Circadian Activity Rhythms and Phase‐Shifting of Cultured Neurons of the Rat Suprachiasmatic Nucleus

The mammalian suprachiasmatic nucleus (SCN) is the major endogenous pacemaker that coordinates various daily rhythms including locomotor activity and autonomous and endocrine responses, through a neuronal and humoral influence. In the present study we examined the behavior of dispersed individual SCN neurons obtained from 1‐ to 3‐day‐old rats cultured on multi‐microelectrode arrays (MEAs). SCN neurons were identified by immunolabeling for the neuropeptides arginine‐vasopressin (AVP) and vasoactive intestinal polypeptide (VIP). Single SCN neurons cultured at low density onto an MEA can express firing rate patterns with different circadian phases. In these cultures we observed rarely synchronized firing patterns on adjacent electrodes. This suggests that, in cultures of low cell densities, SCN neurons function as independent pacemakers. To investigate whether individual pacemakers can be influenced independently by phase‐shifting stimuli, we applied melatonin (10 pM to 100 nM) for 30 min at different circadian phases and continuously monitored the firing rate rhythms. Melatonin could elicit phase‐shifting responses in individual clock cells which had no measurable input from other neurons. In several neurons, phase‐shifts occurred with a long delay in the second or third cycle after melatonin treatment, but not in the first cycle. Phase‐shifts of isolated SCN neurons were also observed at times when the SCN showed no sensitivity to these phase‐shifting stimuli in recordings from brain slices. This finding suggests that the neuronal network plays an essential role in the control of phase‐shifts.     

Effect of NO Synthase Inhibition on Cardiovascular Circadian Rhythms in Wild‐Type and eNOS‐Knock‐Out Mice

Cardiovascular functions (blood pressure [BP], heart rate [HR]) were radiotelemetrically studied in endothelial nitric oxide synthase (NOS) knock‐out mice (eNOS‐/‐) and their wild type C57BL/6 (WT) controls. Studies were performed with and without inhibition of the NOS with the non‐specific inhibitor N^ω‐Nitro‐L‐Arginin‐Methylester (L‐NAME). Six eNOS‐/‐and five WT mice, kept under a light:dark schedule of 12:12 h (lights on 07:00 h), were treated with L‐NAME in tap water containing different concentrations (94, 282, and 940 mg/kg) each for three days. Under control conditions, the eNOS‐/‐mice are mildly hypertensive in comparison to WT. L‐NAME increased systolic [SBP] and diastolic [DBP] blood pressures in WT mice to the levels of eNOS‐/‐mice after two days of L‐NAME application with no dose‐dependency, whereas L‐NAME had no effects on SBP and DBP in eNOS‐/‐mice. In neither mouse strain were the circadian rhythms in BP and HR affected by drug treatment. The similarity of the 24 h BP profiles in eNOS‐/‐and L‐NAME‐treated WT mice support the notion that only the enothelial NOS and not other NOS isoenzymes are of importance for hypertension in the knock‐out mouse strain.     

Effects of Three‐Hour Restricted Food Access during the Light Period on Circadian Rhythms of Temperature,Locomotor Activity,and Heart Rate in Rats

