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1 Laboratoire Rythmes
Biologiques et Chronothérapeutique, Circadian rhythms in circulating
leukocyte and lymphocyte counts persisted with halved amplitudes in
constant light (LL) of 300 lx intensity for 8 wk, whereas circadian
rhythms in body temperature, locomotor activity, and plasma
catecholamines were completely suppressed. Subsequent exposure to
constant darkness (DD) normalized all circadian rhythms within 2 wk.
Rhythms in circulating T lymphocyte subsets were studied in LL or DD
using double labeling with monoclonal antibodies and flow cytometry.
Circadian rhythms were suppressed for leukocytes and lymphocytes but
were maintained for both T helper cells (Th) and T cytotoxic cells (Ts)
lymphocytes after 11 wk in LL. A group 24-h rhythm was only validated
for total lymphocytes after 16 wk in LL. However, individual total, Th, and Ts lymphocytes maintained their usual respective phase
relationships in each rat. The alteration of immune cell circulatory
rhythms likely stemmed from a progressive loss of circadian
synchronization among rats kept in LL. Conversely, after 11 or 16 wk in
DD, leukocytes and lymphocyte subsets circadian rhythms were
maintained. Thus catecholamines do not drive circulatory T cell
rhythms. The loss of coupling between T lymphocyte rhythms and three
major outputs of the circadian system further supports the hypothesis
of an independent immunologic oscillator.
circadian rhythms; desynchronization; body temperature; locomotor
activity; leukocytes; catecholamines
SEVERAL STUDIES HAVE demonstrated circadian (~24 h)
rhythms in circulating counts and functions of lymphoid cells and their subsets in mammals, including humans (19). Circulating T lymphocyte subsets usually reach a maximum during the rest span, whereas circulating natural killer (NK) cells and lytic activity usually peak
near the beginning of the active phase (1, 10, 13). Such circadian
organization of the immune system was hypothesized as a coordinated
clock (20).
Circadian rhythms persist in the absence of any known external periodic
signal and thus are generated by endogenous pacemakers (2, 12, 30). The
circadian period of these so-called free-running rhythms slightly
deviates from 24 h in animals kept in constant darkness (DD) and can be
lengthened or shortened according to species and individual (5, 27).
The suprachiasmatic nucleus (SCN), located in the anterior
hypothalamus, is considered the main central circadian clock of
mammals. It displays circadian rhythms in metabolic and electrical
activity both in vitro and in vivo and drives a great number of
behavioral and physiological rhythms, such as locomotor activity, food
intake, body temperature, and several neuroendocrine secretions (15).
On the other hand, immune responses are modulated by a neuroendocrine
network (31). Specific receptors for epinephrine, norepinephrine,
dopamine, and melatonin were found on the surface of T, B, NK,
and/or other lymphatic cells (11, 32). In addition, a circadian
rhythm in lymphocyte adrenoreceptor density was reported with a peak
around noon and a trough around midnight in human subjects (26). As a
result, melatonin and adrenergic hormones can exert enhancing or
suppressing effects on immune functions, which possibly vary along the
24-h time scale.
Prolonged exposure of rats to constant light (LL) achieved a mean
sixfold decrease in plasma melatonin and a marked dampening but no
abolition of its circadian rhythmicity. Nevertheless, circadian rhythms
in body temperature and locomotor activity were completely suppressed
in these conditions (8). Subsequent DD exposure normalized all three
rhythms within 2 wk. In this model of functional suppression of the
circadian system, prolonged LL could impair the transduction pathways
from the SCN to different outputs or stop the oscillator, thus
resulting in an uncoupling of several components of a multioscillator
system (27, 36).
LL exposure has also been associated with alterations in rodent immune
system (23, 24). The hypothesis of an independent immunologic circadian
oscillator would be supported by the observation of ~24-h rhythms in
immune variables in the absence of catecholamines, body temperature, or
locomotor activity circadian rhythms. The rat model of prolonged LL
exposure was chosen for such purpose, with animals kept in prolonged DD
serving as controls.
