Chronopharmacokinetics of valproic acid following constant-rate administration in mice.

This study was carried out to investigate the circadian rhythm in the pharmacokinetics of valproic acid (VPA). ICR male mice, housed under a light-dark (12 h:12 h) cycle, were used in these studies. In the constant-rate administration study (536.3 or 1,072.6 micrograms/h), osmotic minipumps were implanted subcutaneously in mice. There was a significant circadian rhythm in plasma VPA concentrations: higher values were obtained in the light phase and lower values were found during the dark phase. A significant circadian rhythm also was shown for clearance (CL) of the drug: lower values were obtained in the light phase and higher values were demonstrated in the dark phase. In the intravenous administration study, VPA (50 mg/kg) was injected into a tail vein of the mice. Mean plasma VPA concentrations were significantly higher in mice injected with the drug at 1700 h than at 0100 h. The CL was higher, the volume of distribution (V) was larger, and the area under the curve (AUC) was smaller (p less than 0.05, respectively) in mice injected with the drug at 0100 h than at 1700 h. As the values of CL and V increased similarly during the dark period, there was no effect on half-life (t 1/2) and obviously on the elimination rate constant (K). These findings indicate that the circadian rhythms of plasma VPA concentrations observed after constant-rate administration are due to those of CL and V. To keep drug concentrations constant, the drug release rate from the osmotic minipump should be controlled according to the rhythm of drug pharmacokinetics.     

Chronopharmacokinetics and Chronotoxicity of Lithium in Mice Eating Normal and Low-Sodium Diets

The temporal aspects of the pharmacokinetics and toxicity of lithium were studied in mice eating normal and low-sodium diets. ICR male mice, housed under a lightrdark (LD; 12:12) cycle, were injected with variable doses of lithium chloride i.p. A circadian rhythm was found in lithium clearance after a single administration in mice eating the normal diet showed the maximum value in the early dark phase and the minimum in the early light phase. The repeated administration of lithium did not affect the rhythm of the pharmacokinetics of the drug under the LD cycle. Although the low-sodium diet significantly decreased the lithium clearance, it did not influence the rhythm of the clearance. Higher toxicity was demonstrated in mice injected with the drug at the time of day with lower lithium clearance in the single-dose study but not in the repeated-doses study, regardless of the diet conditions. The low-sodium diet increased the acute and chronic toxicity of lithium. The results indicate that there is a circadian rhythm of acute toxicity and clearance of lithium after a single dose or repeated administration of the drug in mice eating normal and low-sodium diets and that the low-sodium diet increases lithium toxicity by reducing the clearance of the drug without influencing the rhythm characteristics.     

Chronopharmacokinetics and Chronotoxicity of Lithium in Mice Eating Normal and Low-Sodium Diets

The temporal aspects of the pharmacokinetics and toxicity of lithium were studied in mice eating normal and low-sodium diets. ICR male mice, housed under a lightrdark (LD; 12:12) cycle, were injected with variable doses of lithium chloride i.p. A circadian rhythm was found in lithium clearance after a single administration in mice eating the normal diet showed the maximum value in the early dark phase and the minimum in the early light phase. The repeated administration of lithium did not affect the rhythm of the pharmacokinetics of the drug under the LD cycle. Although the low-sodium diet significantly decreased the lithium clearance, it did not influence the rhythm of the clearance. Higher toxicity was demonstrated in mice injected with the drug at the time of day with lower lithium clearance in the single-dose study but not in the repeated-doses study, regardless of the diet conditions. The low-sodium diet increased the acute and chronic toxicity of lithium. The results indicate that there is a circadian rhythm of acute toxicity and clearance of lithium after a single dose or repeated administration of the drug in mice eating normal and low-sodium diets and that the low-sodium diet increases lithium toxicity by reducing the clearance of the drug without influencing the rhythm characteristics.     

