Circadian rhythmus of the excretion of electrolytes and urinary proteins in rats]

Circadian rhythms in urinary water, sodium, potassium and proteins excretion are studied in 45 rats living alone in metabolism cages. Urines are collected during 4 consecutive 6 hours long periods during 2 consecutive days. Large circadian variations of these parameters (especially water and proteins excretion and urinary protein concentration) are described. The influence of feeding rhythms on the circadian urinary excretion rhythms is discussed. It is proposed that nightly renal hemodynamic changes (during meal digestion or with high renin plasma levels) can induce modifications in glomerular filtration rate and electrolytes and macromolecules transglomerular flow.     

Periodic fluctuations of the blood pressure and heart contraction rate in premature newborn infants

To study the postnatal development of circadian rhythm of the blood pressure and the heart rate these parameters were monitored automatically during 24-72 hours at 5h intervals. Fifty infants were investigated at the age 1, 2, 3, and 4 weeks. The results were compared with the cosine curves of different periods (1-48 h) by the IBM PC XT. The fluctuations with all mentioned periods including circadian could be determined in each infant, dominating period being of any duration. In traditional approach the expressiveness of periodical fluctuations is evaluated by the amplitude of cosine curve. Application of the criterion has shown that only amplitude values exceeding 8 mm Hg should be taken into account. Hence, the analysis of BP and HR time series has demonstrated that about 45-50% of the infants have noncircadian rhythms, 20-20% have no significant periodical fluctuations and only 25-30% have circadian rhythm.     

Ultradian and circadian compartmentalization of respiratory and metabolic exchanges in small laboratory vertebrates

Carbon dioxide emission (VCO2) taken as an index of respiratory and metabolic exchanges, was continuously recorded during 4-30 consecutive days in 100 quail, 87 chicks, 347 rats, 665 mice and 70 guinea-pigs which were under controlled environmental parameters. Harmonic analysis, fast Fourier transform, chi-square periodograms, peak and trough intervals were computed with VCO2 values obtained with CO2 concentrations sampled every 20 min on the CO2 recordings. In LD 12:12 alternation, circadian rhythms were observed in all quail, chicks, rats and mice, but only in 80% of the guinea-pigs. Ultradian VCO2 rhythms, with periods which show statistically significant interspecies differences, were assessed. For each of the 5 species these computed periods, which were the same in LL and DD, were: 1.17 h for quail and chickens, 1.25 h for rats, 1.50 h for mice and 1.0 h for guinea-pigs. In LD 12:12 these periods were different during L and D in quail, chicks, rats and mice, but not in guinea-pigs. The amplitudes of these ultradian variations were, according to the species, 10-20% of their mean VCO2 levels. These ultradian rhythms persist in the absence (or masking) of circadian rhythms, e.g. in LD 12:12 in 20% of guinea-pigs and in LL in 87% of Japanese quail and in 23% of Sprague-Dawley rats. Moreover, these ultradian rhythms persist during starvation, locomotor activity restraint and ageing. These ultradian VCO2 cycles which are related to rest-activity variations appear to be basic physiological rhythms with a genetic origin.     

Photoperiod variations of various blood biochemistry constants in the rabbit]

Biochemical blood parameters generally show fluctuations during the day. The aim of this study is to determine the variations of the following parameters: hemoglobin, glucose, triglycerides, ASAT, ALAT, urea and creatinine in New Zealand white (NZW) rabbits, kept under five different circadian rhythms (12/12, 14/10, 10/14, 16/8, 8/16, light/dark). These parameters were analyzed by using a reflection photometer (Reflotron). Most results are found within the physiological range for this species but there are significant differences among cycles. These findings could be explained by the dependence of the biochemical blood parameters on the light/dark cycle. It can be concluded that the most useful cycle to find fewer variations of the biochemical blood parameters in NZW rabbit is 12/12 light/dark cycle.     

