Melatonin rhythm observed throughout a three-cycle bright-light stimulus designed to reset the human circadian pacemaker.

Exposure to light and darkness can rapidly induce phase shifts of the human circadian pacemaker. A type 0 phase response curve (PRC) to light that has been reported for humans was based on circadian phase data collected from constant routines performed before and after a three-cycle light stimulus, but resetting data observed throughout the entire resetting protocol have not been previously reported. Pineal melatonin secretion is governed by the hypothalamic circadian pacemaker via a well-defined neural pathway and is reportedly less subject to the masking effects of sleep and activity than body temperature. The authors reasoned that observation of the melatonin rhythm throughout the three-cycle light resetting trials could provide daily phase-resetting information, allowing a dynamic view of the resetting response of the circadian pacemaker to light. Subjects (n = 12) living in otherwise dim light (approximately 10-15 lux) were exposed to a noncritical stimulus of three cycles of bright light (approximately 9500 lux for 5 h per day) timed to phase advance or phase delay the human circadian pacemaker; control subjects (n = 11) were scheduled to the same protocols but exposed to three 5-h darkness cycles instead of light. Subjects underwent initial and final constant routine phase assessments; hourly melatonin samples and body temperature data were collected throughout the protocol. Average daily phase shifts of 1 to 3 h were observed in 11 of 12 subjects receiving the bright light, supporting predictions obtained using Kronauer's phase-amplitude model of the resetting response of the human circadian pacemaker. The melatonin rhythm in the 12th subject progressively attenuated in amplitude throughout the resetting trial, becoming undetectable for >32 hours preceding an abrupt reappearance of the rhythm at a shifted phase with a recovered amplitude. The data from control subjects who remained in dim lighting and darkness delayed on average -0.2 h per day, consistent with the daily delay expected due to the longer than 24-h intrinsic period of the human circadian pacemaker. Both temperature and melatonin rhythms shifted by equivalent amounts in both bright light-treated and control subjects (R = 0.968; p<0.0001; n = 23). Observation of the melatonin rhythm throughout a three-cycle resetting trial has provided a dynamic view of the daily phase-resetting response of the human circadian pacemaker. Taken together with the observation of strong type 0 resetting in humans in response to the same three-cycle stimulus applied at a critical phase, these data confirm the importance of considering both phase and amplitude when describing the resetting of the human circadian pacemaker by light.     

Efficacy of a single sequence of intermittent bright light pulses for delaying circadian phase in humans

It has been shown in animal studies that exposure to brief pulses of bright light can phase shift the circadian pacemaker and that the resetting action of light is most efficient during the first minutes of light exposure. In humans, multiple consecutive days of exposure to brief bright light pulses have been shown to phase shift the circadian pacemaker. The aim of the present study was to determine whether a single sequence of brief bright light pulses administered during the early biological night would phase delay the human circadian pacemaker. Twenty-one healthy young subjects underwent a 6.5-h light exposure session in one of three randomly assigned conditions: 1) continuous bright light of approximately 9,500 lux, 2) intermittent bright light (six 15-min bright light pulses of approximately 9,500 lux separated by 60 min of very dim light of <1 lux), and 3) continuous very dim light of <1 lux. Twenty subjects were included in the analysis. Core body temperature (CBT) and melatonin were used as phase markers of the circadian pacemaker. Phase delays of CBT and melatonin rhythms in response to intermittent bright light pulses were comparable to those measured after continuous bright light exposure, even though the total exposure to the intermittent bright light represented only 23% of the 6.5-h continuous exposure. These results demonstrate that a single sequence of intermittent bright light pulses can phase delay the human circadian pacemaker and show that intermittent pulses have a greater resetting efficacy on a per minute basis than does continuous exposure.     

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.     

Dynamic resetting of the human circadian pacemaker by intermittent bright light

In humans, experimental studies of circadian resetting typically have been limited to lengthy episodes of exposure to continuous bright light. To evaluate the time course of the human endogenous circadian pacemaker's resetting response to brief episodes of intermittent bright light, we studied 16 subjects assigned to one of two intermittent lighting conditions in which the subjects were presented with intermittent episodes of bright-light exposure at 25- or 90-min intervals. The effective duration of bright-light exposure was 31% or 63% compared with a continuous 5-h bright-light stimulus. Exposure to intermittent bright light elicited almost as great a resetting response compared with 5 h of continuous bright light. We conclude that exposure to intermittent bright light produces robust phase shifts of the endogenous circadian pacemaker. Furthermore, these results demonstrate that humans, like other species, exhibit an enhanced sensitivity to the initial minutes of bright-light exposure.     

