Seasonal variations in circadian rhythms of plasma melatonin in ruin lizards

We examined melatonin profiles of ruin lizards in different seasons (spring, summer, and autumn) under light:dark (LD) and concomitant responses when transferred to continuous darkness (DD) to determine the degree to which previously reported seasonally dependent effects of pinealectomy on locomotor behavior are related to melatonin secretion. The amplitude of the melatonin rhythm and the amount of melatonin produced over 24 h varied with season. In spring, the amount of melatonin produced was greatest and the amplitude 4- 5 times that found in summer or autumn. The degree of self-sustainment of the melatonin rhythm when transferred to DD also varied with season. In DD, melatonin levels remained high but did not exhibit circadian variation in spring. In summer, the melatonin profile persisted virtually unchanged in DD, showing the existence of a circadian rhythm. Finally, in the fall there was no circadian variation in DD and levels remained low. These responses correspond closely to previously reported effects of pinealectomy on locomotor behavior where there is little or no effect of pinealectomy in spring or fall but a profound alteration of locomotor behavior in summer. These results suggest that the seasonally dependent effects of pinealectomy on locomotor behavior in ruin lizards are related to a seasonally mediated change in the degree of self-sustainment of some component of the circadian pace-making system of which melatonin plays some role.     

Influence of seasons on circadian rhythm of acid phosphatase and beta-acetyl-glucosaminidase activity in the mouse salivary gland and liver

Influence of seasons on circadian changes in acid phosphatase and beta-acetylglucosaminidase activity was studied in the liver and submandibular gland of sexually mature mice. Seasonal differences in circadian rhythm was found in both examined enzymes in both organs. These changes were independent from each other and each rhythm was differently subjected to seasonal changes. The lowest seasonal influence was observed in acid phosphatase in the salivary gland. Activity of the other enzymes changed in different degrees, having their acrophases at different times of the day and different rhythm intensity. The highest circadian activity changes measured by amplitude and mesor were observed in spring and summer whereas in the autumn and winter their activity had much weaker rhythms or even they disappeared completely. An attempt was made to explain the observed changes by changes in hormonal background and a certain kind of genetic memory pertaining to laboratory animals.     

Circadian rhythms of locomotor activity and temperature selection in sleepy lizards, <Emphasis Type="Italic">Tiliqua rugosa</Emphasis>

This study examined whether the daily rhythms of locomotor activity and behavioural thermoregulation that have previously been observed in Australian sleepy lizards (Tiliqua rugosa) under field conditions are true circadian rhythms that persist in constant darkness (DD) and whether these rhythms show similar characteristics. Lizards held on laboratory thermal gradients in the Australian spring under the prevailing 12-hour light : dark (LD) cycle for 14 days displayed robust daily rhythms of behavioural thermoregulation and locomotor activity. In the 13-day period of DD that followed LD, most lizards exhibited free-running circadian rhythms of locomotor activity and behavioural thermoregulation. The predominant activity pattern displayed in LD was unimodal and this was retained in DD. While mean levels of skin temperature and locomotor activity were found to decrease from LD to DD, activity duration remained unchanged. The present results demonstrate for the first time that this species’ daily rhythm of locomotor activity is an endogenous circadian rhythm. Our results also demonstrate a close correlation between the circadian activity and thermoregulatory rhythms in this species indicating that the two rhythms are controlled by the same master oscillator(s). Future examination of seasonal aspects of these rhythms, may, however, cause this hypothesis to be modified.     

The effect of dorsal raphe nucleus (DRN) lesions on the locomotor activity rhythm in mice.

The study employed electrical lesions of dorsal raphe nucleus (DRN) to determine the functional significance of those nuclei in the regulation of wheel-running activity rhythm in mice in light/dark (LD 12:12), constant light (LL), and constant dark (DD) conditions. The wheel-running records showed that raphe nucleus lesions resulted in few days' decrease in common activity and amplitude in LD. The activity phase was not compact but in fragmentary form, especially in DD condition. In some animals an earlier onset of activity after DRN lesion in LD was observed. In LL extension of the rhythm period occurred. Destruction of DRN only slightly modulates the wheel-running circadian rhythm in mice.     