Chronotoxicology of florfenicol

Sixty 3-month-old homozygote male mice were studied for circadian rhythmicity in the toxicity of florfenicol overdose. Animals were kept under a regimen of 12h light, 12h darkness (12:12 LD) with food and water available ad libitum. The LD50 (median lethal) dose was determined in a preliminary experiment and was administered to groups of 10 mice at six different clock times (hours) after light onset (HALO): 0, 4, 8, 12, 16, and 20 HALO. Cosinor analysis verified a statistically significant (P < .04) circadian rhythm in the toxic effect (mortality) of florfenicol. Mortality was greatest when the drug was injected 4h after the commencement of the activity span (16 HALO) and least when injected 4h after the start of the diurnal rest span (4 HALO). Mortality was 2.5 times greater when drug injection was given at 16 HALO than at 4 HALO.     

Valproic acid phase shifts the rhythmic expression of Period2::Luciferase

Valproic acid (VPA) is an anticonvulsant used to treat bipolar disorder, a psychiatric disease associated with disturbances in circadian rhythmicity. Little is known about how VPA affects circadian rhythms. The authors cultured tissues containing the master brain pacemaker for circadian rhythmicity, the suprachiasmatic nuclei (SCN), and skin fibroblasts from transgenic PERIOD2::LUCIFERASE (PER2::LUC) mice and studied the effect of VPA on the circadian PER2::LUC rhythm by measuring bioluminescence. VPA (1 mM) significantly phase advanced the PER2::LUC rhythm when applied at a time point corresponding to the lowest (trough, ~ZT 0) PER2::LUC expression but phase delayed the PER2::LUC rhythm when the drug was administered at the time of highest (peak, ~ZT 12) protein expression. In addition, it significantly increased the overall amplitude of PER2::LUC oscillations at time points at or close to ZT 12 but had no effect on period. Real-time PCR analyses on mouse and human fibroblasts revealed that expressions of other clock genes were increased after 2 h treatment with VPA. Because VPA is known to inhibit histone deacetylation, the authors treated cultures with an established histone deacetylation inhibitor, trichostatin A (TSA; 20 ng/mL), to compare the effect of VPA and TSA on molecular rhythmicity. They found that TSA had similar effects on the PER2::LUC rhythm as VPA. Furthermore, VPA and TSA significantly increased acetylation on histone H3 but in comparison little on histone H4. Lithium is another commonly used treatment for bipolar disorder. Therefore, the authors also studied the impact of lithium chloride (LiCl; 10 mM) on the PER2::LUC rhythm. LiCl delayed the phase, but in contrast to VPA and TSA, LiCl lengthened the PER2::LUC period and had no effect on histone acetylation. These results demonstrate that VPA can delay or advance the phase, as well as increase the amplitude, of the PERIOD2::LUCIFERASE rhythm depending on the circadian time of application. Furthermore, the authors show that LiCl delays the phase and lengthens the period of the PER2::LUC rhythm, confirming previous reports on circadian lithium effects. These different molecular effects may underlie differential chronotherapeutic effects of VPA and lithium.     

Circadian time-dependent hepatic and renal toxicities to valproic acid in mice

This study aims to investigate whether hepatic and renal valproic acid (VPA) toxicities varied according to the dosing time in the 24-h scale in mice. VPA was administered by i.p. route to different groups of animals at four different circadian stages (1, 7, 13, and 19 h after light onset (HALO)). Biochemical study and histopathological examinations on liver and kidney sections were performed. The results showed that the hepatic and renal toxicity induced by VPA was time related. Animals treated at 19 HALO showed vacuolar degenerative changes, congestions, and inflammatory areas on liver parenchyma. Lesions within proximal tubules were observed in the kidney in groups treated at 19 HALO. The largest increases in alanine aminotransferase, alkaline phosphatase and plasma creatinine activities were also observed at 19 HALO. The obtained data indicate that the optimal hepatic and renal tolerance is observed when VPA was injected in the middle of the light-rest span of mice.     