The effects of food on biological rhythms may influence the findings of chronopharmacological studies. The present study evaluated the influence of a restricted food access during the rest (light) span of nocturnally active Wistar rats on the 24 h time organization of biological functions in terms of the circadian rhythms of temperature (T), heart rate (HR), and locomotor activity (LA) in preparation for subsequent studies aimed at evaluating the influence of timed food access on the pharmacokinetics and pharmacodynamics of medications. Ten‐wk‐old male Wistar rats were housed under controlled 12:12 h light:dark (LD) environmental conditions. Food and water were available ad libitum, excepted during a 3 wk period of restriction. Radiotelemetry transmitters were implanted to record daily rhythms in T, HR, and LA. The study lasted 7 wk and began after a 21‐d recovery span following surgery. Control baseline data were collected during the first wk (W1). The second span of 3 wk duration (W2 to W4) consisted of the restricted feeding regimen (only 3 h access to food between 11:00 and 14:00 h daily) during the L (rest span) under 12:12 h LD conditions. The third period of 3 wk duration (W5 to W7) consisted of the recovery span with ad libitium normal feeding. Weight loss in the amount of 5% of baseline was observed during W1 with stabilization of body weight thereafter during the remaining 2 wk of food restriction. The 3 h restricted food access during the L rest span induced a partial loss of circadian rhythmicity and the emergence of 12 h rhythms in T, HR, and LA. Return to ad libitum feeding conditions restored circadian rhythmicity in the manner evidenced during the baseline control span. Moreover, the MESORS and amplitudes of the T, HR, and LA 24 h patterns were significantly attenuated during food restriction (p<0.001) and then returned to initial values during recovery. These changes may be interpreted as a masking effect, since T, HR, and LA are known to directly react to food intake. The consequences of such findings on the methods used to conduct chronokinetic studies, such as the fasting of animals the day before testing, are important since they may alter the temporal structure of the organism receiving the drug and thereby compromise findings.     

The Effects of Circadian Rhythmicity and Time‐Awake on a Simple Motor Task

The aim of the study was to assess if a simple motor task, one that required muscle contractions well below maximum, showed evidence of circadian changes and time‐awake. The task consisted of using a larger counter to flick a number of smaller counters to land as near as possible to the center of a target. The closer a counter landed next to the center of the target, the higher the score obtained. Two distances from the target were used (long and short), and 20 counters were flicked at each distance. The task was performed by 72 diurnally active healthy participants at six test sessions distributed every 4 h throughout the day (08:00 h, 12:00 h, … , 04:00 h), so covering a circadian cycle. When performing the sessions, subjects had been awake for about 1, 4, … , 20 h. Before each test session, sublingual temperature was measured, and estimates of the individual's fatigue and alertness were made. Clear normally phased circadian rhythms (p<0.0001) in oral temperature and alertness with mean peak time (i.e., acrophases of 17.2 h and 15.9 h, respectively) and fatigue (i.e., mean acrophase of 3.4 h) were detected. The total scores for both the long and short distances also showed circadian rhythms that peaked slightly before the temperature rhythm (by 2.31±0.91 h and 1.77±0.77 h, mean±SE, respectively), and the number of occasions that the target was missed altogether showed rhythms that were in anti‐phase (mean acrophases=3.8 h and 4.1 h for the long and short distances, respectively) to that of total scores (mean acrophases=16.0 h and 15.2 h for the long and short distances, respectively). With the long and, particularly, short distances, there were generally significant correlations (r<0.0005) between both the measures of accuracy (total score and number of misses) and body temperature and time‐awake. The accuracy of performance at this task seems to show circadian and time‐awake effects, and so makes it of potential value in protocols where repetitive measurements during the course of a day are required.     

Effects of a Two‐Hour Change in Bedtime on the Sleep of Healthy Seniors

Some of the sleep disruption seen in seniors (>65 yrs) may be due to alteration of the circadian pacemaker phase and/or its phase angle with bedtime. The purpose of this study was to determine the effects of 2 h changes in the timing of bedtime (both earlier and later) on the sleep of seniors. Ten healthy seniors (9 F, 1 M, age 70–82 yrs) were each studied individually during three 120 h sessions (each separated by >2 weeks) in a time‐isolation laboratory. On nights 1 and 2, bedtime and rise‐time occurred at the subjects' habitual times; on nights 3–5, bedtime was specified by the experiment, but rise‐time was at the subjects' discretion (without knowledge of clock time). Under the control condition, subjects went to bed at their habitual bedtime (HBT), under the earlier bedtime condition at (HBT?2 h), and under the later bedtime condition at (HBT+2 h). Sleep was polysomnnographically recorded and rectal temperature continuously monitored. Although total sleep time increased in the earlier compared to the later condition (p<0.01), sleep efficiency decreased and wake after sleep onset increased (p<0.01). Subjective ratings of sleep were also worse under the earlier (HBT?2 h) than under later (HBT+ 2 h) condition (p<0.05). Performance did not differ between the earlier and later conditions. The larger the phase angle between actual bedtime and circadian temperature minimum (Tmin), the longer the time spent in bed and total sleep time, and the worse the sleep efficiency and subjective sleep ratings. There were no effects related to the phase angle between Tmin and rise‐time. The relative benefits of longer vs. more efficient sleep in the elderly require further investigation.     