Interseasonal variation in the circadian rhythms of locomotor activity and temperature selection in sleepy lizards, <Emphasis Type="Italic">Tiliqua rugosa</Emphasis>

Few studies in non-mammalian vertebrates have examined how various effectors of the circadian system interact. To determine if the daily locomotor and behavioural thermoregulatory rhythms of Tiliqua rugosa are both controlled by the circadian system in different seasons, lizards were tested in laboratory thermal gradients in four seasons and in constant darkness. Circadian rhythmicity for both rhythms was present in each season, being most pronounced in spring and summer and least evident in autumn. Most lizards displayed a unimodal locomotor activity pattern across all seasons. However, some individuals presented a bimodal locomotor activity pattern in spring and summer. Seasonal variations in the phase relationships of both rhythms to the light:dark (LD) cycle were demonstrated. No seasonal differences in the free-running period lengths of either rhythm were detected, raising the possibility that a single circadian pacemaker drives both rhythms in this species. Our present results demonstrate that both rhythms are similarly controlled by the circadian system in each season. Although seasonal variations in the thermal preferences of reptiles both in the field and laboratory have previously been well documented, this study is the first to demonstrate circadian rhythms of temperature selection in a reptile species in each season.     

Serum cortisol levels in goats exhibit seasonal but not daily rhythmicity

The aim of the study was to find out whether there is a daily rhythm in goat serum cortisol concentrations, whether the concentration profiles differ between normal light:dark and constant dark conditions, and whether any seasonal variations might be detected in daily cortisol secretion patterns. Seven Finnish landrace goats were kept at indoor temperature (18-23°C) under artficial lighting that approximately simulated the annual changes of daylength at 60°N. Blood samples were collected for cortisol measurements by radioimmunoassay at 2h intervals during six times of the year: winter (light:dark 6:18h), early spring (10:14h), late spring (14:10h), summer (18:6h), early fall (14:10h), and late fall (10:14h). Cortisol profiles were determined for two consecutive days, first in light:dark (LD) conditions and then in continuous darkness (DD). There was no significant daily rhythm in serum cortisol levels in any time of the year, nor did the profiles in LD and DD conditions show any differences. A significant seasonal variation was, however, detected among the overall cortisol levels. In winter, the concentrations were higher than in any other season, and from early spring to summer they were at their lowest. Under equal photoperiods, the cortisol levels were higher in fall than spring. The difference between winter and summer was confirmed in the following year in LD conditions. There was no correlation between the serum cortisol and progesterone levels. The results suggest that the possible circadian variation of cortisol secretion in goats is completely masked by external factors, and the lighting conditions do not have immediate effects on the daily secretion patterns. The seasonal variation in the overall cortisol levels is most probably related to the changes in photoperiod, because other conditions were relatively constant during the experiment.     

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

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

Daily and annual variations of free fatty acid, glycerol and leptin plasma concentrations in goats (Capra hircus) under different photoperiods

The aim of the study was to differentiate the impact of lighting conditions and feeding times on the regulation of lipid metabolism of goats under different photoperiods throughout the year. Seven Finnish landrace goats were kept under artificial lighting that simulated the annual changes of photoperiod at 60 degrees N (the longest light period 18 h, the shortest 6 h). Ambient temperature and feeding regime were kept constant. Blood samples were collected six times a year at 2-h intervals for 2 days, first in light/dark (LD) conditions and then after 3 days in constant darkness (DD). Significant daily variations were detected in the concentrations of plasma free fatty acids (FFA) and glycerol throughout the year. The nocturnal decrease and morning rise of FFA levels were related to the photoperiod, while the trough levels of glycerol were associated with the concentrate meal times. In DD conditions, FFA and glycerol rhythms were unstable. A significant seasonal variation was detected in the overall FFA and glycerol levels suggesting decreased lipogenesis in winter, increased lipolysis in spring and high lipogenesis in summer and fall. There was no significant daily rhythm in serum leptin levels, nor did the profiles in LD and DD conditions differ. The leptin level was slightly lower in early fall than in the other seasons, paralleling a small decrease of body mass in the goats after the grazing season. The daily or annual variations of FFA and glycerol levels were not clearly related to leptin concentrations. The results suggest that lipid metabolism of goats is regulated by light even in constant temperature and feeding conditions; however, no significant contribution of leptin levels could be shown.     

