulses

The wheel-running activity rhythm of tree shrews (tupaias; Tupaia belangeri) housed in constant darkness (DD) phase-advanced following a 3-hr light pulse at circadian time (CT) 21. Dark pulses of 3 hr presented to tupaias in bright constant light (LL) did not induce significant phase shifts of the free-running activity rhythm, irrespective of the CT. In dim LL, tupaias showed simultaneous splitting of their circadian rhythm of wheel-running activity, nest-box activity, and feeding behavior. Light pulses of 6 hr and 2300 lux were presented to 13 tupaias with split wheel-running activity rhythms. These light pulses induced immediate phase shifts in the two components of the split rhythm in opposite directions. No differences were observed between the light-pulse phase response curves of the two components. Equally large immediate phase advances were induced in both components by light pulses of 230 lux, but not by 23 lux. The final phase shifts were small at all CTs. In two tupaias, activity rhythms transiently split and re-fused. Analysis of the relative position of the components in one of these indicates asymmetry in the coupling between the components.     

Effects of induced wheel running on the circadian activity rhythms of Syrian hamsters: entrainment and phase response curve

The goal of this study was to provide an example of nonsocial and nonphotic entrainment in Syrian hamsters, together with a corresponding phase response curve (PRC). Fourteen male hamsters were given 2-hr bouts of induced activity (mostly wheel running) at 23.83-hr intervals in constant darkness (DD). The activity onsets of 10 hamsters entrained to this manipulation, with no anticipatory activity present. After entrainment, the rhythms resumed free-running from a time 0.66-3.91 hr after the onset of the last bout of induced activity. Postentrainment free-running periods were shorter than pre-entrainment values. The PRC for 2-hr pulses of induced activity in DD revealed phase advances induced in some animals between circadian time (CT) 4 and CT 11 (approximately the last half of the hamsters' rest period), and delays between CT 23 and CT 3 and between CT 17 and CT 20. The CTs for phase advances are compatible with the phase angle differences observed between rhythm and zeitgeber at the end of entrainment. Many features of the results (not all animals entraining, PRC characteristics, lack of observable anticipation to the daily stimuli, phase relationship between zeitgeber and activity rhythms) are similar to those from a previous study on social entrainment in this species (Mrosovsky, 1988). These similarities reinforce the idea that induced activity and social zeitgebers act on activity rhythms via a common mechanism.     

Circadian phototransduction: phase resetting and frequency of the circadian clock of Gonyaulax cells in red light

Constant red light (RR) influences the Gonyaulax clock in several ways: (1) Phase resetting by white or blue light pulses is stronger under background RR than in constant white light (WW); (2) frequency of the rhythm is less in RR than in WW; and (3) the amplitude of the spontaneous flashing rhythm is greater in RR than in WW. The phase response curve (PRC) to 4-hr white or blue light pulses is of high amplitude (Type 0) for cells in RR, but is of lower amplitude (Type 1) for cells in WW. In all cases, the PRC is highly asymmetrical: The magnitude of advance phase resetting is far higher than that of delay resetting. Consistent with this PRC, Gonyaulax cells in RR (free-running period greater than 24 hr) will entrain to T cycles of between 21 and 26.5 hr. The bioluminescence rhythms exhibit "masking" by blue light pulses while entrained to these T cycles. The fluence response of phase resetting to light-pulse intensity is not linear or logarithmic--rather, it is discontinuous. This feature is consistent with a limit cycle interpretation of Type 0 resetting of circadian clocks. Light pulses that cause large phase shifts also shorten the subsequent free-running period. The phase angle difference between the clock and the previous LD cycle is within 2 hr of the same phase between 16 degrees C and 25 degrees C, as determined from the light PRCs at various temperatures. Several drugs that inhibit mitochondria and/or electron transport will partially inhibit the phase shift by light.     