Animals and Housing
ABSTRACT
Top
Abstract
Introduction
Methods
Results
Discussion
References
INTRODUCTION
Top
Abstract
Introduction
Methods
Results
Discussion
References
MATERIAL AND METHODS
Top
Abstract
Introduction
Methods
Results
Discussion
References
Temperature and Activity Monitoring
Intraperitoneal temperature and locomotor activity were monitored every 10 min by telemetry using Dataquest III software (Datasciences, St. Paul, MN). A TA10TA-F40 sensor (Datasciences) was implanted under ether anesthesia.Experimental Procedures
Experiment 1. Two groups of eight rats each (aged 4 mo and weighing on average 420 g, range 360-540 g, on arrival) were exposed to LL for 8 wk. Intraperitoneal temperature and locomotor activity were monitored in 12 rats (6 per group). All animals from each group had blood samples taken at six different times of day, staggered by 4-8 h over 32 h. Blood was obtained via a jugular catheter implanted under thiopental anesthesia (50 mg/kg) 2 days before the first sampling time point. Six rats from the second group were subsequently switched to DD for 2 wk, then submitted to the same blood-sampling procedure. Blood was also obtained from a third group of 13 age-matched rats, synchronized with an alternation of 12 h of light and 12 h of darkness (LD12:12), with light from 0800 to 2000 (Fig. 1, top). Blood was collected on EDTA-K3, leukocytes were counted immediately, and plasma was stored at
80°C until assay.
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Experiment 2. Twelve rats aged ~4 mo and weighing on average 490 g on arrival (range 420-550 g) were first synchronized with LD12:12 (light from 0700 to 1900) for 2 wk, then randomized into 2 groups, which were both exposed to constant conditions for 17 wk. The first group was maintained in LL, and the second one was kept in DD. Both groups were subsequently switched back to LD12:12 for 16 additional weeks. Blood samples (1 ml) were obtained at six circadian times staggered over 48 h on four occasions in each rat, after both 11 and 16 wk in LL or in DD and after 4 and 16 wk following the switch to LD12:12 (Fig. 1, bottom). Tail vein punctures were performed within 3 min with the use of an immobilization bandage. Blood was collected on EDTA-K3 and assayed immediately for leukocyte and lymphocyte subset counts.
Catecholamine assays. Plasma epinephrine, norepinephrine, and dopamine concentrations were determined with the use of liquid chromatography with electrochemical detection. The mobile phase (50 mM sodium acetate, 20 mM citric acid, 35 mM dibutylamine, 7% methanol) was added with octane sulfonate 3.75 mM for a reverse phase separation on an optical density scanner 5-µm column. Instrumental control and data acquisition, storage, and retrieval functions were performed with a Maxima 820 Chromatography Workstation (Waters, St. Quentin en Yvelines, France). Results were expressed as nanomoles per liter.
Blood cell counts and lymphocyte subsets
analysis. Circulating leukocytes were counted by
Coulter Counter ZM (Coultronics, Margency, France); differential count
was analyzed from blood smears. Lymphocyte subsets were measured with a
flow cytometer (ACR 1500 SP Bruker Spectrospin, Wissembourg, France)
using anti-rat lymphocyte monoclonal agents [CD3+: clone G4.18,
ref. 22015B; CD4+: clone OX-35, ref. 22024D; CD8+: clone OX-8, ref.
22074D; controls by mouse immunoglobulin (Ig)
G1, ref. 03004C, and mouse IgG2a, ref. 03025A; Pharmingen,
Clinisciences, Montrouge, France]. Phenotypes were chosen for
characterizing the lymphocyte subset according to presence or absence
of specific differentiation clusters at the cell surface: CD3+, all T
lymphocytes (T3); CD3+4+, T
helper cells (Th); CD3+8+, T cytotoxic cells (Ts); CD38+, NK
cells. Cell labeling was performed after whole blood lysis (25).