Seasonal variation in endogenous serum melatonin profiles in goats: a difference between spring and fall?

The pineal hormone melatonin serves as a signal of day length in the regulation of annual rhythms of physiological functions and behavior. The duration of high melatonin levels in body fluids is proportional to the duration of the dark period of the day. Due to the direct suppression of melatonin by light, the overt melatonin rhythm may differ from the endogenous rhythm driven by the hypothalamic circadian clock. The aim of this study was to find out possible differences between the overt and endogenous melatonin rhythms in goats during the course of a year. Seven Finnish landrace goats (nonlactating females) were kept under artificial lighting that approximately simulated the annual changes of day length at 60 degrees N. Blood samples for melatonin measurements by radioimmunoassay were collected at 2-h intervals during six seasons: winter (light:dark 6:18 h), early spring (10:14), late spring (14:10), summer (18:6), early fall (14:10), and late fall (10:14). Melatonin profiles were determined for 2 consecutive days, first in light-dark (LD) conditions and then in continuous darkness (DD). In LD conditions, the profiles matched the dark period with one exception: In winter, the mean peak duration was significantly shorter than the scotoperiod. In DD conditions, two types of endogenous melatonin patterns were found: a "winter pattern" (peak duration 13-15 h) in winter, early spring, early fall, and late fall, and a "summer pattern" (duration about 11 h) in late spring and summer. Thus, in equal habitual LD conditions in late spring and early fall (LD 14:10), the endogenous melatonin rhythms were not quite similar: The pattern in late spring resembled that in summer, and the pattern in early fall that in winter. These results suggest that, in addition to the light-adjusted overt melatonin rhythm, the endogenous rhythm of melatonin secretion varies during the course of a year.     

Circadian Locomotor Activity Rhythm in the Freshwater Crab Pseudothelphusa americana (De Saussure, 1857): Effect of Eyestalk Ablation

The circadian rhythm of locomotor activity in the freshwater crab, Pseudothelphusa americana , was studied in aquaria using infrared crossing sensors. Individuals with ablated eyestalks were compared with intact individuals in constant darkness (DD) and in light-dark cycles (LD). Our results showed that intact animals in DD displayed bimodal rhythms. In LD conditions the two peaks were associated with lights on and lights off, respectively. A significant difference in the free running periods before and after LD was observed in all intact animals. After eyestalk ablation (ES-X), the circadian rhythm of locomotor activity disappeared immediately, but reappeared several days later. Diurnal activity was seen in some ES-X animals when exposed to LD. Our results indicate that locomotor activity rhythm in P. americana is driven primarily by oscillators located outside the eyestalks, and that extraretinal photoreceptors mediate either entrainment or masking effects.     

Circadian Locomotor Activity Rhythm in the Freshwater Crab Pseudothelphusa americana (De Saussure, 1857): Effect of Eyestalk Ablation

The circadian rhythm of locomotor activity in the freshwater crab, Pseudothelphusa americana, was studied in aquaria using infrared crossing sensors. Individuals with ablated eyestalks were compared with intact individuals in constant darkness (DD) and in light-dark cycles (LD). Our results showed that intact animals in DD displayed bimodal rhythms. In LD conditions the two peaks were associated with lights on and lights off, respectively. A significant difference in the free running periods before and after LD was observed in all intact animals. After eyestalk ablation (ES-X), the circadian rhythm of locomotor activity disappeared immediately, but reappeared several days later. Diurnal activity was seen in some ES-X animals when exposed to LD. Our results indicate that locomotor activity rhythm in P. americana is driven primarily by oscillators located outside the eyestalks, and that extraretinal photoreceptors mediate either entrainment or masking effects.     