Phase shifts in the circadian rhythm in plasma concentrations of melatonin in rams induced by a 1-hour light pulse

The effect of a 1-hr light pulse, given at night, on the timing of the circadian rhythm in the plasma concentration of melatonin was examined in Soay rams to investigate the mechanisms involved in determining the duration of the nocturnal peak in melatonin secretion. Animals (n = 8) were housed under short days (LD 8:16) or long days (LD 16:8) and received a light pulse at various times of night. They were released into constant dim red light (DD) on day 1. Blood samples were collected hourly for 30 hr from 1000 hr on day 3, and the plasma concentration of melatonin was determined by radioimmunoassay to assess the timing of the melatonin peak. Control animals (n = 8) were maintained under the same conditions but received no light pulse. Under short days, a light pulse given early in the night caused a phase delay in the melatonin peak, and a light pulse given in the late night caused a phase advance. The mean duration of the melatonin peak was slightly reduced following a light pulse in the early or late night, and slightly increased following a pulse given near the middle of the night. Under long days, both light-pulse treatments given at night caused a phase delay in the melatonin peak, but there was no significant change in duration of the melatonin peak. The duration of the melatonin peak at day 3 under DD in the control animals was similar for all treatments, regardless of the previous entraining photoperiod (mean duration: 12.6-14.8 hr) and was similar to that under short days (14.6 hr), but was significantly longer than that under long days (8.2 hr). Information on the phase response curve in the Soay ram and on the period of the circadian oscillator governing the melatonin rhythm (c 23.0 hr under DD) predicts a close phase relationship between the end of the light phase and the onset of the melatonin peak as observed under normal 24-hr LD cycles. The current results also indicate that light acts to entrain the circadian rhythm influencing the onset and offset of melatonin secretion, and thus dictates the duration of the melatonin peak.     

Cell Division Cycles and Orcadian Oscillators in Euglena

The algal flagellate Euglena grown photoautotrophically in L:D 3:3 displays a circadian rhythm of cell division. Oscillatory models for cell cycle (CDC) control (particularly those of the limit cycle variety) include the property of phase perturbation, or resetting. This prediction has been tested in synchronous cultures in which the free-running rhythm has been scanned by 3-hr light signals. A strong (Type 0) phase response curve (PRC), yielding both advances and delays as great as 15 hr, has been derived. A second prediction of the limit cycle model is that there exists a pulse of a critical intensity, which, if given at one specific phase of the rhythm (the singularity point), should result in a phaseless, motionless state in which the rhythmicity disappears. Such a point has been found in Euglena in the late subjective night for light pulses having an intensity ranging from 40 to 700 Ix. Finally, circadian oscillators typically display temperature-compensated period lengths within the physiological range of steady-state temperatures, although the length of the CDC is commonly thought to be highly temperature dependent. We have found that over a range of at least 10°C, the period of the division rhythm is only slightly affected, exhibiting a Q₁₀ of about 1.05-1.20. These observations, therefore, collectively implicate a circadian oscillator in the control of the CDC.     

Melatonin enhances the sensitivity of circadian pacemakers to light in the nocturnal field mouse Mus booduga

The effect of exogenous melatonin (1 mg/kg) on light pulse (LP) induced phase shifts of the circadian locomotor activity rhythm was studied in the nocturnal field mouse Mus booduga. Three phase response curves (PRCs: LP, control, and experimental) were constructed to study the effect of co-administration of light and melatonin at various circadian times (CTs). The LP PRC was constructed by exposing animals free-running in constant darkness (DD) to LPs of 100-lux intensity and 15-min duration, at various CTs. The control and experimental PRCs were constructed by using a single injection of either 50% DMSO or melatonin (1 mg/kg dissolved in 50% DMSO), respectively, administered 5 min before LPs, to animals free-running in DD. A single dose of melatonin significantly modified the waveform of the LP PRC. The experimental PRC had significantly larger areas under advance and delay regions of the PRC compared to the control PRC. This was also confirmed when the phase shifts obtained at various CTs were compared between the three PRCs. The phase delays at three phases (CT12, CT14, and CT16) of the experimental PRCs were significantly greater than those of the control and the LP PRCs. Based on these results we conclude that phase shifting effects of melatonin and light add up to produce larger responses.     