Blood urea content changes in man under hypokinesia

It has been demonstrated that hypokinesia (diminished muscular activity) leads to an increase in blood urea content in man. Against this background the objective of this investigation was to determine blood urea content under hypokinesia (HK) on 17 physically healthy men aged 19-23 yr. They were divided into three groups: the 1st group (5 men) was examined under HK, the 2nd group (4 men) was studied during the background period (BGP) as well as in the readaptation period (RTP), and the 3rd group (8 men) was placed under ordinary conditions and served as control. For the simulation of the hypokinetic effect the men were kept under a rigorous bed rest regime for 16 days. Blood urea, blood creatinine, urine urea, and urine creatinine were measured. The results were processed statistically. The most pronounced increased urea content was observed in the men with an initial low concentration (3.3-4.2 mmole/liter). Variations in the urea concentration were analogous and manifested a reduction during the initial days and an elevation thereafter. Creatinine excretion and clearance were reduced uniformly and significantly during the initial 10 days of HK. It was concluded that diminished muscular activity induced an increase in urea content and a decrease in creatinine clearance in man.     

Comparative effect of theophylline on urinary protein excretion in rats during diurnal and nocturanal periods]

In 3 lots of 15 rats, urinary proteins excretion (U.P.E.) during 6 hours periods is compared : 1) daily and nightly in physiological conditions, where U.P.E. increases nightly (+ 97,47%) : 2) daily, before and after theophylline injection, where U.P.E. increases after theophylline injection (+ 78,62%) ; 3) nightly, before and after theophylline injection, where U.P.E. increase is only tiny (+ 3,53%) after theophylline injection. So, in nightly physiological and in daily experimental conditions the important U.P.E. increases can be in part explained by those of renal functions. This result can explain after nightly theophylline injection (where renal functions are already very higher than in daily period) the so small nightly proteinuria increase with the xanthic diuretic ; it cannot be compared to the daily value, because the renal functions cannot increase beyond some limits. Lastly, the present paper exhibits how a pharmacomolecule (as theophylline here) can act differently daily and nightly and open a new investigation field in chronopharmacology of diuretics.     

Ultradian, circadian and seasonal variations of plasma progesterone and LH concentrations during the luteal phase

The circadian variations in plasma progesterone (P) and LH concentrations were investigated in six women, aged 23-40 years. All were studied in the mid-luteal phase (7 +/- 2 days after LH mid-cycle surge). Experiments were conducted in autumn and in spring. Blood samples were obtained every 15 min for 24 hr. Plasma P and LH concentrations were measured by RIA. Each subject's time-series was analysed using three methods; visual inspection (chronogram), spectral analysis to estimate component periods of rhythms (tau) and cosinor analysis to quantify the rhythms parameters. Marked temporal variations in plasma P concentration were observed in each subject. The maximal variations over a 24-hr period, ranged between 13-58.5 mmol/l. Differences related to sampling time were statistically validated by ANOVA (p less than 0.00001). Significant harmonic periods were detected by spectral analysis but differed among subjects. In all subjects but one, a circadian rhythm was detected. The acrophase location was similar (about 0700 hr) in the four subjects studied in autumn, but ranged from 1940 to 0320 hr in those studied in spring. An ultradian rhythm with tau = 8 hr was also validated in six time-series with similar acrophases (about 0200, 1000, and 1800 hr). Cosinor analysis of pooled data revealed that the 24-hr, 12-hr, and 8-hr rhythms were statistically significant (p = 0.001) in autumn. algebraic sum of these three cosine functions yielded a circadian waveform with peak-times occurring near 0300 and 1130 hr and a trough-time about 2200 hr. In spring, the circadian pattern appeared quite different, and peak-times were found near 0700 and 2000 hr, and trough-times near 0300 and 1500 hr. Furthermore, the 24-hr mean of P was higher in autumn (28.9 +/- 0.4 nmol/l) than in spring (17.2 +/- 0.4 nmol/l), p from ANOVA less than 0.00001. The evidence for a similar circadian LH pattern is not as strong. Seasonal, circadian and ultradian rhythms characterize the physiologic time structure of plasma P concentration in mid-luteal phase.     

Effects of Eleutherococcus on the biorhythm of indices of peripheral blood in dogs

The circadian rhythms of the erythrocytes, hemoglobin and leucocytes concentration in the peripheral blood in dogs in winter and the effect of the eleutherococcus multi-injected at different periods of the day, were studied. The changes in the parameters of the studied biorhythms indices were observed only in the after dinner-injected animals, they being marked by the evident increase of the mesor and amplitude, acrophase shift by 1-2 hours by the beginning of the twenty-four hours.     