Short-term and circadian rhythms in the behaviour of the vole,Microtus agrestis (L.)

Summary The activity behaviour of the vole, Microtus agrestis, has been recorded in order to investigate the relationship between short-term rhythm and circadian rhythm. A simple device was developed, allowing separate monitoring of the time spent in or outside the nest, wheel-running, eating and drinking. Under natural light conditions during summer, a distinct differentiation between a short term rhythm of eating and drinking during the day-time and a circadian rhythm of wheel-running during the night was observed. The short-term rhythm depends closely on metabolic demands (hunger, thirst, excretion). Control of these demands by an endogenous oscillation could not be substantiated. The circadian rhythm of wheel-running activity is, however, controlled by an endogenous oscillation, synchronized by light conditions. It is subjected to seasonal variations. a) The threshold of light intensity below which wheel-running occurs is lowest during summer (<0.5 lx) and is higher during spring and autum (> 5 lx). b) Wheel-running is controlled by a circadian oscillation during summer only whereas it is an integrated part of the short-term rhythm during spring and autumn (experiments during the winter have not yet been performed). Experiments gave evidence that the properties of the cage can deeply influence the amount and pattern of wheel-running activity. It is concluded that wheel-running reflects a certain level of excitation, which may be caused by different behavioural intentions. The seasonal changes of the control of wheel-running activity are discussed with respect to this assumption. The relevancy of locomotor activity patterns as usually recorded in the laboratory to reveal the physiological and ecological significance of endogenously controlled behavioural patterns is discussed.Supported by the Deutsche Forschungsgemeinschaft     

Circadian clock controlling egg hatching in the cricket (Gryllus bimaculatus)

Adult crickets (Gryllus bimaculatus) were maintained under a 12-h light:12-h dark cycle (LD 12:12). After oviposition, their eggs were incubated under different lighting regimens at 23 degrees C, and temporal profiles of egg hatching were examined. When the eggs were incubated in LD 12:12 or in DL 12:12 with a phase difference of 12h from LD 12:12, throughout embryogenesis, 88% to 97% of hatching occurred within 3 h of the dark-light transition on days 17 and 18 of embryogenesis; the phases of the egg-hatching rhythms in the LD 12:12 and DL 12:12 groups differed by about 12 h. In eggs incubated in constant darkness (DD) throughout embryogenesis, a circadian (about 24 h) rhythm of hatching was found, and the phase of the rhythm was similar to that seen in eggs incubated in LD 12:12, but not DL 12:12, throughout embryogenesis. When eggs that had been incubated in DD after oviposition were transferred to DL 12:12 in the middle or later stages of embryogenesis and were returned to DD after three cycles of DL 12:12, the rhythm of hatching synchronized (entrained) to DL 12:12. However, when eggs in the earlier stages of embryogenesis were transferred from DD to DL 12:12 and returned to DD after three cycles, 52% to 94% of hatching did not entrain to DL 12:12. To determine whether photoperiodic conditions to which the parents had been exposed influenced the timing of egg hatching, adult crickets were maintained in DL 12:12, and their eggs were incubated in LD 12:12, DL 12:12, or DD throughout embryogenesis. The egg-hatching rhythm was also found in the eggs incubated under these three lighting regimens. In DD, the phase of the rhythm was similar to that seen in eggs incubated in DL 12:12, not LD 12:12, throughout embryogenesis. The results indicate that in the cricket, the timing of egg hatching is under circadian control and that the circadian rhythm of hatching entrains to 24-h light:dark cycles, but only if the light:dark cycles are imposed midway through embryogenesis. Therefore, by midembryogenesis, a circadian clock has been formed in the cricket, and this is entrainable to light:dark cycles. In addition, the photoperiodic conditions to which the parents (probably the mothers) have been exposed influence the timing of hatching, suggesting that maternal factors may regulate the timing of egg hatching.     

