Diazepam phase shifts the circadian clock of the field mouseMus booduga

Single 2h administration of diazepam (benzodiazepine) in 3.5% ethanol solution was found to evoke advance and delay phase shifts in the locomotor activity rhythm in the field mouseMus booduga. Through such pulsed administration of diazepam at various phases of circadian rhythm a phase response curve could be constructed. Phase advance occurred during early subjective day (CT 2) and phase delays were observed in the remaining phases. The shape of the diazepam phase response curve is similar to the general shape of the phase response curves generated by intraperitoneal injections of other benzodiazepines in hamsters. The phase shifting action of diazepam may be explained by its agonistic action on the neurotransmitter gamma-aminobutyric acid.     

Intensity-dependent phase-adjustments in the locomotor activity rhythm of the nocturnal field mouse Mus booduga

The locomotor activity rhythm of the nocturnal field mouse Mus booduga was monitored under constant darkness (DD) and free-running periods (tau) were estimated. Following a free-run of about 15 days in DD, the animals were exposed to periodic light pulses (LPs) of various intensities (1 lux, 10 lux, 50 lux, 100 lux, and 1,000 lux) and 15 minutes duration for 65 days at intervals of 24 hours to investigate the influence of intensity of light on the phase-angle-difference (psi) between the onset of locomotor activity and the time of LP administration. The experimentally observed values of psi and tau for a LP of 1,000 lux intensity used for 15 minutes every 24 hr, showed a sigmoid shaped relationship with tau. This relationship was similar to that predicted based on the nonparametric model of entrainment, which uses the tau and the LP phase response curve (PRC) constructed using LP of similar duration and intensity. The functional nature of the relationship between psi and tau was not found to change significantly with increasing intensities of LP used to entrain the locomotor activity rhythm. However, psi was significantly modulated by the intensity of LP. These results suggest that the periodic sensitivity of the circadian pacemaker underlying the locomotor activity rhythm in the nocturnal field mouse M. booduga to LPs plays an important role in maintaining a characteristic psi with the zeitgeber and the psi changes in a light intensity-dependent manner.     

Chronobiotic effect of melatonin following phase shift of light/dark cycles in the field mouseMus booduga

The objective of this study was to assess whether melatonin accelerates the re-entrainment of locomotor activity after 6 h of advance and delay phase shifts following exposure to LD 12:12 cycle (simulating jet-lag/shift work). An experimental group of adult male field mice Mus booduga were subjected to melatonin (1 mg/kg) through i.p. and the control group were treated with 50 % DMSO. The injections were administered on three consecutive days following 6h of phase advance and delay, at the expected time of “lights off”. The results show that melatonin accelerates the re-entrainment after phase advance (29%) when compared with control mice. In the 6 h phase delay study, the experimental mice (melatonin administered) take more cycles for re-entrainment (51%) than the control. Further, the results suggest that though melatonin may be useful for the treatment of jet-lag caused by eastward flight (phase advance) it may not be useful for westward flight (phase delay) jet-lag     

Irradiance dependency of UV-A induced phase shifts in the locomotor activity rhythm of the field mouse Mus booduga

In the nocturnal field mouse Mus booduga, the responsiveness of the circadian system to UV-A light of 2.5 W/m² and 30 minutes duration is known to be phase dependent. The results of our experiments indicate that the phase shifts evoked by UV-A at the two phases, CT14 (circadian time 14) and CT20 increases nonlinearly with irradiance. (Chronobiology International, 17(6), 777-782, 2000)     

Entrainment of circadian locomotor activity rhythm of the nocturnal field mouse Mus booduga using daily injections of melatonin

In this paper, we report the effects of daily injections of melatonin on the locomotor activity rhythm of the nocturnal field mouse Mus booduga. The locomotor activity rhythm of 45 animals was first monitored in constant darkness (DD) of the laboratory for about 15 days. The animals were then divided into three groups (experimental, vehicle-treated control, and the nontreated control groups) and subjected to three different treatments. The animals from the experimental group (n=19) were administered daily a single subcutaneous (s.c.) injection of melatonin (1 mg/kg) for about 45 days. The vehicle treated controls (n=13) were administered daily injections of 50% dimethyl sulfoxide (DMSO) for about 45 days, and the nontreated controls (n=13) were handled similar to the other two groups without being administered injections. Following the treatments, the animals were maintained in DD for about 20 days, after which the experiments were terminated. A significantly larger percentage of animals from the experimental group either entrained or showed phase control to daily treatments, compared to the animals from the two control groups. These results suggest that externally administered melatonin can influence the phase of the circadian locomotor activity rhythm of M. booduga. The fact that none of the nontreated controls showed any sign of phase control to daily handling, clearly demonstrates that the entrainment or phase control in the melatonin treated group of animals is caused by melatonin alone and not due to handling.     