Briefly, 1 ml of blood was lysed in 20 ml of a lysing buffer (Ortho),
then washed two times with 20 ml of phosphate-buffered saline with azide (PBSa; 1 g/l) and resuspended in 1.5 ml of PBSa. Cell aliquots (50 µl) and monoclonal antibodies (MAbs) were allowed to incubate 30 min at 4°C, according to previous dilution studies (10 µl of 0.5-0.2 MAb dilution). Cells were then washed twice to eliminate excess of MAbs in PBSa. Percentages of MAb-labeled cells were quantitated by flow cytometry, and lymphocyte subsets counts were then
calculated (total lymphocyte count × proportion of labeled cells).
Statistical analyses. Temperature and activity data were analyzed with a serial section of 7-day time series, shifted with 3-day intervals using Dataquest III software. First power spectrum analysis (Fourier transform) was applied to 30-min average data intervals. Then least-squares cosine regression was applied for test periods, differing by 5 min within the range of the dominant period. This method tests the probability of the null amplitude hypothesis for a chosen period and gives the rhythm parameters (e.g., amplitude, one-half of the difference between peak and trough of fitted cosine function, and acrophase, time of maximum of fitted cosine function). The period that corresponded to the highest percent of rhythm (percent of the variability accounted for by fitted cosine function) was considered as the dominant one if P < 0.001. Lymphocyte subsets (Th, Ts, and NK cells) were analyzed both as raw counts and as percentages of the individual mean of each. By doing so, variability in mean level among rats was eliminated, thus allowing for comparisons of rhythmicity. The statistical significance of differences between groups or time points was tested with analysis of variance. Least-squares cosine regression was used to validate circadian or ultradian rhythms in individual and group data. Individual peak-trough differences (dPT) were also computed and compared with Student's t-test.
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RESULTS |
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Top
Abstract
Introduction
Methods
Results
Discussion
References
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Body Temperature and Locomotor Activity
Mean values in body temperature or in locomotor activity were similar in rats exposed to LD12:12, LL, or DD in both experiments (Table 1). Circadian rhythms were found for both variables in rats kept in LD12:12 (Fig. 2).
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Mean dominant periods of temperature and activity under LL were ~7 h for both variables in experiment 1, and 6 and 4.9 h, respectively, in experiment 2. Such desynchronization in the ultradian domain might result from increased LL exposure in the latter experiment. Under prolonged LL for more than 10 wk, one of three rats had arrythmic locomotor activity, whereas temperature rhythm remained ultradian.
Temperature rhythms had lower amplitudes for rats kept in LL (average = 0.16°C, in the ultradian domain) than for rats kept in DD (average = 0.41°C, circadian) or in LD12:12 (0.65°C, circadian). Conversely, mean amplitudes of locomotor activity rhythms were similar in rats kept in LL (average = 39% of mean) or in DD (36% of mean) despite differences in period domains. They were, however, significantly lower than those of rats kept in LD12:12 (72% of mean). The dominant period in both temperature and activity switched from ultradian in LL to circadian within 2 wk of DD.
Under prolonged DD exposure, body temperature circadian rhythms persisted, yet with approximately halved amplitude compared with LD12:12 (0.33°C and 0.65°C, respectively). The circadian rhythm in locomotor activity was suppressed in one of the three monitored rats after 10 wk of DD exposure. Both body temperature and locomotor activity circadian rhythms were restored and/or normalized within 1 wk after all animals were switched from LL or DD to LD12:12.
Catecholamines
Mean concentrations of epinephrine and norepinephrine were lower in LL- or DD- exposed rats compared with those kept in LD12:12. No such difference was found for dopamine (Table 2).
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Although dopamine plasma concentrations were significantly higher during light than during darkness in rats kept in LD12:12, no sinusoidal 24-h rhythm was validated with cosinor. After LL for 8 wk, the dopamine daily variation was abolished. After the switch to DD, dopamine plasma concentrations were higher from 1600 to 2400 than from 0400 to 1200, without any sinusoidal 24-h rhythm. Thus, although the 24-h variation was restored in DD, its maximum was shifted by 8 h compared with LD12:12.