Changes in melatonin binding sites under artificial light-dark, constant light and constant dark conditions in the masu salmon brain

To test whether the affinity (Kd) and total binding capacity (Bmax) of melatonin receptors exhibit daily and circadian changes in teleost fish whose melatonin secretion is not regulated by intra-pineal clocks, we examined the changes in melatonin binding sites in the brains of underyearling masu salmon Oncorhynchus masou under artificial light-dark (LD), constant light (LL) and constant dark (DD) conditions. In Experiment 1, fish were reared under a long (LD 16:8) or short (LD 8:16) photoperiod for 69 days. Blood and brains were sampled eight times at 3 h intervals. Plasma melatonin levels were high during the dark phase and low during the light phase in both photoperiodic groups. The Bmax exhibited no daily variations. Although the Kd slightly, but significantly, changed under LD 8:16, this may be of little physiological significance. In Experiment 2, fish reared under LD 12:12 for 27 days were exposed to LL or DD from the onset of the dark phase under LD 12:12. Blood and brains were sampled 13 times at 4 h intervals for two complete 24 h cycles. Plasma melatonin levels were constantly high in the DD group and low in the LL group. No significant differences were observed in the Kd and the Bmax between the two groups, and the Kd and the Bmax exhibited no circadian variation either in the LL or DD groups. These results indicate that light conditions have little effect on melatonin binding sites in the masu salmon brain.     

Turkey retina and pineal gland differentially respond to constant environment

Dynamics of rhythmic oscillations in the activity of arylalkylamine N-acetyltransferase (AA-NAT, the penultimate and key regulatory enzyme in melatonin biosynthesis) were examined in the retina and pineal gland of turkeys maintained for 7 days in the environment without daily light-dark (LD) changes, namely constant darkness (DD) or continuous light (LL). The two tissues differentially responded to constant environment. In the retina, a circadian AA-NAT activity rhythm disappeared after 5 days of DD, while in the pineal gland it persisted for the whole experiment. No circadian rhythm was observed in the retinas of turkeys exposed to LL, although rhythmic oscillations in both AA-NAT and melatonin content were found in the pineal glands. Both tissues required one or two cycles of the re-installed LD for the full recovery of the high-amplitude AA-NAT rhythm suppressed under constant conditions. It is suggested that the retina of turkey is less able to maintain rhythmicity in constant environment and is more sensitive to changes in the environmental lighting conditions than the pineal gland. Our results indicate that, in contrast to mammals, pineal glands of light-exposed galliformes maintain the limited capacity to rhythmically produce melatonin.     

Temporal Organization in Locomotor Activity of the Hypogean Loach, Nemacheilus Evezardi, and its Epigean Ancestor

Locomotor activity rhythm in the hypogean population of Nemacheilus evezardi was recorded first under light-to-dark (LD) 12 : 12 h cycle and then DD. The results were compared with that of its epigean counterpart held under comparable regimes. In LD 12 : 12, while hypogean loach exhibited a distinct bimodality in its locomotor activity rhythm, it was altogether absent in the case of epigean population. In hypogean loach, dark-to-light transition peak in LD was observed to free-run under DD. The same was not discernible in case of epigean loach. The locomotor activity rhythm in epigean fish was noticed to free-run in DD either from the dawn peak or dusk peak in LD. It is hypothesized that the hypogean fish still possesses a functional oscillator underlying its overt circadian rhythm in locomotor activity. The ecophysiological significance of these findings is yet to be fully understood.     

