The visual input stage of the mammalian circadian pacemaking system: II. The effect of light and drugs on retinal function.

Acute light pulses as well as long-term light exposure may not only modulate photoreceptive properties, but also induce reversible or irreversible damage to the retina, depending on exposure conditions. Illuminance levels in laboratory animal colonies and manipulations of lighting regimens in circadian rhythm research can threaten retinal structure and physiology, and may therefore modify zeitgeber input to the central circadian system. Given the opportunity to escape light at any time, the nocturnal rat self-selects a seasonally varying "naturalistic skeleton photoperiod" that protects the eyes from potential damage by nonphysiological light exposures. Both rod rod-segment disk shedding and behavioral circadian phase shifts are elicited by low levels of twilight stimulation. From this vantage point, we hypothesize that certain basic properties of circadian rhythms (e.g., Aschoff's rule and splitting) may reflect modulation of retinal physiology by light. Pharmacological manipulations with or without the addition of lighting strategies have been used to analyze the neurochemistry of circadian timekeeping. Drug modulation of light input at the level of the retina may add to or interact with direct drug modulation of the central circadian pacemaking system.     

High-intensity red light suppresses melatonin

Early studies on rodents indicated that the long-wavelength portion of the spectrum (orange- and red-appearing light) could influence circadian and neuroendocrine responses. Since then, both polychromatic and analytic action spectra in various rodent species have demonstrated that long-wavelength light is very weak, if not entirely inactive, for regulating neurobehavioral responses. Since testing of monochromatic light wavelengths above 600 nm is uncommon, many researchers have assumed that there is little to no effect of red light on the neuroendocrine or circadian systems. The aims of the following studies were to test the efficacy of monochromatic light above 600 nm for melatonin suppression in hamsters and humans. Results in hamsters show that 640 nm monochromatic light at 1.1 x 10(17) photons/cm2 can acutely suppress pineal melatonin levels. In normal healthy humans, equal photon density exposures of 1.9 x 10(18) photons/cm2 at 460, 630, and 700 nm monochromatic light elicited a significant melatonin suppression at 460 nm and small reductions of plasma melatonin levels at 630 and 700 nm. These findings are discussed relative to the possible roles of classical visual photoreceptors and the recently discovered intrinsically photosensitive retinal ganglion cells for circadian phototransduction. That physiology, and its potential for responding to red light, has implications for domestic applications involving animal care, the lighting of typical human environments, and advanced applications such as space exploration.     

High‐intensity red light suppresses melatonin

Early studies on rodents indicated that the long‐wavelength portion of the spectrum (orange‐ and red‐appearing light) could influence circadian and neuroendocrine responses. Since then, both polychromatic and analytic action spectra in various rodent species have demonstrated that long‐wavelength light is very weak, if not entirely inactive, for regulating neurobehavioral responses. Since testing of monochromatic light wavelengths above 600 nm is uncommon, many researchers have assumed that there is little to no effect of red light on the neuroendocrine or circadian systems. The aims of the following studies were to test the efficacy of monochromatic light above 600 nm for melatonin suppression in hamsters and humans. Results in hamsters show that 640 nm monochromatic light at 1.1×10¹⁷ photons/cm² can acutely suppress pineal melatonin levels. In normal healthy humans, equal photon density exposures of 1.9×10¹⁸ photons/cm² at 460, 630, and 700 nm monochromatic light elicited a significant melatonin suppression at 460 nm and small reductions of plasma melatonin levels at 630 and 700 nm. These findings are discussed relative to the possible roles of classical visual photoreceptors and the recently discovered intrinsically photosensitive retinal ganglion cells for circadian phototransduction. That physiology, and its potential for responding to red light, has implications for domestic applications involving animal care, the lighting of typical human environments, and advanced applications such as space exploration.     