Circadian rhythms were found for plasma epinephrine and norepinephrine concentrations in rats maintained under LD12:12. Their amplitudes were ~60% of the individual mean, and their acrophases were located near 0600, close to the end of the activity span. After LL for 8 wk, both epinephrine and norepinephrine circadian rhythms were suppressed. Two weeks of DD exposure restored both rhythms and normalized their relative amplitudes (as percent of the 24-h mean) and peak times (Fig. 3).
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Leukocytes
Mean counts of circulating leukocytes were similar in rats kept in LL or DD compared with their age-matched rats synchronized with LD12:12. Animals kept under this latter regimen displayed a circadian rhythm in this variable with a maximum located near 1100, e.g., the early rest span. This rhythm persisted after 8 wk of LL, with a similar location of its maximum. Indeed, if leukocyte rhythm was free running with
24.5 h, ~8 wk would be needed for the maximum to occur at the
same local clock hour. In addition, mean individual dPT was reduced
compared with LD12:12 (42 vs. 62%). DD exposure for 2 wk normalized
the leukocyte circadian rhythm. Amplitude and time of maximum were
similar to those observed in rats synchronized with LD12:12, and mean
dPT was restored to 70 ± 5% (Fig. 4).
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The group leukocyte circadian rhythm was suppressed after LL exposure for 11 or 16 wk. It was not detected after 11 wk in DD either, but was found after 16 wk in DD, with a 17% amplitude. Its maximum was located near 2200, thus ~12 h phase shifted compared with LD12:12. Inspection of individual data suggested free-running rhythms in three of six rats after 11 wk that became synchronized with the other three rats 5 wk later.
The leukocyte rhythm of rats previously exposed to DD was normalized after the switch to LD12:12 for 4 or 16 wk: mean amplitude was ~30% and its maximum was again located near 1100, i.e., in the early rest span. However, rats previously exposed to LL showed persistent loss of synchronization among themselves, despite being exposed to LD12:12 for up to 16 wk.
Lymphocyte Counts and Subsets
Mean circulating total lymphocyte counts were similar in rats kept in LL or DD compared with their age-matched LD rats. A group circadian rhythm was demonstrated in rats exposed either to LD12:12 or to LL for 8 wk or to DD for 2 wk. Amplitudes ranged from ~22% in LD12:12 to 25% in DD and 15% in LL. Maxima were located near 1100.Lymphocyte subpopulations were examined in the second study. Mean counts of lymphocytes and their Th and Ts subsets were similar, irrespective of environmental condition. However, mean (±SE) NK cell count was ~40% lower after exposure to LL for 11 or 16 wk (0.44 ± 0.05 and 0.32 ± 0.04 × 109/ml, respectively) compared with DD for 11 or 16 wk (0.61 ± 0.06 and 0.45 ± 0.04 × 109/ml, respectively).
After LL exposure for 11 wk, circadian rhythms could not be established
for total lymphocyte count but were maintained for both Th and Ts
lymphocytes, with an ~30% amplitude. Maxima were located near 2000 (Fig. 5). After LL exposure for 16 wk, a
circadian rhythm was found for total lymphocytes, but not for Th or Ts
subsets. Inspection of individual time series suggested a loss of
synchronization among rats for Th and Ts cell counts. The phase
relationship of the Th and Ts rhythms appeared maintained after LL for
11 or 16 wk. The median differences (range) of times of peak values
were 4 h (0-8 h) after 11 or 16 wk of LL. NK cell count displayed
significant variations according to sampling hour after LL exposure for
11 or 16 wk as documented with analysis of variance
(P < 0.001 and P < 0.01, respectively). Cosinor
analysis validated ultradian rhythmicity in these conditions ( ~10-12 h).
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A group circadian rhythm in total lymphocyte count was not apparent
after DD exposure for 11 wk, yet it was found 5 wk later, with a
maximum located at 2200 and an amplitude of ~17%. Both Th and Ts
lymphocytes maintained a circadian rhythm after 11 and 16 wk of DD
exposure (Fig. 5). The median differences (range) of times of peak
values were 4 h (0-8 h) after 11 or 16 wk of DD. NK cells
displayed significant hourly variations after both 11 and 16 wk of DD
exposure. Cosinor analysis validated ultradian rhythms with ~12 h
after DD for 11 wk and ~8 h after DD for 16 wk
(P < 0.001 and
P < 0.01, respectively).