Chronobiologic fluctuation of cyclophosphamide induced urinary bladder damage in mice

Cyclophosphamide is the most widely used alkylating agent in clinical medicine. The usefulness of this drug is often limited by its propensity to produce hemorrhagic cystitis. To be active cyclophosphamide must be metabolized by the mixed function oxidase system. It has been previously demonstrated that the oncolytic activity and host lethality of cyclophosphamide are dependent upon circadian fluctuations. When cyclophosphamide is administered i.p. to male mice there is a dose dependent increase in urinary bladder weight. Histopathologic examination of these bladders revealed hemorrhage, edema, inflammation and stretching of the epithelial lining. When administered i.p. at 4-h intervals throughout a 24-h time period, cyclophosphamide produced maximum bladder damage when administered at 0500 and 1700 and little or no damage to the bladder when administered at 0100 or 1300. These studies suggest that cyclophosphamide induced cystitis, a toxicity resulting from the metabolic production of acrolein, may also be dependent upon chronobiologic fluctuations.     

Chronotoxicity of cyclophosphane under different lighting conditions

Study of the chronotoxicity of cyclophosphamide injected to mice at 18, 24, 6 and 12 o'clock has shown that animals kept under the conditions of natural changes of day and night showed the circadian rhythm of the drug toxicity with the maximal survival of the animals after injection at 24 and 6 o'clock and with the minimal survival after injection at 18 o'clock. In animals maintained under the conditions of artificial constant light the toxicity rhythm was perversed within the first hours after injection and improved on subsequent observation. Moreover the earlier death was marked in these animals.     

CHRONOTOXICOLOGY OF FLORFENICOL

Sixty 3-month-old homozygote male mice were studied for circadian rhythmicity in the toxicity of florfenicol overdose. Animals were kept under a regimen of 12h light, 12h darkness (12:12 LD) with food and water available ad libitum. The LD₅₀ (median lethal) dose was determined in a preliminary experiment and was administered to groups of 10 mice at six different clock times (hours) after light onset (HALO): 0, 4, 8, 12, 16, and 20 HALO. Cosinor analysis verified a statistically significant (P <. 04) circadian rhythm in the toxic effect (mortality) of florfenicol. Mortality was greatest when the drug was injected 4h after the commencement of the activity span (16 HALO) and least when injected 4h after the start of the diurnal rest span (4 HALO). Mortality was 2.5 times greater when drug injection was given at 16 HALO than at 4 HALO. (Chronobiology International, 18(3), 567–572, 2001)     

Influence of Feeding Schedule on Chronopharmacological Aspects of Gentamicin in Mice

The effects of the time of day of drug administration on the subchronic toxicity and pharmacokinetics of gentamicin, as well as the role of feeding schedule on circadian rhythms, were investigated in mice. ICR male mice were housed in a light-dark (LD) cycle (12:12) with food and water ad libitum (ALF) or under a time-restricted feeding (TRF) schedule (feeding time: 8 h during the light phase) for 1 day or 14 days before drug administration. The animals were given a single subcutaneous dose of gentamicin 180 mg/kg for the kinetic studies and subcutaneous doses of gentamicin 180 mg/kg/day for 14 days or 220 mg/kg/day for 18 days for the subchronic toxicity studies. A significant dosing-time dependency was shown for mortality and body weight loss, with higher values at midlight and lower ones at the middark (p > 0.05). A significant circadian rhythm was also found for gentamicin kinetics in ALF mice, with the highest clearance at middark and the lowest one at midlight (p > 0.01). The kinetic rhythm of gentamicin coincided well with the toxicity rhythm of the drug. The TRF schedule had a marked influence on the rhythms of gentamicin kinetics and toxicity, showing lowest clearance and higher toxicity at middark. The rhythm of subchronic toxicity of gentamicin seems to be due, at least in part, to the rhythm in kinetics and is strongly influenced by the feeding schedule. Thus, the timing of dosing is an important factor in the kinetics and the subchronic toxicity of gentamicin administration in mice, and the manipulation of feeding schedule can modify the rhythm of the toxicity by changing the rhythm of gentamicin kinetics.     