The circadian system of the Turkish hamster, Mesocricetus brandti: responses to light

The circadian system of the Turkish hamster controlling wheel-running activity responded to single 1-hr light pulses and to repeated 1-hr pulses in a similar way as that of Syrian hamsters studied previously. At constant light of 100 lx, the period length (tau) of the freerunning activity rhythm of Turkish hamsters was longer and the activity time (alpha) was shorter than that of Syrian hamsters. Among individuals, the ability of the system to be entrained by one 1-hr light pulse per cycle was related to the range (advance plus delay amplitude) of the phase response curve (PRC) derived from single light pulses and to the compression of alpha caused by the pulse Zeitgeber. The data support the hypothesis derived from experiments on Syrian hamsters that the range of the PRC is functionally related with alpha, possibly reflecting the phase relations (coupling) between two oscillators.     

Is the PRC for Dark Pulses in LL a Mirror Image of the PRC for Light Pulses in DD in Mice?

The phase-response curves (PRC) for light pulses in continuous darkness (DD) have been described in many mammals, especially in nocturnal rodents. The PRC for dark pulses in continuous light (LL), however, has been described in a few mammals only, in nocturnal for bat and for hamster and in diurnal for Octodon degus, suggesting that this PRC is mirror imaging the PRC for light pulses. Therefore, the effect of 1-h and 3-h lasting dark pulses on the circadian wheel-running activity rhythm of mice in continuous light was investigated and then the PRC for dark pulses in LL was drawn up. For comparison, the effect of 1-h lasting light pulses on the circadian wheel-running activity rhythm of mice in DD was examined and the PRC for light pulses in DD was constructed. It appeared that the PRC for dark pulses, to a certain degree, represents a mirror image of the PRC for light pulses in mice. However, the advance region of this PRC is longer than that of delay. The mechanism of dark pulses action is discussed.     

Is the PRC for Dark Pulses in LL a Mirror Image of the PRC for Light Pulses in DD in Mice?

The phase-response curves (PRC) for light pulses in continuous darkness (DD) have been described in many mammals, especially in nocturnal rodents. The PRC for dark pulses in continuous light (LL), however, has been described in a few mammals only, in nocturnal for bat and for hamster and in diurnal for Octodon degus, suggesting that this PRC is mirror imaging the PRC for light pulses. Therefore, the effect of 1-h and 3-h lasting dark pulses on the circadian wheel-running activity rhythm of mice in continuous light was investigated and then the PRC for dark pulses in LL was drawn up. For comparison, the effect of 1-h lasting light pulses on the circadian wheel-running activity rhythm of mice in DD was examined and the PRC for light pulses in DD was constructed. It appeared that the PRC for dark pulses, to a certain degree, represents a mirror image of the PRC for light pulses in mice. However, the advance region of this PRC is longer than that of delay. The mechanism of dark pulses action is discussed.     

Circadian rhythm of locomotor activity in the Japanese honeybee, Apis cerana japonica

Abstract.  To reveal circadian characteristics and entrainment mechanisms in the Japanese honeybee Apis cerana japonica , the locomotor-activity rhythm of foragers is investigated under programmed light and temperature conditions. After entrainment to an LD 12 : 12 h photoperiodic regime, free-running rhythms are released in constant dark (DD) or light (LL) conditions with different free-running periods. Under the LD 12 : 12 h regime, activity offset occurs approximately 0.4 h after lights-off transition, assigned to circadian time (Ct) 12.4 h. The phase of activity onset, peak and offset, and activity duration depends on the photoperiodic regimes. The circadian rhythm can be entrained to a 24-h period by exposure to submultiple cycles of LD 6 : 6 h, as if the locomotive rhythm is entrained to LD 18 : 6 h. Phase shifts of delay and advance are observed when perturbing single light pulses are presented during free-running under DD conditions. Temperature compensation of the free-running period is demonstrated under DD and LL conditions. Steady-state entrainment of the locomotor rhythm is achieved with square-wave temperature cycles of 10 °C amplitude, but a 5 °C amplitude fails to entrain.     