Total lymphocyte count displayed a normal circadian rhythm 4 wk after the switch from DD to LD12:12, with a mean (±SE) amplitude of 32 ± 5% and peak values at ~1030, i.e., the early rest span. No such rhythm was found in rats previously exposed to LL after being switched to LD12:12.
After 16 wk of LD12:12 exposure, mean total lymphocyte count varied significantly as a function of sampling time in rats previously exposed either to LL or to DD. However, a 24-h rhythm was only detected in those rats previously kept in DD. Inspection of individual time series suggested persistent loss of synchronization in rats previously exposed to LL.
Lymphocyte subsets were also studied after 16 wk in LD12:12. Mean values of Th and Ts lymphocyte counts were similar irrespective of previous LL or DD exposure. However, mean NK cell count remained significantly lower in the rats previously exposed to LL compared with those previously exposed to DD (0.4 ± 0.06 vs. 0.8 ± 0.1 × 109/ml ; P < 0.01). Both Th and Ts lymphocytes displayed significant variations according to sampling time in both groups (Fig. 6). However, by cosinor analysis, a circadian rhythm was validated only in rats previously exposed to DD. NK cells displayed significant variations according to sampling time only in rats previously exposed to DD. Despite persistent interindividual desynchronization for all other parameters studied in rats previously exposed to LL, the peak values of Th and Ts lymphocytes nearly coincided in each animal.1 The fact that individual Th and Ts peak values remained in phase suggests internal synchronization within each rat.
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DISCUSSION |
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Top
Abstract
Introduction
Methods
Results
Discussion
References
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Prolonged exposure to LL induced a complete suppression of circadian periodicity of both body temperature and locomotor activity. Switching the animals to DD restored circadian rhythms within 1 wk. These results are consistent with previous findings (8). Under prolonged DD exposure, persistent but altered circadian rhythms with approximately halved amplitudes were observed. The circadian period was close to 24 h, as was previously reported for another rat strain (12). Furthermore, in one of three rats kept in DD, the circadian rhythm of locomotor activity was suppressed after the 5th week of exposure, whereas that of temperature persisted. Although the possibility of an alteration of rat circadian rhythms in locomotor activity or temperature in LL was occasionally suggested, only a slight dampening of the 24-h activity rhythm was reported after DD exposure for 10 days in some strains of rats, but not in others (5, 7, 9, 12, 14). Under these constant conditions, a transient loss of coupling between temperature and activity rhythms was observed. The dominant period of locomotor activity rhythm shifted from the circadian domain to ultradian 2-5 wk earlier than that of temperature under LL. Conversely, the dominant period of temperature shifted 3-7 days earlier than activity from the ultradian domain to the circadian one in rats switched from LL to DD. Furthermore, a marked dampening but no abolition of plasma melatonin circadian rhythm was observed in LL (8). These reports suggested that prolonged LL exposure could uncouple multiple oscillators, an assumption made earlier (27, 28). The recent demonstration of a retinal clock in hamsters further strengthened the multioscillator theory (34).
The present study was carried out to evaluate the hypothesis of the existence of a special circadian oscillator involved in the regulation of immunologic functions. In a first experiment, we examined leukocyte and catecholamine rhythms in rats kept in LD12:12, LL, or DD. Catecholamines displayed significant circadian rhythms under LD12:12, with highest values near the end of the activity span. This is in agreement with previous reports, which indicate highest sympathetic tone in the activity span (3, 18). Leukocytes and lymphocytes also showed significant circadian rhythms with highest values at the beginning of the early rest span, thus confirming earlier reports (reviewed in Ref. 19). Catecholamine circadian rhythms were suppressed in rats maintained under LL, whereas leukocyte and lymphocyte rhythms persisted with a 50% decrease in amplitude. Two weeks after rats were switched to DD, all rhythms were partly or completely normalized. This suggests a coupling of catecholamine rhythms with the rhythms of body temperature and locomotor activity but an uncoupling from the hematoimmunologic circulatory rhythms under prolonged LL. Nevertheless, leukocyte circadian rhythm was suppressed if LL exposure lasted for 11 or 16 wk as shown in experiment 2, where rats were kept in prolonged constant environmental conditions (LL or DD).