Phase resetting and phase singularity of an insect circannual oscillator

In circadian rhythms, the shape of the phase response curves (PRCs) depends on the strength of the resetting stimulus. Weak stimuli produce Type 1 PRCs with small phase shifts and a continuous transition between phase delays and advances, whereas strong stimuli produce Type 0 PRCs with large phase shifts and a distinct break point at the transition between delays and advances. A stimulus of an intermediate strength applied close to the break point in a Type 0 PRC sometimes produces arrhythmicity. A PRC for the circannual rhythm was obtained in pupation of the varied carpet beetle, Anthrenus verbasci, by superimposing a 4-week long-day pulse (a series of long days for 4 weeks) over constant short days. The shape of this PRC closely resembles that of the Type 0 PRC. The present study shows that the PRC to 2-week long-day pulses was Type 1, and that a 4-week long-day pulse administered close to the PRC’s break point induced arrhythmicity in pupation. It is, therefore, suggested that circadian and circannual oscillators share the same mode in phase resetting to the stimuli.     

Effects of the duper mutation on circadian responses to light

The circadian mutation duper in Syrian hamsters shortens the free-running circadian period (τ(DD)) by 2 hours when expressed on a tau mutant (τ(ss)) background and by 1 hour on a wild-type background. We have examined the effects of this mutation on phase response curves and entrainment. In contrast to wild types, duper hamsters entrained to 14L:10D with a positive phase angle. Super duper hamsters (expressing duper on a τ(ss) background) showed weak entrainment, while τ(ss) animals either completely failed to entrain or showed sporadic entrainment with episodes of relative coordination. As previously reported, wild-type and τ(ss) hamsters show low amplitude resetting in response to 15-minute light pulses after short-term (10 days) exposure to DD. In contrast, super duper hamsters show high amplitude resetting. This effect is attributable to the duper allele, as hamsters carrying duper on a wild-type background also show large phase shifts. Duper mutants that were born and raised in DD also showed high amplitude resetting in response to 15-minute light pulses, indicating that the effect of the mutation on PRC amplitude is not an aftereffect of entrainment to 14L:10D. Hamsters that are heterozygous for duper do not show amplified resetting curves, indicating that for this property, as for determination of free-running period, the mutant allele is recessive. In a modified Aschoff type II protocol, super duper and duper hamsters show large phase shifts as soon as the second day of DD. Despite the amplification of the PRC in super duper hamsters, the induction of Period1 gene expression in the SCN by light is no greater in these mutants than in wild-type animals. Period2 expression in the SCN did not differ between super duper and wild-type hamsters exposed to light at CT15, but albumin site D-binding protein (Dbp) mRNA showed higher basal levels and greater light induction in the SCN of super duper compared to wild-type animals. These results indicate that the duper mutation alters the amplitude of the circadian oscillator and further distinguish it from the tau mutation.     

Seeing the Light in Washington Chronobiology at the 3rd Annual Meeting of the Association of Professional Sleep Societies (Apss)

Chronobiology at the 3rd Annual Meeting of the APSS was, no doubt in the “Sign of Light”. Chronobiological research has certainly moved from DD to BL (bright light). Several factors contributed to the great interest and excitement regarding the effects of light on human beings. The Science publication of Czeisler et al., “Bright light induction of strong (type 0) resetting of the human circadian pacemaker”, which attracted the attention of the media, coincided with the scheduled APSS meeting. Also, a very well-attended one-day workshop in Bethesda, Maryland, on the biologic effects of light was organized by the Society for Light Therapy and Biological Rhythms the day before, and succeeded in raising interest and excitement over the subject. The presentations on the biologic effects of light on the circadian system focused on the phase resetting-response to light of the human circadian system, and on the practical implications of this resetting on seasonal affective disorders (SAD), in sleep-phase-delay and sleep-phase-advance syndromes as well as sleep disturbances associated with jet lag.     