Circadian Rhythm in Gamma Glutamyltranspeptidase and Leucine Aminopeptidase Urinary Activity in Rats

Urinary gamma glutamyltranspeptidase (GGT) and leucine aminopeptidase (LAP), renal tubular brush border enzymes, have been shown to be sensitive indicators of renal tubular functions. This study documents circadian rhythms in the urinary activity of GGT and LAP, statistically validated and quantified by the cosinor method, in 15 male Wistar rats standardized to a LD 12:12 illumination schedule (light from 0800 hr to 2000 hr) and fed ad libitum. The acrophase of the circadian rhythms in urinary GGT and LAP activity occurred at the end of the rest span of the animals: between 1730 and 1915 for GGT (depending on the mode of expression of the activity) and between 1700 and 1910 for LAP. Of striking resemblance in their timing, both these rhythms were also of large amplitude (about 50% of the mesor for urinary GGT activity and about 45% for LAP one). The circadian acrophases of urinary GGT and LAP activity led in timing the circadian rhythms in urine volume and creatinine excretion by about 13hr. Such findings consistent with the circadian variations found by other investigators in GGT in kidney homogenates or in LAP in human urine thus reflect a periodicity in renal tubular function. The reasons for these circadian variations, still unknown at this time, are discussed. The influence recently demonstrated of the hormonal context on protein and enzyme synthesis at the tubule, and its phase relations to urinary enzyme excretion emphasize how much the circadian rhythm in urinary GGT and LAP activity is well included in the murine time structure. Therefore it should be of interest to consider the circadian rhythm in urinary GGT and LAP release as a marker rhythm of predictive value as to the side effects of nephrotoxic drugs.     

Circadian Rhythm in Gamma Glutamyltranspeptidase and Leucine Aminopeptidase Urinary Activity in Rats

Urinary gamma glutamyltranspeptidase (GGT) and leucine aminopeptidase (LAP), renal tubular brush border enzymes, have been shown to be sensitive indicators of renal tubular functions. This study documents circadian rhythms in the urinary activity of GGT and LAP, statistically validated and quantified by the cosinor method, in 15 male Wistar rats standardized to a LD 12:12 illumination schedule (light from 0800 hr to 2000 hr) and fed ad libitum. The acrophase of the circadian rhythms in urinary GGT and LAP activity occurred at the end of the rest span of the animals: between 1730 and 1915 for GGT (depending on the mode of expression of the activity) and between 1700 and 1910 for LAP. Of striking resemblance in their timing, both these rhythms were also of large amplitude (about 50% of the mesor for urinary GGT activity and about 45% for LAP one). The circadian acrophases of urinary GGT and LAP activity led in timing the circadian rhythms in urine volume and creatinine excretion by about 13hr. Such findings consistent with the circadian variations found by other investigators in GGT in kidney homogenates or in LAP in human urine thus reflect a periodicity in renal tubular function. The reasons for these circadian variations, still unknown at this time, are discussed. The influence recently demonstrated of the hormonal context on protein and enzyme synthesis at the tubule, and its phase relations to urinary enzyme excretion emphasize how much the circadian rhythm in urinary GGT and LAP activity is well included in the murine time structure. Therefore it should be of interest to consider the circadian rhythm in urinary GGT and LAP release as a marker rhythm of predictive value as to the side effects of nephrotoxic drugs.     

Daily Rhythms in Blood and Milk Lead Toxicokinetics Following Intravenous Administration of Lead Acetate to Dairy Cows in Winter

The objective was to investigate circadian variations of blood and milk lead toxicokinetics in dairy cows in winter. Twenty lactating Holstein animals were randomly assigned to 4 treatments, corresponding to different hours after onset of light (HALO): 2, 8, 14 and 20. Cows received a single intravenous administration of 2.5 mg/kg lead as lead acetate. Blood and milk samples were taken and analyzed by atomic absorption spectrophotometry. For each toxicokinetic parameter, a one-way analysis of variance was performed to outline the existence of daily variations. Significant differences as a function of HALO were detected in blood for the hybrid constant of elimination (β), half-life of elimination (t_1/2β), area under the curve (AUC) and clearance (Cl) (p &lt; 0.01) and volume of distribution at steady state (V_ss) (p < 0.05). Half-life of elimination was highest when lead acetatae was injected at 2 HALO, and lowest following the 14 HALO administration. Milk data showed significant differences for maximum concentration, AUC and Cl (p &lt; 0.001), volume of distribution and V_ss (p < 0.005). The ratio AUC_milk/ABC_blood was utilized to estimate penetration of lead in milk. It differed significantly along the day (p < 0.01). Blood daily variations could be fitted a circadian rhythms. No circadian rhythms were detected in milk parameters or in the ratio AUC_milk/ABC_blood. It was concluded that blood and milk lead toxicokinetics are distinctly affected by the time of the day.     