Seasonal Modulation of the 8‐and 24‐Hour Rhythms of Ondansetron Tolerance in Mice

Ondansetron (Zophren®) is a serotonin 5HT₃-receptor antagonist used primarily to control nausea and vomiting caused by cytotoxic chemo‐and radio‐therapy. Tolerance to this drug shows both 24 and 8 h periodicities. In this framework, this study aimed to determine whether these ondansetron tolerance rhythms are modulated by season. The chronotoxic effect of a fixed dose (3.5 mg/kg, i.p.) of the drug was investigated with reference to both time of the day and year dependencies. Season‐related studies were performed on 560 male Swiss mice, 10 to 12 wks old, synchronized with L:D=12:12 for three weeks. During a 1 yr span (2005), four 24 h studies were performed with a single dosing time at 1, 7, 13, and 19 hours after light onset (HALO), respectively. Tolerance was assessed daily during a 40‐day span after acute ondansetron treatment. Both χ² test and cosinor methods were used to analyze the time series data. Statistically significant dosing time‐dependent changes were validated in both yearly and daily time scales. The 24 h mean survival rate peaked in spring (92%) compared to fall (72%), the 20% difference being statistically significant (χ² test with p<0.05 and cosinor with p<0.0001 for seasonal rhythm detection and with a peak time, Ø,=April 3±6.6 days). A 24 h rhythm was also detected in each of the seasonal time points. However, the curve pattern was monophasic in fall as well as spring. In fall, a large amplitude (A) circadian rhythm was detected that peaked at 19 HALO, while in the spring, a small circadian rhythm was detected that peaked at 1 HALO. The curve pattern was biphasic in summer (with large A) and in winter (with a small A). The existence of two peaks of equal magnitude in winter (100% survival rate) and in summer (100% and 90%) suggests the presence of both circadian and ultradian rhythms rather than an ultradian component of the 24 h period. The seasonal modulation of ondansetron circadian chronotolerance seems to involve several rhythm parameters: season‐related changes in the 24 h mean (M), amplitude (A), acrophase location (Ø), as well as bimodal curve patterns including the coexistence of rhythms with respectively 24 and 8 h periods in winter and summer. In conclusion, tolerance to ondansetron varies not only according to the 24 and 8 h periods but also according to seasons, which suggests the complexity of ondansetron toxicity rhythms. Seasonal modulation of ondansetron tolerance may also influence the strategies of chemo‐and chrono‐therapy, and it is therefore necessary to take it into account in clinical drug‐delivery protocols to minimize side effects of cytotoxic anticancer and antiemetic agents.     

Effects of Galarmin and Cobra Venom on the Morphofunctional State of the Substantia Nigra in a Rat Model of Parkinson’s Disease

Neurophysiology - Morphofunctional characteristics of the substantia nigra (SN) were compared in four groups of experimental rats: (i) control (sham-operated), (ii) animals with a rotenone model of...     