Relationship between phase resetting and the free-running period in Djungarian hamsters.

A data set of 293 phase shifts was analyzed in order to determine the relationship between phase resetting and the free-running period (tau) in Djungarian hamsters. Phase shifts in response to a 15-min light pulse were assigned to one of two groups (tau short, less than 24 hr; tau long, greater than 24 hr), and two phase response curves (PRCs) were constructed. The two PRCs differed predominantly in the advance region, which extended so far into the subjective day of PRClong that a dead zone was lacking. The functional significance of PRC differences was assessed by computer simulations of entrainment to varying skeleton photoperiods and entrainment to a 12-hr skeleton photoperiod with varying tau's. Results from these simulations confirmed the theoretical predictions by Pittendrigh and Daan: Stability of entrainment under varying photoperiods depended on the ratio of the PRC slopes at the phases illuminated by light (SE/SM). This ratio was always larger than 1 for PRClong. It approached 0 for PRCshort as soon as the evening light illuminated the dead zone; this occurred for entrainment to very short photoperiods. Stability of entrainment to lights-off was in general better for PRClong than for PRCshort, especially if PRClong was used in combination with tau long. This suggests that it can be advantageous for stability of entrainment to lights-off to express a tau greater than 24 hr in combination with a PRC lacking a dead zone. Stability of entrainment under varying tau's was not much different for PRClong or PRCshort. However, stability of entrainment deteriorated for PRClong in combination with short tau's, whereas it deteriorated for PRCshort in combination with long tau's.     

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.     

Phase and period responses to short light pulses in a wild diurnal rodent,Funambulus pennanti

Photic phase response curves (PRCs) have been extensively studied in many laboratory-bred diurnal and nocturnal rodents. However, comparatively fewer studies have addressed the effects of photic cues on wild diurnal mammals. Hence, we studied the effects of short durations of light pulses on the circadian systems of the diurnal Indian Palm squirrel, Funambulus pennanti. Adult males entrained to a light–dark cycle (12?h–12?h) were transferred to constant darkness (DD). Free-running animals were exposed to brief light pulses (250 lux) of 15?min, 3 circadian hours (CT) apart (CT 0, 3, 6, 9, 12, 15, 18 and 21). Phase shifts evoked at different phases were plotted against CT and a PRC was constructed. F. pennanti exhibited phase-dependent phase shifts at all the CTs studied, and the PRC obtained was of type 1 at the intensity of light used. Phase advances were evoked during the early subjective day and late subjective night, while phase delays occurred during the late subjective day and early subjective night, with maximum phase delay at CT 15 (?2.04?±?0.23?h), and maximum phase advance at CT 21 (1.88?±?0.31?h). No dead zone was seen at this resolution. The free-running period of the rhythm was concurrently lengthened (deceleration) during the late subjective day and early subjective night, while period shortening (acceleration) occurred during the late subjective night. The maximum deceleration was noticed at CT 15 (?0.40?±?0.09?h) and the maximum acceleration at CT 21 (0.39?±?0.07?h). A significant positive correlation exists between the phase shifts and the period changes (r?=?0.684, p?=?0.001). The shapes of both the PRC and period response curve (τRC) qualitatively resemble each other. This suggests that the palm squirrel’s circadian system is entrained both by phase and period responses to light. Thus, F. pennanti exhibits robust clock-resetting in response to light pulses.     