Time-Dependent Effects of Trichloroethylene on Motor Activity in Rats

Circadian variations in acute and subacute neurobehavioural effects of trichloroethylene (TRI: 1.2 g/kg i.p.) were investigated in the rat under a light: dark = 12:12 hr cycle. An acute effect of TRI evaluated by decreased muscle tone was maximal during the early dark phase (21:00). A subacute effect of TRI was evaluated by a continuous recording of spontaneous locomotor activity in the rat. The circadian rhythm in spontaneous locomotor activity was extensively impaired by the injection of TRI for three consecutive days. Spectral analysis of spontaneous locomotor activity showed that ultradian periods became more dominant than the circadian period, and the 1//fluctuation of the spectrum disappeared after the injection of TRI. The effect of TRI on the circadian rhythm in spontaneous locomotor activity was circadian-phase dependent, and the treatment of TRI at 09:00 provoked greater circadian rhythm impairment than that at 21:00. The mechanisms of the time-dependent effect of TRI on neurobehaviour are the subject of further investigation.     

Time-Dependent Effects of Trichloroethylene on Motor Activity in Rats

Circadian variations in acute and subacute neurobehavioural effects of trichloroethylene (TRI: 1.2 g/kg i.p.) were investigated in the rat under a light: dark = 12:12 hr cycle. An acute effect of TRI evaluated by decreased muscle tone was maximal during the early dark phase (21:00). A subacute effect of TRI was evaluated by a continuous recording of spontaneous locomotor activity in the rat. The circadian rhythm in spontaneous locomotor activity was extensively impaired by the injection of TRI for three consecutive days. Spectral analysis of spontaneous locomotor activity showed that ultradian periods became more dominant than the circadian period, and the 1//fluctuation of the spectrum disappeared after the injection of TRI. The effect of TRI on the circadian rhythm in spontaneous locomotor activity was circadian-phase dependent, and the treatment of TRI at 09:00 provoked greater circadian rhythm impairment than that at 21:00. The mechanisms of the time-dependent effect of TRI on neurobehaviour are the subject of further investigation.     

Effect of Single Low Intensity Light Pulse Exposure and Melatonin Treatment on the Circadian Variation of Melatonin in Indian Palm Squirrel Funambulus pennanti

The endogenous circadian rhythm of melatonin in mammals provides information regarding the resetting response of the mammalian circadian timing system in response to the changes in light dark cycle. Photoperiodic changes are reported to have acute and chronic effect on melatonin rhythm. Our aim in present experiment was to study the effect of single light pulse of low intensity on the circadian variation of melatonin in Indian palm squirrel. A short pulse of 5min was given to the animals at 22:55 h on day 16th in natural photoperiodic condition of long day length (LD ~ 13.55:10.05) and melatonin levels were estimated at every 4-h interval on ZT scale on day 17th (DD). Observations suggest that the light pulse given on day 16th suppressed the melatonin level on day 17th (DD). Besides this, it was also found that there was phase delay in the peak value of melatonin. Further, we tested the ability of single melatonin injection on the light pulse induced phase shift of acrophase of melatonin in this species F. pennanti. We injected the single physiological dose of melatonin (25 microgram/100 g body wt.) just 5 min prior to the commencement of light pulse (22:50 h) on day 16 and melatonin levels were estimated on day 17th as above. Injection of melatonin prior to light pulse altered the suppressing and phase shifting effect of light in terms of peak concentration of melatonin in squirrels. Above data may lead us to conclude that the biological clock mechanism controlling circadian rhythm of melatonin in this rodent is in response to the phase shifting effect of light and acute melatonin treatment. Further, we may suggest that single melatonin injection has the capability to entrain melatonin rhythm but a dose dependent study is required to facilitate the suggestion.     

Effect of Single Low Intensity Light Pulse Exposure and Melatonin Treatment on the Circadian Variation of Melatonin in Indian Palm Squirrel Funambulus pennanti