Reaction of circadian rhythms of the lymphoid system to deep screening from geomagnetic fields of the earth

C57B1/6 inbred mice were placed in hypomagnetic condition during 14 days constantly. Degree of relaxation of geomagnetic field was 10(4). The increase of the number of eosinophil granulocytes was discovered in peripheral blood of mice. Measures of circadian rhythms of blood's absolute lymphocytosis, absolute number of cells in bone marrow, thymus, spleen and inguinal lymph nodes were safe. Adaptation of lymphoid system to hypomagnetic condition was manifested by desynchronization of circadian rhythmicity on the basis of different sensitivity of lymphoid organs, that realized in strengthening of ultradian rhythms with periods of 15 hours. There are indirect data, that show the increase of speed and/or volume of recirculation of lymphoid cells.     

The internal synchronization of five circadian functions of the rabbit

Five different physiological functions of the rabbit (hard faeces and urine excretion, food and water intake and locomotor activity) were registered during LD 12:12 and during continuous light conditions (LL).

(1) In LD 12:12 a strong synchronization of the five parameters existed. The minima of all functions consistently occurred during the hours of light. The nocturnal percentage of overall 24-hr events was increased significantly in 'hard faeces excretion' (66±8 (S.D.) %), 'water intake' (64±15 (S.D.) %) and 'urine excretion' (58±10 (S.D.) %). The nocturnal percentage of locomotor activity was significantly increased during the dark-hours in 9 out of 14 animals. In the other five individuals prominent peaks were present even during the photoperiod. On the average of all 14 animals 5S±13 (S.D.) % of the 24 hr events of locomotor activity occurred during the night. Despite a trough during the cessation of hard faeces excretion the events of food intake were not elevated significantly during the dark hours.

(2) During LL the synchronization of the five functions within each animal persisted during the complete 90-day LL period. A free-running circadian rhythm with-: = 24.8±0.5 (S.D.) hr was present in the four rabbits kept in LL conditions within 5-16 days after the withdrawal of the zeitgeber.

(3) In addition to the circadian period the power spectrum analysis of data obtained during LD 12:12 revealed significant ultradian periods of an average period length of 11,6 hr (hard faeces and urine excretion), 8 hr (food and water intake, locomotor activity) and 4 hr (food intake, locomotor activity). During the free-run ultradian periods of 8 and 3.2-4.2 hr were significant in almost all parameters.

(4) During LL the level of locomotor activity was reduced for 13±16 (S.D.) %, the events of food intake were increased for 17±12 (S.D.) %.

(5) The reinserted LD 12:12 zeitgeber re-entrained the circadian rhythms within 25-45 days.

(6) These results provided evidence of a predominant nocturnality of the rabbits under investigation.     

The cyanobacterium Oscillatoria brevis β-carotene extract modulates alterations of biochemical and hematological circadian patterns in stress-induced rat