Probing the circadian pacemaker of a mouse using two light pulses

In two separate sets of experiments, the phases of the locomotor activity rhythm of the nocturnal field mouse Mus booduga were probed using two light pulses (LPs). In the first set of experiments, the circadian pacemaker underlying the locomotor activity rhythm was perturbed at circadian time 14 (CT 14) using a resetting light pulse LP1 of 1000 lux intensity and 15 min duration. The phases of the resetting pacemaker were then probed at all even CTs between CT 16 and CT 14 using a PRC probing light pulse LP2 of equal strength. The "LP2 PRC" thus obtained was then compared with the single light pulse PRC in terms of the area under delay (D) and advance (A) zones of the PRCs. The time course and waveform of the two LP PRCs suggest that the LP2 PRC resembled the single LP PRC, displaced by 2 h toward the right. The LP1 PRC had smaller D compared to the single LP PRC (p = 0.007), whereas both the PRCs had A of equal magnitude (p = 0.23). This suggests that the pacemaker phase shifts rapidly after LP perturbations. In the second set of experiments, the LP1 was administered at CT 14. The phase of the pacemaker was then perturbed on day 1 (next cycle after LP1) either 2 h after activity onset (at ca. CT 14 of the transient cycle) or 8 h after activity onset (at ca. CT 20 of the transient cycle) using an LP2 of equal strength. It was observed that the steady-state phase shifts evoked by positioning an LP2, 2 h after activity onset, were positively correlated with the phase shifts observed on day 1. The steady-state phase shifts observed, when the LP2 was positioned, 8 h after activity onset, were negatively correlated with the phase shifts observed on day 1. These results suggest that the transient cycles do not mirror the state of the pacemaker oscillator.     

Circadian effects of light no brighter than moonlight

In mammals, light entrains endogenous circadian pacemakers by inducing daily phase shifts via a photoreceptor mechanism recently discovered in retinal ganglion cells. Light that is comparable in intensity to moonlight is generally ineffective at inducing phase shifts or suppressing melatonin secretion, which has prompted the view that circadian photic sensitivity has been titrated so that the central pacemaker is unaffected by natural nighttime illumination. However, the authors have shown in several different entrainment paradigms that completely dark nights are not functionally equivalent to dimly lit nights, even when nighttime illumination is below putative thresholds for the circadian visual system. The present studies extend these findings. Dim illumination is shown here to be neither a strong zeitgeber, consistent with published fluence response curves, nor a potentiator of other zeitgebers. Nevertheless, dim light markedly alters the behavior of the free-running circadian pacemaker. Syrian hamsters were released from entrained conditions into constant darkness or dim narrowband green illumination (~0.01 lx, 1.3 x 10(-9) W/cm(2), peak lambda = 560 nm). Relative to complete darkness, constant dim light lengthened the period by ~0.3 h and altered the waveform of circadian rhythmicity. Among animals transferred from long day lengths (14 L:10 D) into constant conditions, dim illumination increased the duration of the active phase (alpha) by ~3 h relative to complete darkness. Short day entrainment (8 L:16 D) produced initially long alpha that increased further under constant dim light but decreased under complete darkness. In contrast, dim light pulses 2 h or longer produced effects on circadian phase and melatonin secretion that were small in magnitude. Furthermore, the amplitude of phase resetting to bright light and nonphotic stimuli was similar against dimly lit and dark backgrounds, indicating that the former does not directly amplify circadian inputs. Dim illumination markedly alters circadian waveform through effects on alpha, suggesting that dim light influences the coupling between oscillators theorized to program the beginning and end of subjective night. Physiological mechanisms responsible for conveying dim light stimuli to the pacemaker and implications for chronotherapeutics warrant further study.     