Circannual variation of intestinal cell proliferation in BDF1 male mice on three lighting regimens

BDF, male mice were studied over a 24-hr span in winter, spring, summer and fall. For three weeks prior to study, one-third of the animals were kept under a lighting regimen of 8 hr light alternating with 16 hr of darkness (LD 8:16), one-third on a lighting regimen of LD 12:12 and a remainder on a lighting regimen of LD 16:8. During each study, subgroups of animals on all three lighting regimens were killed at 4-hr intervals over a 24-hr span. Twenty minutes prior to being killed, the animals received 5yCi of [³Hthymidine/0.2 ml/20 gm of body weight intraperitoneally. The thymidine uptake in the DNA of the colon and of the small intestine were studied as an index of cell proliferation. A circadian rhythm in [³H]-thymiduie uptake in the colon was found and validated by cosinor analysis. This rhythm was similar in acrophase and amplitude in the animals kept on LD 8:16 and LD 12:12. Also in the mice on LD 16:8, there was a statistically significant circadian rhythm of ('HJ-thymidtne uptake in the DNA of the colon during all four seasons. The acrophases of this rhythm, however, varied widely suggesting free running. A circadian rhythm of pHJ-thymidine uptake in small intestine was less consistent. In animals on all three lighting regimens, however, a circannual variation of f'HJ-thymidine uptake in DNA in colon and small intestine was found with the highest uptake during summer. This study indicates that a lighting regimen of LD 16:8 does not reliably synchronize the circadian rhythm of [³H]-thymidine uptake in the colon. It further shows a circannual rhythm of this function in the colon and in the small intestine which persists under three lighting regimens (LD 8:16, 12:12 and 16:8) maintained for three to four weeks prior to being killed.     

Seasonal and task-related variation in free running activity rhythms in honey bees (Apis mellifera)

We measured seasonal variation in the locomotor behavior of newly emerged adult honey bee workers in the laboratory. Analyses of bees from 12 colonies, 7 of which were tested once and 5 tested more than once, revealed seasonal changes in the free-running period (FRP) of the rhythm for locomotor behavior, with an increase from spring to summer. At the same time there was a decrease in the age at onset of circadian rhythmicity. There were no seasonal changes in overall levels of locomotor activity. Temperature and photoperiod, the only factors known to mediate plasticity in the insect clock, cannot account for the observed seasonal variation because bees were maintained under constant conditions. In a second experiment we found no differences in the FRP of nurses and foragers obtained from colonies maintained in a 12 h light: 12 h dark illumination regime. These findings suggest that exposure to unknown cues during preadult stages may affect the circadian behavior of adult bees. Received 7 April 2005; revised 30 August 2005; accepted 1 September 2005.     

Circadian rhythms in heart rate, motility, and body temperature of wild-type C57 and eNOS knock-out mice under light-dark, free-run, and after time zone transition

The nitric oxide (NO) system is involved in the regulation of the cardiovascular system in controlling central and peripheral vascular tone and cardiac functions. It was the aim of this study to investigate in wild-type C57BL/6 and endothelial nitric oxide synthase (eNOS) knock-out mice (eNOS-/-) the contribution of NO on the circadian rhythms in heart rate (HR), motility (motor activity [MA]), and body temperature (BT) under various environmental conditions. Experiments were performed in 12:12 h of a light:dark cycle (LD), under free-run in total darkness (DD), and after a phase delay shift of the LD cycle by -6 h (i.e., under simulation of a westward time zone transition). All parameters were monitored by radiotelemetry in freely moving mice. In LD, no significant differences in the rhythms of HR and MA were observed between the two strains of mice. BT, however, was significantly lower during the light phase in eNOS-/- mice, resulting in a significantly greater amplitude. The period of the free-running rhythm in DD was slightly shorter for all variables, though not significant. In general, rhythmicity was greater in eNOS-/- than in C57 mice both in LD and DD. After a delay shift of the LD cycle, HR and BT were resynchronized to the new LD schedule within 5-6 days, and resynchronization of MA occurred within 2-3 days. The results in telemetrically instrumented mice show that complete knock-out of the endothelial NO system—though expressed in the suprachiasmatic nuclei and in peripheral tissues—did not affect the circadian organization of heart rate and motility. The circadian regulation of the body temperature was slightly affected in eNOS-/- mice.