The endogenous circadian rhythm of melatonin in mammals provides information regarding the resetting response of the mammalian circadian timing system in response to the changes in light dark cycle. Photoperiodic changes are reported to have acute and chronic effect on melatonin rhythm. Our aim in present experiment was to study the effect of single light pulse of low intensity on the circadian variation of melatonin in Indian palm squirrel. A short pulse of 5min was given to the animals at 22:55 h on day 16th in natural photoperiodic condition of long day length (LD ~ 13.55:10.05) and melatonin levels were estimated at every 4-h interval on ZT scale on day 17th (DD). Observations suggest that the light pulse given on day 16th suppressed the melatonin level on day 17th (DD). Besides this, it was also found that there was phase delay in the peak value of melatonin. Further, we tested the ability of single melatonin injection on the light pulse induced phase shift of acrophase of melatonin in this species F. pennanti . We injected the single physiological dose of melatonin (25 microgram/100 g body wt.) just 5 min prior to the commencement of light pulse (22:50 h) on day 16 and melatonin levels were estimated on day 17th as above. Injection of melatonin prior to light pulse altered the suppressing and phase shifting effect of light in terms of peak concentration of melatonin in squirrels. Above data may lead us to conclude that the biological clock mechanism controlling circadian rhythm of melatonin in this rodent is in response to the phase shifting effect of light and acute melatonin treatment. Further, we may suggest that single melatonin injection has the capability to entrain melatonin rhythm but a dose dependent study is required to facilitate the suggestion.     

Influence of Bcg Administration Time on the in-Vivo Migration of Leukocytes

The temporal variation in the migration of polymorphonuclear leukocytes (PMN) induced by live BCG was studied in the mouse. Ten μl of a 5 ± 10⁶ live BCG/ml suspension or sterile saline were placed on a cell trap immediately before its subcutaneous implantation at different clock times: 0100, 0500, 0900, 1300, 1700 and 2100 in animals synchronized with L(0700-1900): D(1900-0700). Eight hours later, the cell trap was removed, prepared for histological identification and counted. PMN counts in the cell trap were maximal 480 min after implantation. Nonspecific migration was thought to occur and the peak value of leukocytes of 22.8 ± 6.1 cells/10,000 μm² was obtained when the saline cell-trap was implanted at 0500. In the BCG-treated mice, a circadian rhythm was observed in the migration of leukocytes. The acrophase was at 1700. The results support the hypothesis that the circadian stage of antigen encounter influences the magnitude of the immune response.     

Influence of Bcg Administration Time on the in-Vivo Migration of Leukocytes

The temporal variation in the migration of polymorphonuclear leukocytes (PMN) induced by live BCG was studied in the mouse. Ten μl of a 5 ± 10⁶ live BCG/ml suspension or sterile saline were placed on a cell trap immediately before its subcutaneous implantation at different clock times: 0100, 0500, 0900, 1300, 1700 and 2100 in animals synchronized with L(0700-1900): D(1900-0700). Eight hours later, the cell trap was removed, prepared for histological identification and counted. PMN counts in the cell trap were maximal 480 min after implantation. Nonspecific migration was thought to occur and the peak value of leukocytes of 22.8 ± 6.1 cells/10,000 μm² was obtained when the saline cell-trap was implanted at 0500. In the BCG-treated mice, a circadian rhythm was observed in the migration of leukocytes. The acrophase was at 1700. The results support the hypothesis that the circadian stage of antigen encounter influences the magnitude of the immune response.     

Behavioral and Neurochemical Effects of Fencamfamine on Rats: A Chronobiologic Approach

Fencamfamine (FCF) is a psychostimulant classified as an indirect dopaminergic agonist. Circadian rhythms of some behavioral and neurochemical parameters were investigated in control rats and in rats which had been treated with a single dose of FCF across the 24-hr span. Rats were entrained to light/dark (LD) 12:12, lights on from 0700 to 1900. In behavioral experiments (performed in March) the rats were injected intraperitoneally with saline or FCF (3.5 mg/kg) at one of six times: 0900, 1300, 1700, 2100, 0100 or 0500. Fifteen minutes after treatment the duration of sniffing, rearing and locomotion was recorded during 120 min. Controls showed circadian rhythms for sniffing and rearing with acrophases at 2255 and 0118, respectively. In animals treated with FCF, only locomotion displayed significant circadian variation with acrophase at 1912. Two-way analysis of variance (ANOVA) showed a statistically significant circadian time-dependent effect of FCF on all behavioral parameters studied; the increase of sniffing, rearing and locomotion induced by FCF was higher in rats treated during the rest phase. In the biochemical studies (performed between March-June), rats were treated (i.p.) with saline or FCF (10 mg/kg) at one of four times: 0900, 1700, 2100 or 0100. The levels of homovanillic acid (HVA) in the striatum and tuberculum olfactorium, 5-hydroxyindolacetic acid (5-HIAA) in the cerebellum and 3-methoxy-4-hydroxypheniglycol (MHPG) in the frontal cortex were determined. Controls showed circadian rhythms for HVA (striatum), MHPG (frontal cortex) and 5-HIAA (cerebellum) with acrophases at 2233, 1955 and 1029, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)     