Restricted feeding-induced entrainment of activity rhythm and peripheral clock rhythm

Sleep and Biological Rhythms - Daily restricted feeding entrains the circadian rhythm of mouse clock gene expression in the central nervous system excluding the suprachiasmatic nucleus, as well as...     

Influence of the corticosterone rhythm on photic entrainment of locomotor activity in rats

The question of involvement of glucocorticoid hormones as temporal signals for the synchronization of the timekeeping system was addressed in rats with different corticosterone status. The authors showed that adrenalectomy had no effects on the synchronization of wheel-running activity rhythms to a steady-state LD 12:12 cycle, regardless of whether it was compensated for by a corticosterone replacement therapy that either reinstated constant plasma concentrations of the hormone or mimicked its natural rhythm. However, after a 12-h phase shift (daylight reversal), the lack of circulating corticosterone induced a significant shortening of the resynchronization rate (less than 3 days vs. 7 days). Normalization required restoration of a rhythmic corticosterone secretion that was synchronized to the new photoperiod. Under constant darkness, the corticosterone rhythm did not show any synchronizing effect, providing evidence that it participates in entrainment of the locomotor activity rhythm through modulation of light effects. It is proposed that, under stable lighting conditions, circulating glucocorticoids contribute to stabilizing activity rhythms by reinforcing resistance of the circadian timing system to variations of the photoperiod. Experimental evidence that serotonergic neurons are involved in relaying their modulatory effects to the clock is also presented.     

Screening for novel ENU-induced rhythm, entrainment and activity mutants

Chemical mutagenesis has provided an opportunity to develop and expand the repertoire of behavioural mutants for gene function studies. With this in mind, we have established a screen in mice for mutations affecting circadian rhythms, entrainment to light and other wheel-running parameters. The screen consists of an assessment of mouse wheel-running activity in a 12:12 h light/dark cycle for 7-10 days followed by assessment in constant darkness for up to 20 days. Responses to light are assessed using two protocols; a 15 minute light pulse given at circadian time 16 on the tenth day in constant darkness and an additional 12 h of light upon transition from light/dark conditions to constant darkness. To date, approximately 1300 progeny of chemically mutagenised mice have been screened. Computer-aided assessment of wheel-running parameters has helped in identifying abnormal phenotypes in approximately 5% of all animals screened. Inheritance testing of mice with abnormal phenotypes has confirmed the number of robustly inherited mutant phenotypes to be 1% of the total screened. Confirmed mutants including those affecting free-running period, light-responsiveness and wheel-running endurance have been identified. Thus far, low-resolution map positions have been established for four mutants by completing genome scans in backcross progeny. Mutant loci do not correspond with those previously associated with wheel-running behaviour. This result confirms that phenotype-driven approaches such as this should continue to provide material for mammalian gene function studies.     

History-dependent changes in entrainment of the activity rhythm in the Syrian hamster (Mesocricetus auratus)

The authors have studied the activity rhythm of Syrian hamsters exposed to square LD cycles with a 22-h period (T22) with the aim of testing the effects of the previous history on the rhythmic pattern. To do so, sequential changes of different lighting environments were established, followed by the same LD condition. Also, the protocol included T22 cycles with varying lighting contrasts to test the extent to which a computational model predicts experimental outcomes. At the beginning of the experiment, exposure to T22 with 300 lux and dim red light occurring respectively at photophase and scotophase (LD300/dim red) mainly generated relative coordination. Subsequent transfer to cycles with approximately 0.1-lux dim light during the scotophase (LD300/0.1) promoted entrainment to T22. However, a further reduction in light intensity to 10 lux during the photophase (LD10/0.1) generated weak and unstable T22 rhythms. When, after that, animals were transferred again to the initial LD300/dim red cycles, the amplitude of the rhythm still remained very low, and the phases were very unstable. Exposure to constant darkness partially restored the activity rhythm, and when, afterwards, the animals were submitted again to LD300/dim red cycles, a robust T22 rhythm appeared. The results demonstrate history-dependent changes in the hamster circadian system because the locomotor activity pattern under the same T22 cycle can show relative coordination or unstable or robust entrainment depending on the prior lighting condition. This suggests that the circadian system responds to environmental stimuli depending on its previous history. Moreover, computer simulations allow the authors to predict entrainment under LD300/0.1 cycles and indicate that most of the patterns observed in the animals due to the light in the scotophase can be explained by different degrees of coupling among the oscillators of the circadian system.     