Managing stress can prevent serious health problems. The present study aimed to evaluate the possible protective effect of β-carotene (βC), as a natural cyanobacterial product, against stress-induced alterations in biochemical and hematological circadian patterns. Male albino rats were subjected to chronic unpredictable stress (CUS) for 21 days. Rats were randomly divided into four groups (20 rats/group), viz. control, CUS exposed, βC-treated, and βC-treated + CUS-exposed groups. Before CUS exposure, Oscillatoria brevis βC extract was administered (10 mg/kg), intraperitoneally. Blood samples were collected at four Zeitgeber times (ZT: 3, 9, 15 and 21), 5 rats/time point, to monitor circadian profiles of aspartate transferase (AST), alanine transferase (ALT), total protein (TP), glucose (Glu), urea, creatinine, lactate dehydrogenase (LDH), creatinine kinase-MB (CK-MB), hemoglobin (Hb), hematocrite (HCV), red blood cell (RBCs), and white blood cell counts (WBCs). Results revealed that these parameters expressed circadian patterns. CUS exposure significantly (p < 0.05) increased AST, ALT, Glu, urea, creatinine, LDH, CK-MB, and HCV. On the other hand, it significantly decreased Hb, RBCs, and WBCs (p < 0.05). Furthermore, under CUS-exposure the peak time (acrophase) of circadian rhythms of all parameters was variably shifted. On the other hand, βC administration modulated these alternations, where data analysis confirmed a significant decrease in AST, ALT, Glu, urea, creatinine, LDH, CK-MB, and HCV, however, a significant increase in TP, Hb, RBCs, and WBCs (p < 0.05) was observed. Taken together, it can be concluded that βC administration caused restoration of acrophase and level of these parameters thus regulating their circadian rhythms in CUS-induced rats.     

Daily rhythms in blood and milk lead toxicokinetics following intravenous administration of lead acetate to dairy cows in summer

The objective of this study was to investigate circadian variations of blood and milk lead toxicokinetics in dairy cows in summer. Twenty lactating Holstein animals were randomly assigned to four treatments corresponding to different hours after onset of light (HALO): 2, 8, 14, and 20. Cows received a single intravenous administration of 2.5 mg/kg lead as lead acetate. Blood and milk samples were taken and analyzed by atomic absorption spectrophotometry. For each toxicokinetic parameter, a one-way analysis of variance (ANOVA) was performed to outline the existence of daily variations. Significant blood differences as a function of HALO were found for the hybrid constant of distribution (α), hybrid constant of elimination (β), elimination half-life (), area under the curve (AUC), volume of distribution at steady state (V_ss) and clearance (Cl_B) (p<0.05). Half-life of elimination presented two peaks at 2 and 14 HALO. Milk data showed significant differences for maximum concentration and AUC (p<0.05). The ratio AUC_milk/AUC_blood was utilized to estimate penetration of lead in milk. It differed significantly throughout the day (p<0.05). Milk data for the significant parameters could be fitted to circadian rhythms. No circadian rhythms were detected in blood parameters or in the ratio AUC_milk/AUC_blood.     

Circadian rhythms in Neurospora crassa: farnesol or geraniol allow expression of rhythmicity in the otherwise arrhythmic strains frq10, wc-1, and wc-2

In Neurospora crassa, the circadian rhythm can be seen in the bd (band) strain as a series of "bands" or conidiation (spore-forming) regions on the surface of an agar medium. Certain mutations at 3 different genes (frq, wc-1, or wc-2) lead to the loss of the circadian rhythm. In this study, it was found that the addition of 10(-4) to 10(-5) M of geraniol or farnesol restored rhythmic banding to strains that lack a circadian rhythm due to mutations in any 1 of these 3 genes. These 3 conditionally arrhythmic strains now exhibited robust, free-running conidiation rhythms. Their rhythms were neither temperature-compensated nor obviously sensitive to light, so the full properties of a circadian rhythm were not restored. At 20 degrees C, in growth tubes, farnesol treatment gave periods of 28, 26, and 22 h for the frq10, wc-1, and wc-2 strains, respectively. Geraniol treatment at 20 degrees C gave periods of 23, 25.5, and 24.5 h for the frq10, wc-1, and wc-2 strains, respectively. A PRC for temperature pulses (1 h, 20 to 40 degrees C) for the frq10 strain grown in the presence of geraniol showed strong resetting (type 0), suggesting that a temperature-sensitive oscillator was present. Farnesol and geraniol are related to known intermediates in the steroid (or mevalonate) pathway. These data are interpreted in terms of a 2-oscillator model, in which farnesol/geraniol activate or amplify a remaining oscillator (a postulated frq-less oscillator).     