Effects of constant darkness and constant light on circadian organization and reproductive responses in the ram

The relationship between circadian rhythms in the blood plasma concentrations of melatonin and rhythms in locomotor activity was studied in adult male sheep (Soay rams) exposed to 16-week periods of short days (8 hr of light and 16 hr of darkness; LD 8:16) or long days (LD 16:8) followed by 16-week periods of constant darkness (dim red light; DD) or constant light (LL). Under both LD 8:16 and LD 16:8, there was a clearly defined 24-hr rhythm in plasma concentrations of melatonin, with high levels throughout the dark phase. Periodogram analysis revealed a 24-hr rhythm in locomotor activity under LD 8:16 and LD 16:8. The main bouts of activity occurred during the light phase. A change from LD 8:16 to LD 16:8 resulted in a decrease in the duration of elevated melatonin secretion (melatonin peak) and an increase in the duration of activity corresponding to the changes in the ratio of light to darkness. In all rams, a significant circadian rhythm of activity persisted over the first 2 weeks following transfer from an entraining photoperiod to DD, with a mean period of 23.77 hr. However, the activity rhythms subsequently became disorganized, as did the 24-hr melatonin rhythms. The introduction of a 1-hr light pulse every 24 hr (LD 1:23) for 2 weeks after 8 weeks under DD reinduced a rhythm in both melatonin secretion and activity: the end of the 1-hr light period acted as the dusk signal, producing a normal temporal association of the two rhythms. Under LL, the 24-hr melatonin rhythms were disrupted, though several rams still showed periods of elevated melatonin secretion. Significant activity rhythms were either absent or a weak component occurred with a period of 24 hr. The introduction of a 1-hr dark period every 24 hr for 2 weeks after 8 weeks under LL (LD 23:1) failed to induce or entrain rhythms in either of the parameters. The occurrence of 24-hr activity rhythm in some rams under LL may indicate nonphotoperiodic entrainment signals in our experimental facility. Reproductive responses to the changes in photoperiod were also monitored. After pretreatment with LD 8:16, the rams were sexually active; exposure to LD 16:8, DD, or LL resulted in a decline in all measures of reproductive function. The decline was slower under DD than LD 16:8 or LL.(ABSTRACT TRUNCATED AT 400 WORDS)     

A phase response curve for circannual rhythm in the varied carpet beetle <Emphasis Type="Italic">Anthrenus verbasci</Emphasis>

We know that entrainment, a stable phase relationship with an environmental cycle, must be established for a biological clock to function properly. Phase response curves (PRCs), which are plots of phase shifts that result as a function of the phase of a stimulus, have been created to examine the mode of entrainment. In circadian rhythms, single-light pulse PRCs have been obtained by giving a light pulse to various phases of a free-running rhythm under continuous darkness. This successfully explains the entrainment to light-dark cycles. Some organisms show circannual rhythms. In some of these, changes in photoperiod entrain the circannual rhythms. However, no single-pulse PRCs have been created. Here we show the PRC to a long-day pulse superimposed for 4 weeks over constant short days in the circannual pupation rhythm in the varied carpet beetle Anthrenus verbasci. Because the shape of that PRC closely resembles that of the Type 0 PRC with large phase shifts in circadian rhythms, we suggest that an oscillator having a common feature in the phase response with the circadian clock, produces a circannual rhythm.     

Light pulses entrain the circadian activity rhythm of a diurnal rodent (Ammospermophilus leucurus)

Circadian rhythms of wheel-running activity of the antelope ground squirrel (Ammospermophilus leucurus) were entrained by light-dark cycles (LD: 100 1x vs total darkness) with periods (T) between ca 23.75 and 24.75 hr. Two 1-hr light pulses per cycle ('skeleton photoperiods') with T = 24.25 hr as well as one 1-hr light pulse per cycle with Ts of 23.75 and 24.25 hr were effective in entraining the circadian activity rhythms in at least 50% of the antelope ground squirrels. Phase and period responses to single 1-hr light pulses were measured which depend on the initial phase and period of the rhythm. It is concluded that discrete (phasic) light input contributes to the mechanism of entrainment to LD cycles in diurnal rodents.     