Daily Rhythms of P‐glycoprotein Expression in Mice

Recent studies have shown the gene expression of several transporters to be circadian rhythmic. However, it remains to be elucidated whether the expression of P‐glycoprotein, which is involved in the transport of many medications, undergoes 24 h rhythmicity. To address this issue, we investigated daily profiles of P‐glycoprotein mRNA and protein levels in peripheral mouse tissues. In the liver and intestine, but not in the kidney, Abcb1a mRNA expression showed clear 24 h rhythmicity. On the other hand, Abcb1b and Abcb4, the other P‐glycoprotein genes, did not exhibit significant rhythmic expression in the studied tissues. In the intestine, levels of whole P‐glycoprotein also exhibited a daily rhythm, with a peak occurring in the latter half of the light phase and a trough at the onset of the light phase. Consistent with the day‐night change of P‐glycoprotein level, the ex vivo accumulation of digoxin, an Abcb1a P‐glycoprotein substrate, into the intestinal segments at the onset of dark phase was significantly lower than it was at the onset of the light phase. Thus, Abcb1a P‐glycoprotein expression, and apparently its function, are 24 h rhythmic at least in mouse intestine tissue. This circadian variation might be involved in various chronopharmacological phenomena.     

Time‐of‐Day Effects on Myoelectric and Mechanical Properties of Muscle During Maximal and Prolonged Isokinetic Exercise

The aim of this study was to examine the time‐of‐day (TOD) effects in myoelectric and mechanical properties of muscle during a maximal and prolonged isokinetic exercise. Twelve male subjects were asked to perform 50 maximal voluntary contractions (MVC) of the knee extensor muscles at a constant angular velocity of 2.09 rad · sec^?1, at 06∶00 and 18∶00 h. Torque and electromyographic (EMG) parameters were recorded for each contraction, and the ratio between these values was calculated to evaluate variations of the neuromuscular efficiency (NME) with fatigue and with TOD. The results indicated that maximal torque values (T₄₅Max) was significantly higher (7.73%) in the evening than in the morning (p<0.003). The diurnal variation in torque decrease was used to define two phases. During the first phase (1st to the 26th repetition), torque values decreased fast and values were higher in the evening than in the morning, and during the second phase (27th to the 50th repetition), torque decreased slightly and reached a floor value that appeared constant with TOD. The EMG parameters (Root Mean Square; RMS) were modified with fatigue, but were not TOD dependent. The NME decrease–significantly with fatigue, showing that peripheral factors were mainly involved in the torque decrease. Furthermore, NME decrease was greater at 18∶00 than at 06∶00 h for the vastus medialis (p<0.05) and the vastus lateralis muscles (p<0.002), and this occurred during the first fatigue phase of the exercise. In conclusion, the diurnal variation of the muscle fatigue observed during a maximal and prolonged isokinetic exercise seems to reflect on the muscle, with a greater contractile capacity but a higher fatigability in the evening compared to the morning.     

Circadian Time‐Effect of Orally Administered Loratadine on Plasma Pharmacokinetics in Mice

Little is known about the chronopharmacokinetics of loratadine, a long‐acting tricyclic antihistamine H₁ widely used in the treatment of allergic diseases. Hence, the pharmacokinetics of loratadine and its major metabolite, desloratadine, were investigated after a 20 mg/kg dose of loratadine had been orally administered to comparable groups of mice (n=33), synchronized for three weeks to 12 h light (rest span)/12 h dark (activity span). The drug was administered at three different circadian times (1, 9, and 17 h after light onset [HALO]). Multiple blood samples were collected over 48 h, and plasma concentrations of loratadine and desloratadine were determined by high performance liquid chromatography. There were no significant differences in T_max of loratadine and desloratadine between treatment‐time different groups. However, the elimination half‐life (t1/2) of the parent compound and its metabolite was significantly longer (p<0.01) following administration at 9 HALO (t1/2 loratadine and desloratadine 5.62 and 4.08 h at 9 HALO vs. 4.29 and 2.6 h at 17 HALO vs. 3.26 and 3.27 at 1 HALO). There were relevant (p<0.05) differences in C_max between the three treated groups for loratadine and desloratadine; 133.05±3.55 and 258.07±14.45 ng/mL at 9 HALO vs. 104.5±2.61 and 188.62±7.20 ng/mL at 1 HALO vs. 94.33±20 and 187.75±10.79 ng/mL at 17 HALO. Drug dosing at 17 HALO resulted in highest loratadine and desloratadine total apparent clearance values: 61.46 and 15.97 L/h/kg, respectively, whereas loratadine and desloratadine clearances (CL) were significantly slower (p<0.05) at the other administration times (loratadine and desloratadine CL was 57.3 and 14.22 L/h/kg at 1 HALO vs. 43.79 and 12.89 L/h/kg at 9 HALO, respectively). The area under the concentration‐time curve (AUC) of loratadine and desloratadine was significantly (p<0.05) greater following drug administration at 9 HALO (456.75 and 1550.57 (ng/mL) · h, respectively); it was lowest following treatment at 17 HALO (325.39 and 1252.53 (ng/mL) · h, respectively). These pharmacokinetic data indicate that the administration time of loratadine significantly affected its pharmacokinetics: the elimination of loratadine and its major metabolite desloratadine.     