Glomerular filtration rate and blood pressure monitoring in awake baboons

Minimally invasive techniques were used to collect urine with an external catheter together with automated intermittent monitoring of arterial blood pressure in awake male baboons. Using endogenous creatinine, 24-hour creatinine clearances were measured for 2 to 3 consecutive days in four intact and in four uninephrectomized baboons. Despite large differences in urinary volume and sodium excretion, reproducibility of 24-hour creatinine clearances was within 15% in 15 of 19 studies obtained from 6 of 8 animals. Arterial blood pressure was monitored intermittently at 30 to 60 minute intervals over 24 hours with a Dinamap monitor and recorder. Mean blood pressure averaged 71 +/- 4.4 to 89 +/- 5.5 mm Hg in different animals. Blood pressure tended to be lower at night than during the day. In separate studies using 15 to 60 minute urine collection periods, inulin clearance was compared in awake and in anesthetized animals with endogenous or exogenous creatinine clearance measured simultaneously. The clearance of creatinine systematically exceeded the clearance of inulin, even in intact animals with a normal serum creatinine. The creatinine-to-inulin clearance ratio averaged 1.16 +/- 0.03 at a serum concentration of 0.7 to 0.8 mg/dl; 1.27 +/- 0.03 at a serum creatinine of 1.0 to 1.1 mg/dl and 1.56 +/- 0.04 at a serum creatinine greater than 10 mg/dl. All values exceed unity significantly (p less than 0.001). Thus, renal function, including inulin clearance, can be measured in awake baboons. Duplicate or triplicate 24-hour urine collections are needed to assess the reliability of creatinine excretion. However, creatinine clearance overestimates glomerular filtration rate, as it does in humans.     

Circadian rhythms of food and 1% NaCl intake, urine and electrolyte excretion, plasma renin activity and insulin concentration in adrenalectomized rats

The circadian rhythms of food and 1% NaCl intake, and urine, Na+, Cl- and K+ excretion were followed up in male Wistar rats before and one week after bilateral adrenalectomy at 4-hour intervals during two consecutive days. The circadian rhythms of plasma renin activity (PRA) and plasma immunoreactive insulin (IRI) were evaluated after decapitation of both intact and adrenalectomized rats at 08, 16 and 24 h. To all rats 1% NaCl was offered instead of drinking water. Adrenalectomy did not cause any significant phase shift in the cosine curves approximating the data collected at 4-hour intervals. The circadian rhythms showed the same relationships before and after the operation: the rhythms of food intake, K+ excretion and saline intake preceded significantly the rhythms of urine, Na+ and Cl- excretion. Adrenalectomy induced an increase in mean PRA and shifted its minimal value from 08 to 24 h. After the operation mean IRI decreased and the minimal value shifted from 16 to 24 h. It was concluded that adrenal glands do not play an important role in the synchronization of the circadian rhythms of food and 1% NaCl intake, urine and synchronization of the circadian rhythms of food and 1% NaCl intake, urine and electrolyte excretion with the illumination cycle, but play a relevant role in the synchronization of the circadian rhythms of PRA and IRI in the rat.     

Rhythmic Behaviour of W. Bancrofti Microfilaraemia in Human Population at Raipur

The temporal dynamics of microfilariae (mf) in human blood, and the biting activity and mf density in the vector were studied. Finger-prick blood samples (in duplicate) were collected from 14 microfilaraemic subjects at two-hourly intervals over a single 24-hour timescale. Samples were taken from 39 subjects at 22:00 in the first week of every month for a period of 17 consecutive months. The blood slides were dried, stained and examined for presence of microfilariae. Data were analyzed with cosinor rhythmometry. A statistically validated circadian rhythm was detected in mf density in 86% of the microfilaraemic subjects. For the circannual rhythm, the peak timing for microfilariae density was recorded in July, although the spread of the peak between subjects was quite wide, extending between May 17 and September 07. In a separate study, temporal variations in biting activity, mf density in the vector and in human blood were studied. The dynamics of parasite transmission were found to be sequential, with the peaks in the circadian rhythms appearing in the order: mf in human blood, biting activity and mf density in the vector. The biting periodicity appeared to be much more stable in all subjects; the peak appeared at 02:37 with a range between 00:39 and 03:22. The peak in mf frequency was earlier by 3 hours and 25 minutes, whereas the peak in mf frequency in the vector was later by about 2 hours 4 minutes. The observed synchronization between the rhythms of vector and host appears to be essential for effective transmission of diseases. Therefore, the application of chronobiologic principles may have a bearing in the success and failure of any future strategies aimed at the prevention, cure and management of filariasis.     