Molecular cloning, mRNA expression, and immunocytochemical localization of a putative blue-light photoreceptor CRY4 in the chicken pineal gland

In non-mammalian vertebrates, the pineal gland contains an endogenous circadian oscillator and serves as a photosensitive neuroendocrinal organ. To better understand the pineal phototransduction mechanism, we focused on the chicken putative blue-light photoreceptive molecule, Cryptochrome4 (cCRY4). Here we report the molecular cloning of pineal cCry4 cDNA, the in vivo expression of cCry4 mRNA, and the detection of cCRY4 protein. cCry4 is transcribed in a wide variety of chick tissues out of which the pineal gland and retina contain high levels of cCry4 mRNA. In the pineal gland, under 12 h light : 12 h dark cycles, the levels of both cCry4 mRNA and cCRY4 protein showed diurnal changes, and in cultured chick pineal cells, the cCry4 mRNA level was not only up-regulated by light but also controlled by circadian signals. Immunoblot analysis with a cCRY4-specific antibody detected cCRY4 in a soluble fraction of the pineal lysate. Immunocytochemistry revealed that cCRY4 was expressed in many parenchymal cells and a limited number of stromal cells. These cCRY4 features strikingly contrast with those of the chick pineal photoreceptor pinopsin, suggesting a possible temporal and/or spatial duplicity of the pineal photoreceptive system, the opsin- and CRY-based mechanisms.     

Dim light adaptation attenuates acute melatonin suppression in humans

Abstract Studies in rodents with retinal degeneration indicated that neither the rod nor the cone photoreceptors obligatorily participate in circadian responses to light, including melatonin suppression and photoperiodic response. Yet there is a residual phase-shifting response in melanopsin knockout mice, which suggests an alternate or redundant means for light input to the SCN of the hypothalamus. The findings of Aggelopoulos and Meissl suggest a complex, dynamic interrelationship between the classic visual photoreceptors and SCN cell sensitivity to light stimuli, relative to various adaptive lighting conditions. These studies raised the possibility that the phototransductive physiology of the retinohypothalamic tract in humans might be modulated by the visual rod and cone photoreceptors. The aim of the following two-part study was to test the hypothesis that dim light adaptation will dampen the subsequent suppression of melatonin by monochromatic light in healthy human subjects. Each experiment included 5 female and 3 male human subjects between the ages of 18 and 30 years, with normal color vision. Dim white light and darkness adaptation exposures occurred between midnight and 0200 h, and a full-field 460-nm light exposure subsequently occurred between 0200 and 0330-h for each adaptation condition, at 2 different intensities. Plasma samples were drawn following the 2-h adaptation, as well as after the 460-nm monochromatic light exposure, and melatonin was measured by radioimmunoassay. Comparison of melatonin suppression responses to monochromatic light in both studies revealed a loss of significant suppression after dim white light adaptation compared with dark adaptation (p < 0.04 and p < 0.01). These findings indicate that the activity of the novel circadian photoreceptive system in humans is subject to subthreshold modulation of its sensitivity to subsequent monochromatic light exposure, varying with the conditions of light adaptation prior to exposure.     

Exogenous melatonin reduces the resynchronization time after phase shifts of a nonphotic zeitgeber in the house sparrow (Passer domesticus)

Continuous melatonin administration via silastic implants accelerates the resynchronization of the circadian locomotor activity rhythm in house sparrows (Passer domesticus) after exposure to phase shifts of a weak light-dark cycle. Constant melatonin might induce this effect either by increasing the sensitivity of the visual system to a light zeitgeber or by reducing the degree of self-sustainment of the circadian pacemaker. To distinguish between these two possible mechanisms, two groups of house sparrows, one carrying melatonin implants and the other empty implants, were kept in constant dim light and subjected to advance and delay shifts of a 12-h feeding phase. The resynchronization times of their circadian feeding rhythm following the phase shifts were significantly shorter when the birds carried melatonin implants than when they carried empty implants. In a second experiment, melatonin-implanted and control birds were released into food ad libitum conditions 2 days after either a delay or an advance phase shift. The number of hours by which the activity rhythms had been shifted on the second day in food ad libitum conditions was assessed. Melatonin-implanted house sparrows had significantly larger phase shifts in their circadian feeding rhythm than control birds. This is in accordance with the first experiment since a larger phase shift at a given time reflects accelerated resynchronization. Additionally, the second experiment also excludes any possible masking effects of the nonphotic zeitgeber. In conclusion, constant melatonin accelerates resynchronization even after phase shifts of a nonphotic zeitgeber, indicating that constant high levels of melatonin can reduce the degree of self-sustainment of the circadian pacemaker independent of any effects on the photoreceptive system.     