The detailed individual and group patterns of circulating lymphocyte subsets were further examined in this second study. Only mean counts of NK cells were significantly lower under LL compared with DD. Circadian rhythms were suppressed for leukocytes and total lymphocytes but were maintained for Th and Ts subsets after LL exposure for 11 wk. Furthermore, the maxima in Th and Ts were coincident, as was observed in DD. Conversely, a low-amplitude group lymphocyte circadian rhythm was detected after 16 wk of exposure to LL, whereas group circadian rhythms of leukocytes and Th and Ts subsets were suppressed. Inspection of individual time series suggested a loss of synchronization among rats, whereas the synchronization of different rhythms within each rat was maintained. Total lymphocytes and T subsets displayed circadian rhythms after 16 wk of DD exposure. The lack of a group rhythm in total lymphocytes after DD exposure for 11 wk might be related to (transient) desynchronization among animals and/or interindividual variability.
The persistence of immunologic rhythms despite suppressed temperature and activity circadian rhythms, which was observed in LL, demonstrates an uncoupling between respective output variables from the immune system and from the circadian system. Furthermore, the presence of lymphocyte circadian rhythms up to 3 wk after the suppression of epinephrine, norepinephrine, and dopamine rhythms indicates that catecholamines do not drive these immunologic rhythms. Catecholamines could, however, influence the rhythm amplitude, because mean individual dPT in leukocytes and lymphocytes were reduced under LL. Similarly, plasma corticosterone rhythm was suppressed after LL exposure of Wistar rats for 4-12 wk (16, 33). Taken together, these results suggest that plasma glucocorticoids are not essential for the expression of these immunologic rhythms either, a finding that also applies to human subjects (21).
Thus circadian rhythms in immunologic variables remained synchronized at a group level in an environment (LL for 8-11 wk) that abolished circadian rhythms in body temperature, locomotor activity, and plasma catecholamines, as well as, reportedly, plasma corticosterone. Further exposure to LL likely produced desynchronization of persistent immunologic rhythms among individual rats, i.e., their periods differed. However, due to the limited number of time points per animal, firm conclusions from individual immunologic time series cannot be drawn from these experiments.
Continuous LL exposure can be considered as a functional pinealectomy, because light exposure reduces plasma melatonin levels (22). This condition, however, does not suppress its circadian rhythmicity (8). Surgical pinealectomy, like pharmacological pinealectomy with evening propanolol, decreased immune responses to sheep red blood cells or allogeneic lymphocytes in mice (17, 22, 23, 29). Surgical pinealectomy also reduced the night peak of the proliferation of bone marrow granulomonocytic colony-forming units, interleukin-2 production by spleen cells, and NK cell activity. Such hematoimmunologic rhythm disturbances were corrected with melatonin supplementation (17).
Rats that were previously exposed to DD for 16 wk recovered normal circadian rhythms in leukocytes or total lymphocytes after being switched to LD12:12 for 4 or 16 wk. Conversely, neither variable displayed any group 24-h rhythm in those animals previously exposed to LL, although individual changes in Th and Ts lymphocytes maintained their usual phase relationship. Such LL-induced immunologic rhythms impairment persisted up to 16 wk after rats were switched to LD12:12, whereas body temperature and activity rhythms had returned to normal. This suggests a long-lasting effect of LL on the immune system. Circulating lymphocyte subsets are currently determined in human subjects as a means to assess immune status. In particular, the extent of alteration of the circadian amplitude of circulating Th subset varied as a function of disease stage in human immunodeficiency virus-infected patients, despite the fact that cortisol and other hormonal rhythms were maintained (4, 35). Indeed, circadian rhythms in circulating T subsets likely result from a rhythmic release of immature T cells from lymphoid organs, which in turn results from circadian waves of lymphoid proliferation and differentiation steps (6). These latter ones are also stimulated (regulated) by antigen encounter (reviewed in Refs. 19 and 20). Therefore we believe that, although the present study restricted its scope to rhythms in circulating lymphocyte subsets, the results support the hypothesis of an autonomous immunologic oscillator, which can be severely impaired with LL exposure beyond 3 mo.