The effects of the light-dark cycle and its inversion on the susceptibility of mice to lead acetate

The effects of the light-dark (LD) cycle and its inversion on the susceptibility of mice to lead were investigated. Groups of mice were injected once with a high (200 mg Pb/kg body weight) or low (10 mg Pb/kg body weight) dose of lead acetate at one of four-hourly intervals, i.e., 09.00 h, 13.00 h, 17.00 h, 21.00 h, 01.00h, and 05.00 h, on the days before and after a phase shift in the LD cycle. The main indices used to measure the susceptibility of the mice were dying rate and percent mortality of ICR mice following injection together with the Pb concentration (Pb-B) and -aminolevulinic acid dehydratase (ALAD) activity in the blood. In the present study, we ascertained the following facts: (1) Dying rate (time to death following injection) and Pb-B exhibited a circadian rhythm with a peak initiated by injection in the dark period. (2) Injection with a low dose of lead within a few days after an LD inversion significantly decreased the level of ALAD activity and increased the Pb in the blood in comparison with control animals kept under a non-inverted LD cycle. Both the dying rate and the percent mortality following a high dose of lead, however, were hardly affected.     

Circadian modification of ethanol damage in utero to mice

This study evaluates the ethanol toxicity for fetal development at different circadian stages. Pregnant mice were given a single intraperitoneal ethanol injection on day 7, 8, or 10 of gestation at one of four circadian stages (0700, 1300, 1900, or 0100 hr). The dams were killed on the day before term (day 18). Prenatal exposure to ethanol resulted in an increased number of resorptions, reduced fetal body weight, and produced an increased incidence of external alterations. The severity of damage was related to the dose, the period of gestation, and particularly to the circadian stage at the time of treatment. Ethanol had the greatest effect on the embryo of a mouse when administered at the mid-dark span. Consequently exposure to a single dose of ethanol at one time or another along the 24-hr time scale during organogenesis has important implications for the substantially increased risk.     

Anticonvulsant profile and teratogenic evaluation of potent new analogues of a valproic acid urea derivative in NMRI mice

BACKGROUND: Valproic acid (VPA) is used to treat epilepsy and bipolar disorders, as well as for migraine prophylaxis. However, its clinical use is limited by two life-threatening side effects: hepatotoxicity and teratogenicity. To develop a more potent and safer second-generation VPA drug, the urea derivatives of four VPA analogs (2-ethyl-3-methylpentanoyl urea, 2-ethylhexanoyl urea, 2-ethyl-4-methylpentanoyl urea, and 2-methylbutanoyl urea) were synthesized. METHODS: Four CNS-active analogs of a VPA urea derivative testedthe anticonvulsant activity in the maximal electroshock seizure test (MES) and subcutaneous metrazol seizure threshold test (scMet). Teratogenic effects of these compounds were evaluated in NMRI mice susceptible to VPA-induced teratogenicity by comparison with VPA. RESULTS: All four VPA analogs showed superior anticonvulsant activity over VPA. Compared with VPA, which induced neural tube defects (NTDs) in fetuses at 1.8 and 3.6 mmol/kg, the analog derivatives induced no NTDs at any concentration up to 4.8 mmol/kg (except for a single abnormality at 3.6 mmol/kg with 2-ethyl-3-methylpentanoyl urea). Skeletal examination also revealed that the acylurea derivatives induced vertebral and rib abnormalities in fetuses markedly less frequently than VPA. Our results confirmed that the analogue derivatives are significantly less teratogenic than VPA in NMRI mice. CONCLUSIONS: The CNS-active VPA analogs containing a urea moiety, which have better anticonvulsant potency and lack teratogenicity, are good potential candidates as second-generation VPA antiepileptic drugs. Birth Defects Res (Part B) 86:394–401, 2009. © 2009 Wiley-Liss, Inc.