Effect of Light Intensity on the Phase and Period Response Curves in the Nocturnal Field Mouse Mus booduga

The effect of light intensity on the phase response curve (PRC) and the period response curve (τRC) of the nocturnal field mouse Mus booduga was studied. PRCs and τRCs were constructed by exposing animals free-running in constant darkness (DD), to fluorescent light pulses (LPs) of 100 lux and 1000 lux intensities for 15min duration. The waveform of the PRCs and τRCs evoked by high light intensity (1000 lux) stimuli was significantly different compared to those constructed using low light intensity (100 lux). Moreover, a weak but significant correlation was observed between phase shifts and period changes when light stimuli of 1000 lux intensity were used; however, the phase shifts and period changes in the 100 lux PRC and τRC were not correlated. This suggests that the intensity of light stimuli affects both phase and period responses in the locomotor activity rhythm of the nocturnal field mouse M. booduga. These results indicate that complex mechanisms are involved in entrainment of circadian clocks, even in nocturnal rodents, in which PRC, τRC, and dose responses play a significant role.     

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.     

The timing of the human circadian clock is accurately represented by the core body temperature rhythm following phase shifts to a three-cycle light stimulus near the critical zone

A double-stimulus experiment was conducted to evaluate the phase of the underlying circadian clock following light-induced phase shifts of the human circadian system. Circadian phase was assayed by constant routine from the rhythm in core body temperature before and after a three-cycle bright-light stimulus applied near the estimated minimum of the core body temperature rhythm. An identical, consecutive three-cycle light stimulus was then applied, and phase was reassessed. Phase shifts to these consecutive stimuli were no different from those obtained in a previous study following light stimuli applied under steady-state conditions over a range of circadian phases similar to those at which the consecutive stimuli were applied. These data suggest that circadian phase shifts of the core body temperature rhythm in response to a three-cycle stimulus occur within 24 h following the end of the 3-day light stimulus and that this poststimulus temperature rhythm accurately reflects the timing of the underlying circadian clock.     

Circadian phototransduction and the regulation of biological rhythms

The vertebrate circadian system that controls most biological rhythms is composed of multiple oscillators with varied hierarchies and complex levels of organization and interaction. The retina plays a key role in the regulation of daily rhythms and light is the main synchronizer of the circadian system. To date, the identity of photoreceptors/photopigments responsible for the entrainment of biological rhythms is still uncertain; however, it is known that phototransduction must occur in the eye because light entrainment is lost with eye removal. The retina is also rhythmic in physiological and metabolic activities as well as in gene expression. Retinal oscillators may act like clocks to induce changes in the visual system according to the phase of the day by predicting environmental changes. These oscillatory and photoreceptive capacities are likely to converge all together on selected retinal cells. The aim of this overview is to present the current knowledge of retinal physiology in relation to the circadian timing system.     

Non-photic modulation of phase shifts to long light pulses

Circadian rhythms can be reset by both photic and non-photic stimuli. Recent studies have used long light exposure to produce photic phase shifts or to enhance non-photic phase shifts. The presence or absence of light can also influence the expression of locomotor rhythms through masking; light during the night attenuates locomotor activity, while darkness during the day induces locomotor activity in nocturnal animals. Given this dual role of light, the current study was designed to examine the relative contributions of photic and non-photic components present in a long light pulse paradigm. Mice entrained to a light/dark cycle were exposed to light pulses of various durations (0, 3, 6, 9, or 12 h) starting at the time of lights-off. After the light exposure, animals were placed in DD and were either left undisturbed in their home cages or had their wheels locked for the remainder of the subjective night and subsequent subjective day. Light treatments of 6, 9, and 12 h produced large phase delays. These treatments were associated with decreased activity during the nocturnal light and increased activity during the initial hours of darkness following light exposure. When the wheels were locked to prevent high-amplitude activity, the resulting phase delays to the light were significantly attenuated, suggesting that the activity following the light exposure may have contributed to the overall phase shift. In a second experiment, telemetry probes were used to assess what effect permanently locking the wheels had on the phase shift to the long light pulses. These animals had phase shifts fully as large as animals without any form of wheel lock, suggesting that while non-photic events can modulate photic phase shifts, they do not play a role in the full phase-shift response observed in animals exposed to long light pulses. This paradigm will facilitate investigations into non-photic responses of the mouse circadian system.     