Perspectives
The persistence of circadian rhythms in immunologic parameters despite suppression of three main outputs from the circadian clock supports the hypothesis of the existence of an independent oscillator involved in the circadian regulation of immunologic functions. This "immunologic" oscillator was revealed after 2 mo of LL exposure in rats.It can be speculated that this oscillator is synchronized or reset with different environmental cues than those which calibrate the hypothalamic SCN, which is the main central biological clock. Cyclic antigen exposure as provided by environment or food may indeed play such a role. The lack of external cycles in antigen exposure or their "wrong" timing, with regard to efficiency of the circadian coordination system involving the SCN and the putative immunologic oscillator, might indeed contribute to internal desynchronization and disruption of circadian coordination and facilitate several disease processes, such as infection, allergy, autoimmunity, or cancer. How the putative immunologic oscillator and the SCN synchronize each other and which mediators are involved will be a fascinating area to explore.
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ACKNOWLEDGEMENTS |
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We thank M. Comisso, G. Debotte, E. Filipski, S. van den Heiligenberg, D. Melotée, and M. Tampellini for helpful technical assistance. We further thank C. Mormont for insightful criticisms and comments.
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FOOTNOTES |
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This work was supported by grant Contrat de Recherche Externe de l' Institut National de la Santé et de la Recherche Médicale 92-0913, by grant Réseau de Recherche de l' Institut National de la Santé et de la Recherche Médicale 4R007A, by the Association pour la Recherche sur le Temps Biologique et la Chronothérapie, and by the Association Nouvelles Recherches Biomédicales, Hôpital Paul Brousse, Villejuif, France.
Parts of this work were presented at the 85th Annual Meeting of American Association for Cancer Research, San Francisco, CA, April 10-13, 1994, and at the World Conference on Chronobiology, September 6-10, 1995, Ferrara, Italy.
1 Supplementary figures, available upon request from F. Lévi, show changes of circulating Th and Ts lymphocyte counts along the 24-h time scale in the same rats kept in LL for 11 and 16 wk and then switched to LD12:12 for 16 wk. The figures illustrate the internal synchronization of Th and Ts rhythms in all three conditions within each individual rat. The individual dPT is reduced in LL as opposed to LD12:12 for both lymphocyte subset rhythms.
Address for reprint requests: F. Lévi, Laboratoire Rythmes Biologiques et Chronothérapeutique, ICIG et Université Paris XI, Hôpital Paul Brousse, 14 Ave. Paul Vaillant Couturier, 94807 Villejuif Cedex, France.
Received 25 October 1996; accepted in final form 27 August 1997.
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Abstract
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Discussion
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), or
under LL (
) or DD (
) (experiment
1). Under LD12:12, epinephrine and
norepinephrine displayed circadian rhythms with respective amplitudes
of 61 and 58% of the individual mean and maxima located near 0600;
dopamine plasma concentrations were higher during light than during
darkness (5.4 ± 1 vs. 2.9 ± 0.5 nmol/l, respectively;
P = 0.02). In LL, all rhythms were
suppressed. In DD, epinephrine and norepinephrine circadian rhythms
were restored with respective amplitudes of 68 and 42% of individual
mean and acrophases located at ~0630; dopamine concentrations were
higher from 1600 to 2400 than from 0400 to 1200 (5.3 ± 0.5 vs. 3.8 ± 0.4 nmol/l, respectively; P = 0.03).
) or DD (