Accuracy of human circadian entrainment under natural light conditions: model simulations

The patterns of light intensity to which humans expose their circadian pacemakers in daily life are very irregular and vary greatly from day to day. The circadian pacemaker can adjust to such irregular exposure patterns by daily phase shifts, such as summarized in a phase response curve. It is demonstrated in this paper on the basis of computer simulations applying actually recorded human light exposure patterns that the pacemaker can substantially improve its accuracy by an additional response to light: For that purpose, it should additionally change its angular velocity (and consequently its period tau) in response to light. Reductions of tau in response to light in the morning and increases of tau in response to light in the evening can lead to an increase in entrained pacemaker accuracy with about 25%. Circadian pacemakers have evolved as accurate internal representations of external time, and investigated diurnal mammals all seem to respond to light by changing the period of their circadian pacemaker (in addition to shifting phase). The authors suggest that also human circadian systems take advantage of this possibility and that their pacemakers respond to light by shifting phase and changing period. As a consequence of this postulated mechanism, the simulations demonstrate that the period of the pacemaker under normally entrained conditions is 24 h. The maximum accuracy corresponds to a day-to-day standard deviation of the time of phase 0 of circa 15 min. This is considerably more accurate than the light signal humans usually perceive.     

Conditioning in the Orcadian System

Light is the dominant environmental cue for entrainment of circadian rhythms. In mammals, light entrains rhythms by resetting the phase of a circadian pacemaker located in the hypothalamic suprachiasmatic nucleus (SCN). Until recently, the mechanism responsible for pacemaker resetting by light was thought to be exclusively sensitive to photic cues. New experiments indicate, however, that this mechanism is more plastic than once thought; is amenable to conditioned stimulus control; and is capable of acquiring, through conditioning, new response capabilities. These experiments showed that, in rats, a neutral stimulus paired with light in Pavlovian conditioning trials is capable of eliciting cellular and behavioral effects characteristic of circadian clock phase resetting by light, expression of Fos protein in the ventrolateral region of the SCN, and phase shifts of free-running rhythms. These novel results open up a previously unappreciated perspective on photic phase resetting and entrainment of circadian rhythms. Specifically, they suggest that the process normally initiated by light to reset the clock can be modified by learning and events in the environment that reliably precede the onset of light can assume the resetting function of light.     

Photic resetting of the human circadian pacemaker in the absence of conscious vision

Ocular light exposure patterns are the primary stimuli for entraining the human circadian system to the local 24-h day. Many totally blind persons cannot use these stimuli and, therefore, have circadian rhythms that are not entrained. However, a few otherwise totally blind persons retain the ability to suppress plasma melatonin concentrations after ocular light exposure, probably using a neural pathway that includes the site of the human circadian pacemaker, suggesting that light information is reaching this site. To test definitively whether ocular light exposure could affect the circadian pacemaker of some blind persons and whether melatonin suppression in response to bright light correlates with light-induced phase shifts of thecircadian system, the authorsperformed experiments with 5 totally blind volunteers using a protocol known to induce phase shifts of the circadian pacemaker in sighted individuals. In the 2 blind individuals who maintained light-induced melatonin suppression, the circadian system was shifted by appropriately timed bright-light stimuli. These data demonstrate that light can affect the circadian pacemaker of some totally blind individuals--either by altering the phase of the circadian pacemaker or by affecting its amplitude. They are consistent with data from animal studies demonstrating that there are different neural pathways and retinal cells that relay photic information to the brain: one for conscious light perception and the other for non-image-forming functions.     

Bright light during lactation alters the functioning of the circadian system of adult rats

To examine the role of light in the maturation of the circadian pacemaker, twelve groups of rats were raised in different conditions of exposure to constant bright light (LL) during lactation: both duration and timing of LL were varied. We studied the motor activity rhythm of the rats after weaning, first under LL and then under constant darkness (DD). In DD, two light pulses [at circadian time 15 (CT15) and CT22] were applied to test the response of the pacemaker. Greater exposure to LL days during lactation increased the number of rhythmic animals and the amplitude of their motor activity rhythm in the LL stage and decreased the phase delay due to the light pulse at CT15. The timing of LL during lactation affected these variables too. Because the response of the adult to light depended on both the number and timing of LL days during lactation, the exposure to light at early stages may influence the development of the circadian system by modifying it structurally or functionally.     