Effect of light intensity on the phase and period response curves in the nocturnal field mouse Mus booduga

The effect of light intensity on the phase response curve (PRC) and the period response curve (τRC) of the nocturnal field mouse Mus booduga was studied. PRCs and τRCs were constructed by exposing animals free-running in constant darkness (DD), to fluorescent light pulses (LPs) of 100 lux and 1000 lux intensities for 15min duration. The waveform of the PRCs and τRCs evoked by high light intensity (1000 lux) stimuli was significantly different compared to those constructed using low light intensity (100 lux). Moreover, a weak but significant correlation was observed between phase shifts and period changes when light stimuli of 1000 lux intensity were used; however, the phase shifts and period changes in the 100 lux PRC and τRC were not correlated. This suggests that the intensity of light stimuli affects both phase and period responses in the locomotor activity rhythm of the nocturnal field mouse M. booduga. These results indicate that complex mechanisms are involved in entrainment of circadian clocks, even in nocturnal rodents, in which PRC, τRC, and dose responses play a significant role.     

Photic sensitivity ranges of hamster pupillary and circadian phase responses do not overlap

Mammalian retinal photoreceptors form an irradiance detection system that drives many nonvisual responses to light such as pupil reflex and resetting of the circadian clock. To understand the role of pupil size in circadian light responses, pupil diameter was pharmacologically manipulated and the effect on behavioral phase shifts at different irradiance levels was studied in the Syrian hamster. Dose-response curves for steady-state pupil size and for behavioral phase shifts were constructed for 3 pupil conditions (dilated, constricted, and control). Retinal irradiance was calculated from corneal irradiance, pupil size, retinal surface area, and absorption of ocular media. The sensitivity of photic responses to retinal irradiance is approximately 1.5 log units higher than to corneal irradiance. When plotted against corneal irradiance, pharmacological pupil constriction reduces the light sensitivity of the circadian system, but pupil dilation has no effect. As expected, when plotted against retinal irradiance all dose-response curves superimposed, confirming that the circadian system responds to photon flux on the retina. Pupil dilation does not increase the circadian response to increasing irradiance, since the response of the circadian system attains saturation at irradiance levels lower than those required to induce pupil constriction. The main finding shows that due to the different response sensitivities, the effect of pupil constriction on the light sensitivity of the circadian system in the hamster under natural conditions is virtually negligible. We further suggest the existence of distinct modulating mechanisms for the differential retinal irradiance sensitivity of the pupil system and the circadian system, which enables the different responses to be tuned to their specific tasks while using similar photoreceptive input.     

TheBulla ocular circadian pacemaker

In an effort to understand the cellular basis of entrainment of circadian oscillators we have studied the role of membrane potential changes in the neurons which comprise the ocular circadian pacemaker of Bulla gouldiana in mediating phase shifts of the ocular circadian rhythm. We report that: 1. Intracellular recording was used to measure directly the effects of the phase shifting agents light, serotonin, and 8-bromo-cAMP on the membrane potential of the basal retinal neurons. We found that light pulses evoke a transient depolarization followed by a smaller sustained depolarization. Application of serotonin produced a biphasic response; a transient depolarization followed by a sustained hyperpolarization. Application of a membrane permeable analog of the intracellular second messenger cAMP, 8-bromo-cAMP, elicited sustained hyperpolarization, and occasionally a weak phasic depolarization. 2. Changing the membrane potential of the basal retinal neurons directly and selectively with intracellularly injected current phase shifts the ocular circadian rhythm. Both depolarizing and hyperpolarizing current can shift the phase of the circadian oscillator. Depolarizing current mimics the phase shifting action of light, while hyperpolarizing current produces phase shifts which are transposed approximately 180 degrees in circadian time to depolarization. 3. Altering BRN membrane potential with ionic treatments, depolarizing with elevated K+ seawater or hyperpolarizing with lowered Na+ seawater, produces phase shifts similar to current injection. 4. The light-induced depolarization of the basal retinal neurons is necessary for phase shifts by light. Suppressing the light-induced depolarization with injected current inhibits light-induced phase shifts. 5. The ability of membrane potential changes to shift oscillator phase is dependent on extracellular calcium. Reducing extracellular free Ca++ from 10 mM to 1.3 X 10(-7) M inhibits light-induced phase shifts without blocking the photic response of the BRNs. The results indicate that changes in the membrane potential of the pacemaker neurons play a critical role in phase shifting the circadian rhythm, and imply that a voltage-dependent and calcium-dependent process, possibly Ca++ influx, shifts oscillator phase in response to 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.     