Effects of photoperiod on rat motor activity rhythm at the lower limit of entrainment

The experiment described here studied the rat motor activity pattern as a function of the photoperiod of circadian light-dark cycles in the limits of entrainment (22-and 23-h periods). In most cases, the overt rhythm showed 2 circadian components: 1 that followed the external LD cycle and a 2nd rhythm that was free run. The expression of these components was directly dependent on the photoperiod, and there was a gradual transition in the manifestation of 1 or the other. The component with a period equal to that of the external cycle was more manifested under long photoperiods, while the other 1 was more expressed during short photoperiods. Also, the period of the free-running component was longer under T22 than T23. For each period, the free-running component was longer under a longer photoperiod. At first sight, the presence of these 2 components in most of the rats might appear to be due to the fact that in the limits of entrainment, some rats do not entrain and thus show a free-running rhythm plus masking. However, the gradation observed in the different patterns of the overt motor activity rhythm, especially those patterns related to the different balance between the 2 components and the length of the period of the free-running component under LD as a function of the photoperiod, suggests that the circadian system can be functionally dissociated.     

Circadian rhythm entrainment in flies and mammals

Circadian rhythms are a fundamental adaptation of living cells to the daily and seasonal fluctuation in light and temperature. Circadian oscillations persist in constant conditions; however, they are also phase-adjusted (entrained) by day-night cycles. It is this entrainability that provides for the proper phasing of the program, to the sequence of external changes that it has evolved to exploit. Synchronization of circadian oscillators with the outside world is achieved because light, temperature, or other external temporal cues, have acute effects on the levels of one or more of the clock's components. The consequences are ripples through the interconnected molecular loops, leading to a stable phase realignment of the endogenous rhythm generator and the external conditions. This review summarized the evolving knowledge of the different types, modes, and molecular processes of entrainment in flies and mammals.     

Dynamics of discrete entrainment of the circadian rhythm in the rat pineal N-acetyltransferase activity during transient cycles

To elucidate entrainment of a pacemaker controlling the N-acetyltransferase (NAT) rhythm in the rat pineal gland, we studied the phase response curves (PRCs) of this rhythm. We exposed 50- to 60-day-old male Wistar rats maintained in a light-dark cycle (LD 12:12) to a 1-min light pulse at different times before midnight or at various times throughout the whole night. We then released them into constant darkness and studied the morning NAT decline during the night when rats were pulsed before midnight, as well as the evening NAT rise and the morning decline after 4 days following the pulses. The PRC for the first NAT decline and the PRCs for the NAT rise and decline after 4 days were compared with published transient PRCs (Illnerová and Van?cek, 1982b), in order to obtain a complete picture of the dynamics of the NAT rhythm entrainment during the transient cycles. Phase delays in the NAT rise due to a pulse before midnight were complete (i.e., identical to those of day 4) on day 1. Phase delays in the NAT decline were almost complete on day 1, while incomplete phase delays were observed on day 0. Phase advances in the NAT rise and decline due to a pulse past midnight had different dynamics: Advances in the decline were complete on day 1, while advances in the rise were absent on day 1 and much smaller than in the decline on day 4. The results are discussed in terms of a two-component (E-M) pacemaker controlling the NAT rhythm. The NAT rise may reflect the phase of the E-component, while the decline reflects the M-component. Phase delays of the E-component are accomplished within one cycle, and so are phase advances of the M-component. However, although delays of E already result in delays of M one cycle after the pulse, it takes several transient cycles before advances of M begin to induce advances of E.     