Rapid Phase Resetting of a Mammalian Orcadian Rhythm by Brief Light Pulses

The phase-shift (Δψ) responses of the circadian rhythm in the field mouse Mus booduga to brief light pulses (LPs) of 15 minutes duration and 1000 lux intensity were measured in 90 experiments. In each experiment, a resetting light pulse LP₁ was administered at CT14 (CT, circadian time), and a scanning light pulse LP₂ was then variously administered in separate experiments at CT16, CT20, and CT22 in the same and in the next circadian cycle. The Δψ obtained in all these two-pulse experiments did not differ significantly from theoretical values computed on the assumption that LP₁ reset the phase response curve (PRC) rapidly. In each case, the steady-state Δψ observed after LP₁ and LP₂ differed significantly from the Δψ obtained at the same CT in determination of the single-pulse PRC (control) and also differed significantly from the values on the assumption of no Δψ in the PRC following LP₁. These results indicate that the circadian pacemaker of M. booduga, as measured by its PRC, is substantially reset within 2h after a light pulse at CT14. (Chronobiology International 14(6), 537–548, 1997)     

Quantifying human circadian pacemaker response to brief, extended, and repeated light stimuli over the phototopic range

The authors' previous models have been able to describe accurately the effects of extended (approximately 5 h) bright-light (>4000 lux) stimuli on the phase and amplitude of the human circadian pacemaker, but they are not sufficient to represent the surprising human sensitivity to both brief pulses of bright light and light of more moderate intensities. Therefore, the authors have devised a new model in which a dynamic stimulus processor (Process L) intervenes between the light stimuli and the traditional representation of the circadian pacemaker as a self-sustaining limit-cycle oscillator (Process P). The overall model incorporating Process L and Process P is intended to allow the prediction of phase shifts to photic stimuli of any temporal pattern (extended and brief light episodes) and any light intensity in the photopic range. Two time constants emerge in the Process L model: the characteristic duration for necessary bright-light pulses to achieve their full effect (5-10 min) and the characteristic stimulus-free (dark) interval that can be tolerated without incurring an excessive penalty in phase shifting (30-80 min). The effect of reducing light intensity is incorporated in Process L as an extension of the time necessary for the light pulse to be fully realized (a power-law relation between time and intensity). This new model generates a number of new testable hypotheses, including the surprising prediction that 24-h cycles consisting of 8 h of darkness and 16 h of only approximately 3.5 lux would be capable of entraining a large fraction of the adult population (approximately 45%). Experimental data on the response of the human circadian system to lower light intensities and briefer stimuli are needed to allow for further refinement and validation of the model proposed here.     

Photoperiod differentially modulates photic and nonphotic phase response curves of hamsters

Circadian pacemakers respond to light pulses with phase adjustments that allow for daily synchronization to 24-h light-dark cycles. In Syrian hamsters, Mesocricetus auratus, light-induced phase shifts are larger after entrainment to short daylengths (e.g., 10 h light:14 h dark) vs. long daylengths (e.g., 14 h light:10 h dark). The present study assessed whether photoperiodic modulation of phase resetting magnitude extends to nonphotic perturbations of the circadian rhythm and, if so, whether the relationship parallels that of photic responses. Male Syrian hamsters, entrained for 31 days to either short or long daylengths, were transferred to novel wheel running cages for 2 h at times spanning the entire circadian cycle. Phase shifts induced by this stimulus varied with the circadian time of exposure, but the amplitude of the resulting phase response curve was not markedly influenced by photoperiod. Previously reported photoperiodic effects on photic phase resetting were verified under the current paradigm using 15-min light pulses. Photoperiodic modulation of phase resetting magnitude is input specific and may reflect alterations in the transmission of photic stimuli.     

Light pulses induce "singular" behavior and shorten the period of the circadian phototaxis rhythm in the CW15 strain of Chlamydomonas.

While measuring action spectra for phase-shifting the circadian clock of Chlamydomonas, we observed that light pulses started near the phase response curve (PRC) "breakpoint" caused a reduction of the amplitude of the phototactic rhythm and two unexpected effects: (1) nonmonotonic fluence response curves (FRCs), and (2) shortening of the period of the subsequent free-running rhythm. The reduction of the rhythm's amplitude is dependent upon both the fluence and wavelength of the light pulse. The results are consistent with the amplitude being dependent upon the perceived "strength" of the stimulus, and with the nonmonotonic FRCs and reduced amplitude reflecting a light-induced change of the pacemaker's state variables to a region of the phase plane close to the "singularity." The period change that is evoked by single stimuli exhibits novel characteristics: large changes in period and a phase specificity that correlates with "singular" behavior. These period changes also appear to be a function of the stimulus strength, but indirectly; the magnitude of the period change is most strongly correlated with the magnitude of the light-induced phase shift. These results are interpreted in the context of limit cycle models of circadian clocks, and are used to suggest new tactics for measuring action spectra of light-induced clock resetting.     