Social influences on mammalian circadian rhythms: animal and human studies

While light is considered the dominant stimulus for entraining (synchronizing) mammalian circadian rhythms to local environmental time, social stimuli are also widely cited as 'zeitgebers' (time-cues). This review critically assesses the evidence for social influences on mammalian circadian rhythms, and possible mechanisms of action. Social stimuli may affect circadian behavioural programmes by regulating the phase and period of circadian clocks (i.e. a zeitgeber action, either direct or by conditioning to photic zeitgebers), by influencing daily patterns of light exposure or modulating light input to the clock, or by associative learning processes that utilize circadian time as a discriminative or conditioned stimulus. There is good evidence that social stimuli can act as zeitgebers. In several species maternal signals are the primary zeitgeber in utero and prior to weaning. Adults of some species can also be phase shifted or entrained by single or periodic social interactions, but these effects are often weak, and appear to be mediated by social stimulation of arousal. There is no strong evidence yet for sensory-specific nonphotic inputs to the clock. The circadian phase-dependence of clock resetting to social stimuli or arousal (the 'nonphotic' phase response curve, PRC), where known, is distinct from that to light and similar in diurnal and nocturnal animals. There is some evidence that induction of arousal can modulate light input to the clock, but no studies yet of whether social stimuli can shift the clock by conditioning to photic cues, or be incorporated into the circadian programme by associative learning. In humans, social zeitgebers appear weak by comparison with light. In temporal isolation or under weak light-dark cycles, humans may ignore social cues and free-run independently, although cases of mutual synchrony among two or more group-housed individuals have been reported. Social cues may affect circadian timing by controlling sleep-wake states, but the phase of entrainment observed to fixed sleep-wake schedules in dim light is consistent with photic mediation (scheduled variations in behavioural state necessarily create daily light-dark cycles unless subjects are housed in constant dark or have no eyes). By contrast, discrete exercise sessions can induce phase shifts consistent with the nonphotic PRC observed in animal studies. The best evidence for social entrainment in humans is from a few totally blind subjects who synchronize to the 24 h day, or to near-24 h sleep-wake schedules under laboratory conditions. However, the critical entraining stimuli have not yet been identified, and there are no reported cases yet of social entrainment in bilaterally enucleated blind subjects. The role of social zeitgebers in mammalian behavioural ecology, their mechanisms of action, and their utility for manipulating circadian rhythms in humans, remains to be more fully elaborated.     

Phototransduction Influences Metabolic Flux and Nucleotide Metabolism in Mouse Retina

Production of energy in a cell must keep pace with demand. Photoreceptors use ATP to maintain ion gradients in darkness, whereas in light they use it to support phototransduction. Matching production with consumption can be accomplished by coupling production directly to consumption. Alternatively, production can be set by a signal that anticipates demand. In this report we investigate the hypothesis that signaling through phototransduction controls production of energy in mouse retinas. We found that respiration in mouse retinas is not coupled tightly to ATP consumption. By analyzing metabolic flux in mouse retinas, we also found that phototransduction slows metabolic flux through glycolysis and through intermediates of the citric acid cycle. We also evaluated the relative contributions of regulation of the activities of α-ketoglutarate dehydrogenase and the aspartate-glutamate carrier 1. In addition, a comprehensive analysis of the retinal metabolome showed that phototransduction also influences steady-state concentrations of 5′-GMP, ribose-5-phosphate, ketone bodies, and purines.     