Alterations of the Characteristics of the Circadian Rest‐Activity Rhythm of Cancer In‐Patients

The aim of the present study was to evaluate the characteristics of the circadian rest‐activity rhythm of cancer patients. Thirty‐one in‐patients, consisting of 19 males and 12 females, were randomly selected from the Regional Cancer Center, Pandit Jawaharlal Nehru Medical College, Raipur, India. The rest‐activity rhythm was studied non‐invasively by wrist actigraphy, and compared with 35 age‐matched apparently healthy subjects (22 males and 13 females). All subjects wore an Actiwatch (AW64, Mini Mitter Co. Inc., USA) for at least 4–7 consecutive days. Fifteen‐second epoch length was selected for gathering actigraphy data. In addition, several sleep parameters, such as time in bed, assumed sleep, actual sleep time, actual wake time, sleep efficiency, sleep latency, sleep bouts, wake bouts, and fragmentation index, were also recorded. Data were analyzed using several statistical techniques, such as cosinor rhythmometry, spectral analysis, ANOVA, Duncan's multiple‐range test, and t‐test. Dichotomy index (I<O) and autocorrelation coefficient (r24) were also computed. The results validated a statistically significant circadian rhythm in rest‐activity with a prominent period of 24 h for most cancer patients and control subjects. Results of this study further revealed that cancer patients do experience a drastic alteration in the circadian rest‐activity rhythm parameters. Both the dichotomy index and r24 declined in the group of cancer patients. The occurrence of the peak (acrophase, Ø) of the rest‐activity rhythm was earlier (p<0.001) in cancer patients than age‐ and gender‐matched control subjects. Results of sleep parameters revealed that cancer patients spent longer time in bed, had longer assumed and actual sleep durations, and a greater number of sleep and wake bouts compared to control subjects. Further, nap frequency, total nap duration, average nap, and total nap duration per 1 h awake span were statistically significantly higher in cancer patients than control subjects. In conclusion, the results of the present study document the disruption of the circadian rhythm in rest‐activity of cancer in‐patients, with a dampening of amplitude, lowering of mean level of activity, and phase advancement. These alterations of the circadian rhythm characteristics could be attributed to disease, irrespective of variability due to gender, sites of cancer, and timings of therapies. These results might help in designing patient‐specific chronotherapeutic protocols.     

Effects of 2‐Chloro‐2′‐Deoxyadenosine on the Cell Cycle in the Human Leukemia EHEB Cell Line

To explain why 2‐chloro‐2′‐deoxyadenosine (CdA) is unable to block DNA synthesis and cell cycle progression, and paradoxically enhances progression from G1 into S phase in the CdA‐resistant leukemia EHEB cell line, we studied its metabolism and effects on proteins regulating the transition from G1 to S phase. A low deoxycytidine kinase activity and CdATP accumulation, and a lack of p21 induction despite p53 phosphorylation and accumulation may account for the inability of CdA to block the cell cycle. An alternative pathway involving pRb phosphorylation seems implicated in the CdA‐induced increase in G1 to S phase progression.     