Phase sensitivity analysis of circadian rhythm entrainment

As a biological clock, circadian rhythms evolve to accomplish a stable (robust) entrainment to environmental cycles, of which light is the most obvious. The mechanism of photic entrainment is not known, but two models of entrainment have been proposed based on whether light has a continuous (parametric) or discrete (nonparametric) effect on the circadian pacemaker. A novel sensitivity analysis is developed to study the circadian entrainment in silico based on a limit cycle approach and applied to a model of Drosophila circadian rhythm. The comparative analyses of complete and skeleton photoperiods suggest a trade-off between the contribution of period modulation (parametric effect) and phase shift (nonparametric effect) in Drosophila circadian entrainment. The results also give suggestions for an experimental study to (in)validate the two models of entrainment.     

Temperature entrainment of the circadian cuticle deposition rhythm in Drosophila melanogaster

The cuticle deposition rhythm, which is observed in the apodeme of the furca in the thorax, is controlled by a peripheral circadian clock in the epidermal cells and entrained to light-dark (LD) cycles via CRYPTOCHROME (CRY) in Drosophila melanogaster. In the present study, we examined the effects of temperature (TC) cycles and the combination of LD and TC cycles on entrainment of the cuticle deposition rhythm. The rhythm was entrained to TC cycles, whose period was 28 h. In T = 21 and 24 h, the rhythm was entrained to TC cycles in some individuals. CRY is not necessary for temperature entrainment of the cuticle deposition rhythm because the rhythm in cry(b) (lacking functional CRY) was entrained to TC cycles. Temperature entrainment of the rhythm was achieved even when the thoraxes or furcae were cultured in vitro, suggesting that the mechanism for temperature entrainment is independent of the central clock in the brain and the site of the thermoreception resides in the epidermal cells. When LD and TC cycles with different periods were applied, the rhythm was entrained to LD cycles with a slight influence of TC cycles. Thus, the LD cycle is a stronger zeitgeber than the TC cycle. The variance of the number of the cuticle layers decreased in the flies kept under LD and TC cycles with the same period in which the thermophase coincided with the photophase. Therefore, we conclude that LD and TC cycles synergistically entrain the rhythm. Synergistic effects of LD and TC cycles on entrainment were also observed even when the thoraxes were cultured in vitro, suggesting that the light and temperature information is integrated within the peripheral circadian system.     

Photic entrainment of daily rhythm pattern of locomotor activity in sea bass (Dicentrarcus labrax)

The aim of this study was to evaluate the circadian rhythm of daily locomotor activity in sea bass exposed to three different artificial lighting regimes including 12/12, 24/0, and 0/24 L/D periods, and scheduled daytime feeding. Daily rhythm of locomotor activity during 12/12 L/D was observed, whereas locomotor activity recorded during 24/0 and 0/24 L/D resulted arrhythmic. Statistical differences in mesor values among the photoperiods and between light and dark phases of each photoperiod were found. During the 12/12 and 0/24 L/D, the fish were active mostly during the light phase. During 24/0 L/D, a phase inversion occurred. The results showed that photoperiod is a dominant synchronizer in sea bass, and that the appetite in this fish follows daily rhythms which does not match the imposed feeding schedule, suggesting the absence or the lability of internal process that couples feeding cycles and biological rhythms.     

The effect of pinealectomy on entrainment of the locomotor activity rhythm in sparrows maintained on various short days

Summary Pinealectomy of house sparrows on 3L:21D (3 h light per 24 h) resulted in a significant increase in the time between the onset of perch-hopping activity and lights on (on) as well as the time between the offset of activity and lights of (off). The daily variance in on and off was also increased following the removal of the pineal gland. On longer light cycles (i.e., 5L:19D; 7L:17D), neither on or off, nor the variance of on or off was different between sham-pinealectomized and pinealectomized sparrows. Upon returning the birds to an ultrashort light cycle, 1L:23D, off, as well as the variance in on and off were again found to be significantly larger in the pinealectomized birds when compared to sham-operated controls. These results indicate that the effects of pinealectomy on the entrained rhythm of locomotor activity are most pronounced when birds are exposed to a weak entraining agent, such as an ultrashort light: dark cycle. In view of the observation that pinealectomy can alter the phase relationship between activity onset and offset, it is suggested that the pineal gland may be involved in the coupling of the oscillators that regulate activity onset and offset.     