Ten-Year-Replicated Circadian Profiles for 36 Physiological,Serological and Urinary Variables in Healthy Men

At 3-hr intervals over a 24-hr span, 36 systemic, serologic and urinary variables were examined in 7 men in their mid 20's in the Spring of 1969, and again in the same 7 men in the Spring of 1979 under a similar chronobiologic protocol, using the same chemical and numerical analytical procedures. The variables examined for rhythms by cosinor were: vital signs—blood pressure (systoliC., diastoliC., pulse pressure and mean arterial pressure), heart rate, intraocular pressure (left and right), oral temperature; serum components—albumin, albumin/globulin ratio, total bilirubin, calcium, carbon dioxide, chlorides, bilirubin, cholesterol, globulin, glucose, potassium, sodium, sodium/potassium ratio, transaminase, triglycerides, total protein, urea nitrogen; and urine components—calcium, calcium/magnesium ratio, creatinine, magnesium, pH, potassium, sodium, sodium/potassium ratio, urea clearance, urea nitrogen, volume and zinc. Although all subjects appeared clinically healthy in 1969 and in 1979, certain inter-study differences were observed in a number of rhythm parameters of different variables. Statistically significant increases in mesor for the group as a whole were observed forserum Ca, cholesterol, Cl, CO₂, K, Na, and while statistically significant mesor decreases for a group as a whole were noted in serum glucose and transaminase. Statistically significant increases in amplitude for the group as a whole were observed in serum chloride and urinary Na/K ratio, while statistically signficiant decreases were observed in amplitude for blood pressure, heart rate, serum albumin, A/G ratio, globulin, glucose, protein, sodium and transaminase. For the group as a whole, a statistically significant advance in acrophase was observed in serum transaminase, while a statistically significant delay in acrophase was observed for serum A/G ratio, globulin, glucose, potassium, protein, sodium and for urinary magnesium. Statistically significant by sign test, but not by cosinor, was a numerical mesor increase for urinary urea clearance, a numerical decrease in mesor for urinary zinc; a numerical amplitude decrease for serum cholesterol; and a numerical delay in acrophase for oral temperature and serum cholesterol, CO², and globulin in all men examined. Only mesor changes in serum cholesterol and urinary Ca/Mg were positively correlated with the change in body size over the 10-year span between studies.

From a circadian chronobiologic perspective, the immense amount of data uniquely reviewed in this report across a 10-year span in seven healthy individuals serves a useful beginning to the study of the effects of normal aging upon commonly measured physiologic and biochemical variables and, more importantly, upon the circadian rhythm characteristics of these variables. A great deal of supposition about what happens to the mesor, amplitude and acrophase of an individual's circadian rhythms in a variety of endpoints has been based upon transverse studies of short duration and relatively few longterm studies. The further accumulation of data such as presented here and similar long-term longitudinal time series can have no adequate substitute for truly understanding whether reproducible age-related changes in circadian rhythms occur as individuals age.

With these qualifications and with the further qualification that the timing of our observations within the aging process (mid-20's and mid-30's) may be suboptimal for conclusions about aging, very interesting trends definitely appear worth comment. There is some evidence in these data that the flattening of circadian rhythms may really accompany advancing age. In grouped data, this fall in amplitude may be secondary to an isolated fall in predictable swing around the mesor or a combination of this and increased variability of the acrophase with or without amplitude changes. The data are not robust enough to be sure of the relative contribution of these two components. In any event, the circadian amplitude of each and every physiologic variable studied demonstrated a tendency to fall between the mid-20's and mid- 30's. This tendency toward a flattening of circadian variability is also a very prominent property of many of the serum chemistries which were measured. The circadian patterns of excretion of substances in the urine change much less between the mid-20's and mid-30's in our subjects. These findings may indicate a separate effect of aging especially upon metabolic hepatic variables and upon nephrologic circadian rhythms. Cardiovascular rhythms seem to change more in parallel with hepatic metabolic rhythms in contradistinction to the kidney-related serum and urinary rhythms.

Further, ongoing statistical analyses may hopefully turn up interesting and relevant cross-correlations among the individual data themselves in each study year and between the 10-year span, as well as with rhythm (mesor, amplitude and acrophase) and other physiologic characteristics of each subject. Planned re-observation of what happens to the circadian time structure of these seven individuals in their mid-40's will prove invaluable to further sorting out of the effects of aging upon circadian time structure.