Effects of α-Tocopherol on Carbon Tetrachloride Metabolism in Rat Liver Microsomes

α-tocopherol is the major lipid-soluble radical-scavenging antioxidant in rat liver. It has long been used as a putative protective agent in CC1₄ induced liver injury but with variable results. We have used a-tocopherol loaded rat liver microsomes to study the effect of this vitamin on CC1₄ metabolism in vitro. As expected, a-tocopherol inhibits CC1₄-dependent microsomal lipid peroxidation and, at a very high concentration, will inhibit the covalent binding of CC1,- to microsomal protein by up to 50%. No inhibitory effect was observed towards CC1₃ production as measured by the electron spin resonance technique of spin-trapping but this apparent discrepancy may represent a limitation of the technique. The high levels required to inhibit covalent binding probably preclude the likelihood of a-tocopherol significantly affecting that phenomenon at endogenous concentrations but may be relevant to other experiments employing high doses of a-tocopherol as an experimental hepatoprotective agent.     

Biological Rhythms of Biochemical Serum Parameters in a Brazilian Population: a Three‐Year Study

A marked decrease in analytical and post‐analytical variability has been achieved in clinical laboratories by the use of automated analytical systems. Current studies are now focused on the origin of pre‐analytical variability, such as that due to intra‐individual differences and biological rhythms. The objective of this work was to evaluate the occurrence of biological rhythms in several biochemical serum parameters in a Brazilian population. A retrospective study (1996 to 1998) was carried out to collect the test results within the reference intervals of adults, from 21 to 50 yr of age (average age of 36 yr) attending the outpatient clinics of the Teaching Hospital at the University of Campinas, São Paulo, Brazil. The reference sample was 52.9% male and 47.1% female and encompassed 15,036 calcium, 7,478 phosphorus, 53,641 urea, 58,315 creatinine and 6,433 uric acid determinations (140,903 in total). Significant annual rhythms were detected in serum calcium (p≤0.001), with maximum and minimum values in fall and spring, and in serum creatinine (p≤0.002), with maximum and minimum values in summer and winter. The other parameters did not present significant annual rhythmicity. The seasonal rhythms present in the serum concentrations of calcium and creatinine observed in this large population study, although of small amplitude, should be considered a component of the pre‐analytical variation of these clinical laboratory tests.     

Effects of Transient and Continuous Wheel Running Activity on the Upper and Lower Limits of Entrainment to Light‐Dark Cycles in Female Hamsters

The entrainment limits to light‐dark cycles can be modified by the experimental conditions under which they are tested. Among the factors that may influence entrainment is the amount of wheel running exerted by the animal. In the present work, the effects of transitory and continuous wheel running on entrainment to light‐dark cycles were tested using a range of T cycles at the entrainment limits. Four groups of female hamsters were submitted to 1 h stepwise changes in T cycles. Two groups were exposed to T cycles of which the period was shortened at the lower limit from T22 to T18, and the other two groups were exposed to cycles that lengthened at the upper limit from T27 to T32. One of the groups at the lower limit and one at the upper limit had continuous access to a running wheel, while the others had the wheel locked, except at certain T when a lack of period control by T cycle appeared. The study demonstrates that access to running wheel widens the limits of entrainment to LD cycles. Specifically, the following observations were made: the effects of wheel running for entrainment were more evident in the groups with continuous access to wheel, as they did entrain to T19 and T32; continuous access to a wheel produced aftereffects only after T19, but not under T32; and when animals without a wheel showed relative coordination, unlocking the wheel favored entrainment in all the animals at T31, but in only 1 out 6 at T19. All of these indicate a different effect of the wheel running on the upper and lower limits of entrainment.     