Circadian Rhythm of Blood Ethanol Clearance Rates in Rats: Response to Reversal of the L/D Regimen and to Continuous Darkness and Continuous Illumination

Phase relationships of the circadian rhythms of blood ethanol clearance (metabolic) rates and body temperature were studied in rats successively exposed to 4 illumination regimens: LD (light from 0800-2000 hr), DL (light from 2000-0800 hr), constant darkness (DD) and, lastly, constant light (LL). After a 4-wk standardization to each regimen, body temperatures were taken at 9 × 4-hr intervals to establish baseline circadian profiles. One week later, groups (N = 8) received 1.5 g/kg ethanol (i.p.) at 6 equally spaced timepoints during a 24-hr span, when temperatures were again measured. Ethanol clearance rates were estimated from decreasing blood ethanol levels sampled every 20 min from 60-200 min after dosing, and the resultant elimination curves were subjected to cosinor analysis. These studies show for the first time that the high amplitude circadian rhythm in ethanol metabolism persists under constant conditions of illumination (DD and LL), demonstrating that it may well be a truly internal circadian rhythm and not a response to exogenous cues of the light/dark cycle. During both LD and DL, maximal and minimal ethanol clearance rates fell near the end of the dark and light phases, respectively, and followed circadian peak and trough control temperatures by approximately 6 hr. A fixed internal phase relationship between the core body temperature and the circadian rhythm in ethanol metabolism is demonstrated, thus establishing the rhythm in body temperature as a suitable and convenient internal marker rhythm for studies of the metabolism of low-to-moderate ethanol doses. These studies demonstrate that the phase relationships of blood ethanol clearance rate and body temperature can be manipulated by the illumination regimen selected, an observation of both basic and practical importance.     

Psi-Mutation Affects Phase Angle Difference, Free-Running Period and Phase Shifts in Aedes krombeini (Stegomyia)

A new mutation, designated as psi-mutant, affecting the timing of the circadian oviposition rhythm was discovered the in natural population of Aedes krombeini . This mutation advanced the phase of the oviposition median in an entraining light-dark cycle of 12:12 h by ca. 7.0 h and shortened the free running period t in constant darkness (DD) by ca. 4.0 h. Early oviposition in psi-mutants was also observed when while free-running in DD they were subjected to 24-h temperature cycles (29°C for 12 h and l8°C for l2 h). When the phase response curves (PRCs) for light pulses against DD as background were measured, the PRC for the psi-mutant had large delaying phase shifts, whereas, that of the wild strain had small delaying phase shifts.     

Psi-Mutation Affects Phase Angle Difference,Free-Running Period and Phase Shifts in Aedes krombeini (Stegomyia)

A new mutation, designated as psi-mutant, affecting the timing of the circadian oviposition rhythm was discovered the in natural population of Aedes krombeini. This mutation advanced the phase of the oviposition median in an entraining light-dark cycle of 12:12 h by ca. 7.0 h and shortened the free running period t in constant darkness (DD) by ca. 4.0 h. Early oviposition in psi-mutants was also observed when while free-running in DD they were subjected to 24-h temperature cycles (29°C for 12 h and l8°C for l2 h). When the phase response curves (PRCs) for light pulses against DD as background were measured, the PRC for the psi-mutant had large delaying phase shifts, whereas, that of the wild strain had small delaying phase shifts.     

Nonphotic enhancement of adjustment to new light-dark cycles: masking interpretation discounted

The adjustment of hamsters to advanced light-dark (LD) cycles can be greatly accelerated by scheduling a single 3-hr bout of extra activity in a novel running wheel, starting about 7 hr before the time when the animals become active in the preceding LD cycle. The present experiments were designed to provide stronger evidence that this effect depends on a shift in the pacemaker rather than on masking. It was shown that when hamsters were put into continuous darkness (DD) 1 day after the exercise-accelerated phase shift, their free-running rhythms took off from a time nearer to the onset of darkness in the new LD cycle than in the preceding LD cycle. An incidental finding was that in DD the free-running period of the hamsters with the accelerated phase shifts was longer than that of the control animals. Further evidence that the 3-hr exercise pulse had produced a greater phase advance than that occurring in undisturbed control animals was obtained by giving a light pulse at the same clock time to all animals after they had been in DD for 8 days. The animals that had previously exercised for the additional 3-hr phase-advanced in response to the light pulse, while the undisturbed control animals phase-delayed.