Photobiology of the Gonyaulax circadian system. I. Different phase response curves for red and blue light

Phase responses to red and blue light pulses were measured at different times during the circadian cycle (phase response curves, PRC) in the marine unicellular dinoflagellate Gonyaulaxpolyedra Stein. Pulses were given during a 24-h period of darkness; thereafter, cultures were released into constant dim red light for the assessment of phase and period. The results confirmed earlier findings that the Gonyaulax circadian system receives light signals via two distinct input pathways. During the subjective day and for the first 3 h of the subjective night, red and blue light pulses led to identical phase responses. For the rest of the circadian cycle, however, phase responses to pulses of either red or blue light differed drastically both in their amplitude and direction (advances or delays). Thus, the Gonyaulax light PRC is generated by two distinct light responses. One of these represents responses via a light input that is responsive both to red and blue light mainly producing small delays. The other represents responses of a primarily blue-sensitive input system leading to large advances restricted to the subjective night. Via feed-back, the blue-sensitive light input appears to be under the control of the circadian system. Received: 27 November 1996/Accepted: 30 January 1997     

Photic and nonphotic circadian phase resetting in a diurnal primate, the common marmoset

Despite the considerable literature on circadian entrainment, there is little information on this subject in diurnal mammals. Contributing to this lack of understanding is the problem of separating photic from nonphotic (behavioral) phase-resetting events in diurnal species. In the present study, photic phase resetting was obtained in diurnal common marmosets held under constant dim light (DimDim; <0.5 lx) by using a 20-s pulse of bright light to minimize time available for behavioral arousal. This stimulus elicited phase advances at circadian time (CT) 18-22 and phase delays at CT9-12. Daily presentation of these 20-s pulses produced entrainment with a phase angle of approximately 11 h (0 h = activity onset). Nonphotic phase resetting was obtained under DimDim with the use of a 1-h-induced activity pulse, consisting of intermittent cage agitation and water sprinkling, delivered in total darkness to minimize photic effects. This stimulus caused phase delays at CT20-24, and entrainment to a scheduled daily regimen of these pulses occurred with a phase angle of approximately 0 h. These results indicate that photic and nonphotic phase-response curves (PRCs) of marmosets are similar to those of nocturnal rodents and that nonphotic PRCs are keyed to the phase of the suprachiasmatic nucleus pacemaker, not to the phase of the activity-rest cycle.     

Light pulses suppress responsiveness within the mouse photic entrainment pathway

We have characterized a decrease in photic responsiveness of the mammalian circadian entrainment pathway caused by light stimulation. Phase delays of the running-wheel activity rhythm were used to quantify the photic responsiveness of the circadian system in mice (C57BL/6J). In an initial experiment, the authors measured the responsiveness to single "saturating" light pulses ("white" fluorescent light; approximately 1876 [microW; 15 min). In two additional experiments, the authors measured responses to this stimulus at several time points following a saturating pulse at CT 14 or CT 16. Data from these experiments were analyzed in two manners. Experiment 2 was analyzed assuming that the phase of the circadian pacemaker was unchanged by an initial pulse, and Experiment 3 was analyzed assuming that the initial pulse induced an instantaneous phase delay. Results reveal a significant reduction in responsivity to light that persists for at least 2 h and possibly up to 4 h after the initial stimulus. Immediately after the stimulus, the responsiveness of the photic entrainment pathway was reduced to levels < or = 12% of normal. After 2 h, the responsiveness was < or = 42% of normal, and by 4 h, responsiveness had recovered to levels that were < or = 60% of normal (levels not statistically different from controls). By the following circadian cycle, responsiveness was more completely recovered, although the magnitude of some phase delays remained < or = 85% of normal. These major reductions in the magnitude of phase delays (and phase response curve amplitude) caused by saturating light pulses confound interpretations of two-pulse experiments designed to measure the rate of circadian phase delays. In addition, the time course for this reduced responsiveness may reflect the time course of cellular and molecular events that underlie light-induced resetting of the mammalian circadian pacemaker.