The entrainment of ventilation frequency to exercise rhythm

To investigate whether ventilation frequency could be entrained to a sub-harmonic of the exercise rhythm, 19 experimentally naive male volunteers were tested during steady state bicycle ergometry and arm cranking under conditions of constant applied workload. Each exercise was performed at two separate ventilatory loads, one within the linear range and the other in the curvilinear range of ventilatory response to exercise. A preferred exercise rhythm was initially adopted (4 min.) followed by forced incremented and decremented rhythm changes each lasting 3 min during a 12 min exercise period. Ventilation, pedal pulse train and heart rate were sampled at 17 Hz on a PDP 11/23 computer. Ratios of limb frequency to dominant respiratory frequency were determined following Fourier analysis of these signals. Data that lay within +/- 0.05 of an integer and half-integer ratio were accepted as indices of entrainment, provided that the observed entrained scores were statistically significant. Ventilation frequency showed a clear, but intermittent tendency to entrain with limb frequency. This tendency was greater during bicycle ergometry, possibly as a consequence of task familiarisation, although both exercise entrainments were independent of workload. No difference between preferred versus varied exercise rhythm was evident, but more entrainment (p less than 0.01) was observed during a decremental change in exercise rhythm. These responses do not appear to support an appreciable role for limb-based afferents in the control of entrainment. The results of this study provide evidence that exercise rhythm has some regulatory role in the control of breathing during moderate rhythmical laboratory-based exercise ergometry.     

Light and temperature entrainment of a locomotor rhythm in honeybees

Abstract. The circadian locomotor (walking) rhythms of forager honeybees (Apis mellifera ligustica L.) were entrained to eight different 24 h light-dark cycles. The phases of activity onset, peak activity, and offset were correlated with the lights-off transition, suggesting lights-off as the primary zeitgeber for the rhythm. Further support for this hypothesis was provided by LD 1:23 experiments, in which entrainment occurred when the light pulse was situated at the end, but not at the beginning, of the subjective photophase. Steady-state entrainment of the locomotor rhythm was achieved with square-wave temperature cycles of 10^oC amplitude under constant dark: most of the activity occurred within the early thermophase. Smaller amplitude temperature cycles yielded relative coordination of the rhythm. Interactions of temperature and light-dark cycles resulted in entrainment patterns different from those elicited in response to either cycle alone or those formed by a simple combination of the two separate responses. Furthermore, temperature cycles having amplitudes insufficient for entrainment of the rhythm nevertheless modified the pattern of entrainment to light - dark cycles, suggesting a synergism of light and temperature effects on the underlying circadian clock system.     

Cold entrainment of the annual cycle of ovarian activity in the lizard Lacerta vivipara: thermoperiodic rhythm versus hibernation.

A constant warm thermoperiod maintains ovarian quiescence in the lizard Lacerta vivipara, whereas a 4-month artificial hibernation rapidly induces synchronized vitellogenesis after transfer to warmth. The present study examined the possibility of a thermoperiodic regulation of the ovarian cycle and the formal properties of an internal temporal program. These questions were addressed using 24-hr thermoperiodic conditions that combined a long or a short thermophase (6 or 2 hr of basking) with a warm (19-21 degrees C), a cool (5-15 degrees C), or a cold (3-7 degrees C) cryophase. Lizards were exposed to the natural photocycle or to LD 12:12. Occurrence and timing of vitellogenesis completion were monitored using immunodetection of plasma vitellogenin and laparotomies. Cold remained stimulatory when given intermittently with a 24-hr periodicity. However, under long-thermophase conditions, lizards responded poorly to cool cryophases but fully to cold ones (72.7-100% vitellogenesis). Thus a certain amount of cold must be provided during each 24-hr cycle in order to be effective through the succession of thermocycles. Reduction of the daily heat input from 6 to 2 hr modulated the stimulating effects of cold cryophases: The median date for the beginning of vitellogenesis occurred 1 month earlier, but the number of responding females decreased from 100% to 40%. The thermoperiodic regulation of the ovarian cycle also relies upon a precise heat-cold balance per nycthemeral unit. This ensures the entrainment of an internal rhythm, since the timing of reproductive responses varies with the date of transfer from the inhibitory warm thermoperiod to the inducing thermoperiod (long thermophase, cold cryophase). At least half the females started vitellogenesis within 1-2 months after a late transfer (winter solstice) instead of 6 months after an early one (autumn equinox), and the median date for onset differed by 1 month between the two groups. However, autumn transfer was the only one to induce a group response in close agreement with the natural timing.