Photic and nonphotic inputs to the diurnal circadian clock

Diurnal animals occupy a different temporal niche from nocturnal animals and are consequently exposed to different amounts of light as well as different dangers. Accordingly, some variation exists in the way that diurnal animals synchronize their internal circadian clock to match the external 24-hour daily cycle. First, though the brain mechanisms underlying photic entrainment are very similar among species with different daily activity patterns, there is evidence that diurnal animals are less sensitive to photic stimuli compared to nocturnal animals. Second, stimuli other than light that synchronize rhythms (i.e. nonphotic stimuli) can also entrain and phase shift daily rhythms. Some of the rules that govern nonphotic entrainment in nocturnal animals as well as the brain mechanisms that control nonphotic influences on rhythms do not appear to apply to diurnal animals, however. Some evidence supports the idea that arousal or activity plays an important role in entraining rhythms in diurnal animals, either during the light (active) or dark (inactive) phases, though no consistent pattern is seen. GABAergic stimulation induces phase shifts during the subjective day in both diurnal and nocturnal animals. In diurnal Arvicanthis niloticus (Nile grass rats), SCN GABA_A receptor activation at this time results in phase delays while in nocturnal animals phase advances are induced. It appears that the effect of GABA at this circadian phase results from the inhibition of period gene expression in both diurnal and nocturnal animals. Nonetheless, the resulting phase shifts are in opposite directions. It is not known what stimuli or behaviours ultimately induce changes in GABA activity in the SCN that result in alterations of circadian phase in diurnal grass rats. Taken together, studies such as these suggest that it may be problematic to apply the principles governing nocturnal nonphotic entrainment and its underlying mechanisms to diurnal species including humans.     

Circadian organization and photoreception in an Australian dasyurid marsupial (Sminthopsis macroura)

Much is known about the formal properties of circadian rhythm regulation and the physiological substrates underlying rhythmicity in nocturnal rodents, but relatively few studies have addressed circadian rhythm regulation in other mammalian taxonomic groups. In this study, some formal and functional aspects of circadian organization in a nocturnal dasyurid marsupial, the stripe-faced dunnart (Sminthopsis macroura), were analyzed. To determine phasic responses to discrete pulses of light, dunnarts were placed in constant darkness (DD) and were periodically administered pulses of bright light at different times of the animals' circadian day. Analysis of phase shifts in response to light indicated a phase response curve that was similar to responses observed in nocturnal rodents. To determine the possibility of extraretinal photoreception mediating photic entrainment, dunnarts were anesthetized and orbitally enucleated while maintained in a light-dark regimen (LD 14:10). All blinded dunnarts free-ran with periods (tau) that were similar to those observed in DD, indicating that entrainment is mediated through ocular photoreception. However, the data also indicated a decrease in activity in blind dunnarts during the last 3-5 hr of the dark phase, raising the possibility of some retention of photoreceptive capacities.     

A "melanopic" spectral efficiency function predicts the sensitivity of melanopsin photoreceptors to polychromatic lights

Photoreception in the mammalian retina is not restricted to rods and cones but extends to a small number of intrinsically photosensitive retinal ganglion cells expressing the photopigment melanopsin. These mRGCs are especially important contributors to circadian entrainment, the pupil light reflex, and other so-called nonimage-forming (NIF) responses. The spectral sensitivity of melanopsin phototransduction has been addressed in several species by comparing responses to a range of monochromatic stimuli. The resultant action spectra match the predicted profile of an opsin:vitamin A-based photopigment (nomogram) with a peak sensitivity (λ(max)) around 480 nm. It would be most useful to be able to use this spectral sensitivity function to predict melanopsin's sensitivity to broad-spectrum, including "white," lights. However, evidence that melanopsin is a bistable pigment with an intrinsic light-dependent bleach recovery mechanism raises the possibility of a more complex relationship between spectral quality and photoreceptor response. Here, we set out to empirically determine whether simply weighting optical power at each wavelength according to the 480-nm nomogram and integrating across the spectrum could predict melanopsin sensitivity to a variety of polychromatic stimuli. We show that pupillomotor and circadian responses of mice relying solely on melanopsin for their photosensitivity (rd/rd cl) can indeed be accurately predicted using this methodology. Our data therefore suggest that the 480-nm nomogram may be employed as the basis for a new photometric measure of light intensity (which we term "melanopic") relevant for melanopsin photoreception. They further show that measuring light in these terms predicts the melanopsin response to light of divergent spectral composition much more reliably than other methods for quantifying irradiance or illuminance currently in widespread use.