Age‐Related Effects on the Biological Clock and its Behavioral Output in a Primate

In humans, activity rhythms become fragmented and attenuated in the elderly. This suggests an alteration of the circadian system per se that could in turn affect the expression of biological rhythms. In primates, very few studies have analyzed the effect of aging on the circadian system. The mouse lemur provides a unique model of aging in non‐human primates. To assess the effect of aging on the circadian system of this primate, we recorded the circadian and daily rhythms of locomotor activity of mouse lemurs of various ages. We also examined age‐related changes in the daily rhythm of immunoreactivities for vasoactive intestinal polypeptide (VIP) and arginine‐vasopressin (AVP) in suprachiasmatic nucleus neurons (SCN), two major peptides of the biological clock. Compared to adult animals, aged mouse lemurs showed a significant increase in daytime activity and an advanced activity onset. Moreover, when maintained in constant dim red light, aged animals exhibited a shortening of the free‐running period compared to adult animals. In adults, AVP immunoreactivity (ir) peaked during the second part of the day, and VIP ir peaked during the night. In aged mouse lemurs, the peaks of AVP ir and VIP ir were significantly shifted with no change in amplitude. AVP ir was most intense at the beginning of the night; whereas, VIP ir peaked at the beginning of the daytime. A weakened oscillator could account for the rhythmic disorders often observed in the elderly. Changes in the daily rhythms of AVP ir and VIP ir may affect the ability of the SCN to transmit rhythmic information to other neural target sites, and thereby modify the expression of some biological rhythms.     

Exposure to T‐Cycles of 22 and 23 h during Lactation Modifies the Later Dissociation of Motor Activity and Temperature Circadian Rhythms in Rats

Early environmental conditions may affect the development and manifestation of circadian rhythms. This study sought to determine whether the maintenance of rats under different T‐cycles during lactation influences the subsequent degree of dissociation of the circadian rhythms of motor activity and core body temperature. Two groups of 22 day‐old Wistar rats were kept after weaning under T‐cycles of 22 h (T22) or 23 h (T23) for 70 days. Subsequently, they were kept in constant darkness (DD). Half of the animals in each group were born and reared under these experimental conditions, while the other half were reared until weaning under 24 h LD cycles (T24). Rats transferred from T24 to T22 or T23 showed two circadian components in motor activity and temperature, one entrained by light and the other free‐running. In T22, there was also desynchronization between temperature and motor activity. Rats submitted to T23 from birth showed higher stability of the 23 h component than rats transferred from T24 to T23 after weaning. However, in comparison to rats born under T24 and subsequently changed to T22, animals submitted to T22 from birth showed shorter values of the period of the non‐light‐dependent component during T22, more aftereffects when transferred to DD, and a lack of desynchronization between motor activity and temperature. The results suggest that T‐cycles in the early environment may modify overt rhythms by altering the internal coupling of the circadian pacemaker.     

Age‐related Changes in the Circadian and Homeostatic Regulation of Human Sleep

The reduction of electroencephalographic (EEG) slow‐wave activity (SWA) (EEG power density between 0.75–4.5 Hz) and spindle frequency activity, together with an increase in involuntary awakenings during sleep, represent the hallmarks of human sleep alterations with age. It has been assumed that this decrease in non‐rapid eye movement (NREM) sleep consolidation reflects an age‐related attenuation of the sleep homeostatic drive. To test this hypothesis, we measured sleep EEG characteristics (i.e., SWA, sleep spindles) in healthy older volunteers in response to high (sleep deprivation protocol) and low sleep pressure (nap protocol) conditions. Despite the fact that the older volunteers had impaired sleep consolidation and reduced SWA levels, their relative SWA response to both high and low sleep pressure conditions was similar to that of younger persons. Only in frontal brain regions did we find an age‐related diminished SWA response to high sleep pressure. On the other hand, we have clear evidence that the circadian regulation of sleep during the 40 h nap protocol was changed such that the circadian arousal signal in the evening was weaker in the older study participants. More sleep occurred during the wake maintenance zone, and subjective sleepiness ratings in the late afternoon and evening were higher than in younger participants. In addition, we found a diminished melatonin secretion and a reduced circadian modulation of REM sleep and spindle frequency—the latter was phase‐advanced relative to the circadian melatonin profile. Therefore, we favor the hypothesis that age‐related changes in sleep are due to weaker circadian regulation of sleep and wakefulness. Our data suggest that manipulations of the circadian timing system, rather than the sleep homeostat, may offer a potential strategy to alleviate age‐related decrements in sleep and daytime alertness levels.